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    Research Progress in Genomic Selection Breeding Technology for Crops
    WANG Xin, XU Yi-yi, XU Yang, XU Chen-wu
    Biotechnology Bulletin    2024, 40 (3): 1-13.   DOI: 10.13560/j.cnki.biotech.bull.1985.2023-1079
    Abstract2980)   HTML105)    PDF(pc) (1596KB)(1308)       Save

    Genome selection(GS)breeding builds a genetic model based on the association between genotypes of molecular markers on the whole genome and phenotypes of the training population, and then estimates the breeding values or predicts the phenotypes of the candidate population with known genotypes, so as to achieve efficient and accurate selection of the population for breeding. Compared with the commonly used molecular marker-assisted selection breeding, GS breeding does not require marker significance testing, and is particularly suitable for quantitative traits controlled by minor polygenes. It can shorten breeding cycle and reduce breeding cost, and has become a cutting-edge technology in the field of animal and plant breeding. However, for quantitative traits such as crop yield that are greatly affected by environment, it is still bottleneck issue to improve the accuracy of genomic prediction. This article first analyzes the main factors that affect the efficacy of GS in crop breeding, and then elaborates on the research progress of GS technology in crop breeding from the aspects of models with non-additive effects, population construction schemes, multi-trait and multi-environment prediction, multi-omic prediction and the current status of breeding chip technology. Then the article points out the issues and development prospects of the research, and provides the strategies and ideas for further research on crop GS breeding technology.

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    Paenibacillus polymyxa New Strain X-11 and Its Growth-promoting Effects on Tomato and Rice
    LIU Wen-zhi, HE Dan, LI Peng, FU Ying-lin, ZHANG Yi-xin, WEN Hua-jie, YU Wen-qing
    Biotechnology Bulletin    2024, 40 (9): 249-259.   DOI: 10.13560/j.cnki.biotech.bull.1985.2024-0253
    Abstract2598)   HTML5)    PDF(pc) (4822KB)(194)       Save

    【Objective】A plant-growth-promoting rhizobacterium strain(named X-11)was screened from rhizosphere soil of wheat(Triticum aestivum L). Which may provide a basis for the application of the strain in plant growth-promoting.【Method】Strain X-11 was identified by morphological characteristics, physiological and biochemical characteristics, 16S rDNA sequence and genome sequencing, and its effects on the growth and defense enzyme activities of tomato(Solanum lycopersicum)and rice(Oryza sativa)were measured. 【Result】The colonies of strain X-11 are round, swell, neat edge, translucent, and sticky. And the cells are rod-shaped, can form spores, gram negative reaction. And it has the functions of fixing nitrogen, reducing nitrate, producing indole and siderophore, degrading organic phosphorus. Strain X-11 was classified as Paenibacillus polymyxa group based on 16S rDNA sequence, while it was revealed to be a new strain with 55.7% genetic similarity to its closest relative strain through gene comparison in non-redundant database. Tomato(Solanum lycopersicum)seeds soaked with X-11 bacterial suspension increased germination rate and radicle length significantly. Moreover, peroxidase(POD)activity in the leaves significantly increased by X-11 bacterial suspension irrigating roots(R)and irrigating roots combined with spraying leaf surface(R+L). Meanwhile, the R+L treatment increased superoxide dismutase(SOD)activity by 34.58%. X-11 bacterial suspension increased the seedling root length by 6.8% when applied at one-leaf phase of rice(Oryza sativa). Additionally, the SOD activity of the seedlings increased by 90.4% and 51.8%, when the X-11 bacterial suspension was applied at vertical phase or one-leaf phase of rice. 【Conclusion】 Strain X-11 was identified as a new strain of P. polymyxa, which is the effective plant-growth promoting rhizobacterium with multiple functions and has potential for development and application in plant growth promotion.

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    Roles of MYB Transcription Factor in Regulating the Responses of Plants to Stress
    HU Ya-dan, WU Guo-qiang, LIU Chen, WEI Ming
    Biotechnology Bulletin    2024, 40 (6): 5-22.   DOI: 10.13560/j.cnki.biotech.bull.1985.2023-1186
    Abstract2518)   HTML66)    PDF(pc) (2607KB)(1744)       Save

    MYB is one of the biggest multifunctional transcription factors(TFs)families in plants. It is widely involved in regulating multiple process at the transcription level, including plant growth and development, signal transduction of various phyhormones and response to abiotic and biotic stresses. The N-terminal of MYB contained a typical MYB domain, which is divided into different subgroups according to the number of R repeats in the domain. However, the C-terminal domain is very different, thus it is functionally diverse. Many studies have been shown that MYB can bind to the downstream target gene promoter cis-acting elements MYBCORE and AC-box alone or through interaction with other proteins after being activated by environmental signals, and participate in regulating the expressions of downstream target genes, thereby regulating plant tolerances to stresses. Additionally, MYB responded to abiotic and biotic stresses by regulating signaling pathways such as abscisic acid(ABA), brassinolide(BR), jasmonic acid(JA)and reactive oxygen species(ROS). In this review, the structure, classification, and action type of the MYB families in plants were summarized, and the regulatory mechanisms of MYB in the response to abiotic and biotic stresses such as salt, drought, extreme temperature, nutrient deficiency, heavy metals, and pathogenic bacteria, were mainly reviewed. The key direction of future studies were also prospected. This paper may provide genetic resources and theoretical support for genetic improvement of stress resistance of crops and biological breeding in the future.

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    Application of Gene-editing Technology for Germplasm Innovation and Genetic Improvement in Cotton
    HOU Wen-ting, SUN Lin, ZHANG Yan-jun, DONG He-zhong
    Biotechnology Bulletin    2024, 40 (7): 68-77.   DOI: 10.13560/j.cnki.biotech.bull.1985.2023-1054
    Abstract2514)   HTML12)    PDF(pc) (1283KB)(389)       Save

    Cotton is a globally important cash crop and a crucial raw material for the textile industry. Breeding superior cultivars is the primary approach for increasing cotton yield, fiber quality and agronomic benefits. While traditional breeding methods are increasingly limited in improving the genetic traits of crops, gene editing technology offers opportunities to promote germplasm innovation and genetic improvement in cotton. Gene editing is a technology that utilizes engineered nucleases to precisely edit the DNA sequences of an organism's genome by deleting, modifying, inserting or replacing individual or multiple nucleotides in specific target genes. This article reviews the principles of three major gene editing systems, namely ZFNs, TALENs, and CRISPR, aiming to better understand how gene editing technology can be used to enhance cotton growth, development, and stress tolerance. Specifically, a comprehensive overview of the CRISPR gene editing system, which has gained significant attention, is provided, summarizing its current applications in improving cotton stress tolerance and other desirable traits. In addition, this article analyzes the shortcomings and limitations of gene editing technology, emphasizes the need for further research of optimizing and developing gene editing system with intellectual property right as well as increasing its accuracy and safety, and thus leverage its application in germplasm innovation and genetic improvement in cotton.

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    Responses of Sorghum Rhizosphere Soil Bacterial Communities to Salt Stress
    GAO Yu-kun, ZHANG Jian-dong, YANG Pu-yuan, CHEN Dong-ming, WANG Zhi-bo, TIAN Yi-jin, Zakey Eldinn. E. A. Khlid, CUI Jiang-hui, CHANG Jin-hua
    Biotechnology Bulletin    2024, 40 (4): 203-216.   DOI: 10.13560/j.cnki.biotech.bull.1985.2023-0685
    Abstract2440)   HTML13)    PDF(pc) (8197KB)(150)       Save

    Objective】This study aims to investigate the changes in the bacterial community structure of sorghum roots under salt stress, as well as the characteristics of the rhizosphere bacterial co-occurrence network structure.【Method】Two sorghum varieties, HN16(salt-sensitive)and GLZ(salt-tolerant), with varying levels of salt tolerance were cultivated in pots and subjected to salt stress. The root microbiome of these plants was then analyzed using 16S amplicon sequencing technology.【Result】With the aggravation of salt stress, the contents of total phenols and flavonoids in the sorghum roots gradually increased, and the appropriate salt stress induced the synthesis of phenolic acids to improve the salt tolerance of sorghum. The results of the microbial community structure analysis showed that Proteobacteria, Actinobacteria, Acidobacteria, Bacteroidetes and Chloroflexi were the dominant phyla in sorghum rhizosphere soils. Comparative analysis of samples showed that the bacterial composition of sorghum rhizosphere under salt stress was less affected by growth stage, and the bacterial community structure of sorghum rhizosphere changed with the aggravation of salt stress. Through WGCNA analysis of all filters OTUs, a total of 12 gene co-expression module were identified, pink module had significantly positive correlation with salinity and greenyellow module had significantly positive correlation with period and the contents of total phenols and flavonoids after salt treatment. The bacterial co-occurrence network was more complex in low salt stress soils than in higher salt stress soils, reflected by more nodes and more edges. The 13 network key OTUs were identified, OTU8480, OTU6866, OTU3247 and OTU3499 was the keyOTUs in S0 network; OTU6895, OTU4206, OTU6470, OTU1810 and OTU4916 in S3 network; OTU4217, OTU8426, OTU4847, and OTU6066 were the key OTUs in S7 network. 【Conclusion】The composition of rhizosphere bacteria in both varieties of sorghum has been modified in response to salt stress. After salt stress, with more complex co-occurrence network and tighter connections between OTUs of rhizosphere soil bacterial communities.

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    Editing Features of High-fidelity CRISPR/Cas9 System in Poultry Cells
    JIAO Dan-rong, MA Meng-xue, HE Bai-shui, XIE Long, ZUO Er-wei
    Biotechnology Bulletin    2024, 40 (10): 191-197.   DOI: 10.13560/j.cnki.biotech.bull.1985.2024-0673
    Abstract2304)   HTML14)    PDF(pc) (4033KB)(48)       Save

    【Objective】 The emergence of high-fidelity editors in recent years is expected to widely reduce the off-target effect that occurs during gene editing in poultry with the conventional CRISPR/Cas9 system. However, there is still a lack of data support for the application of these high-fidelity editors in poultry. This study aims to further evaluate their on-target editing effects and off-target characteristics in poultry cells, so as to provide an effective reference for molecular genetic breeding and germplasm resource utilization in poultry.【Method】 Five high-fidelity Cas9 variants, which have been reported with high accuracy, were selected and co-transfected with the designed sgRNAs in chicken DF-1 cells, and the positive cells were collected for gene sequencing, and the editing efficiency and off-target effect of different high-fidelity editors were compared in parallel.【Result】 eCas9 maintained high editing activity while demonstrating low off-target effects in multiple sgRNAs tested against the avian influenza resistance-related gene ANP32B and the reproduction-related gene DAZL. Meanwhile, SuperFiCas9 had the lowest off-target effect among multiple tested high-fidelity variants, but a significant decrease in editing efficiency occurred in gene editing of poultry DF-1 cells.【Conclusion】 A variety of high-fidelity editors have shown editing activity in poultry cells, among which the eCas9 editor has high efficiency and low off-target properties, and can be used as a preferred high-fidelity editing tool for the genetic improvement of economically important traits and molecular breeding work in chickens.

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    Alternative Translation of Wheat Enolase-encoding Gene ENO2 and Its Prokaryotic Expression
    ZHANG Na, LIU Meng-nan, QU Zhan-fan, CUI Yi-ping, NI Jia-yao, WANG Hua-zhong
    Biotechnology Bulletin    2024, 40 (5): 112-119.   DOI: 10.13560/j.cnki.biotech.bull.1985.2023-1223
    Abstract2102)   HTML9)    PDF(pc) (5168KB)(122)       Save

    Objective】The objective of this study is to identify the genes(TaENO2s)encoding the enolase family member ENO2 in the important crop plant of wheat(Triticum aestivum L.)and to determine the alternative translation feature and protein enolase activity of TaENO2s.【Method】Wheat TaENO2s were identified by bioinformatics. One of the identified TaENO2s was selected as a representative for protoplast-based exogenous expression and characterization of alternative translation products. This selected TaENO2 was also subjected to prokaryotic expression for purification of recombinant protein. The enolase activity of the purified recombinant protein was determined with in vitro assays. 【Result】The hexaploid wheat genome had three homeologous TaENO2s which sequences encoded a conserved enolase catalytic center and had a putative alternative translation start site at the internal ATG codon encoding the residue M94(ATGM94)of the enolase sequence. When exogenously expressed in protoplasts, the representative TaENO2 encoded two proteins, one full-length form of cytoplasmic and nuclear enolase protein and one N-terminal truncated form of nuclear transcriptional repressor TaMBP-1, while the variant of the same gene containing mutated ATGM94 only encoded the full-length form of enolase protein. Soluble recombinant GST-TaENO2 protein expressed in Escherichia coli was successfully purified and verified to have an in vitro enzymatic activity to catalyze the conversion from 2-phosphoglycerate(2-PGA)to phosphoenolpyruvate(PEP). 【Conclusion】The wheat genome has three TaENO2 genes encoding the enolase protein ENO2. Under the condition of exogenous expression, TaENO2 encodes two protein products via mRNA alternative translation, one enolase protein translated from the first start codon and one TaMBP-1 protein translated from the internal start codon ATGM94. The prokaryotically expressed recombinant protein GST-TaENO2 possesses in vitro enolase activity.

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    The Growth Promoting Effect of Bacillus amyloliquefaciens SQ-2 on Rice
    LI Xue, LI Rong-ou, KONG Mei-yi, HUANG Lei
    Biotechnology Bulletin    2024, 40 (2): 109-119.   DOI: 10.13560/j.cnki.biotech.bull.1985.2023-0770
    Abstract2081)   HTML13)    PDF(pc) (5138KB)(264)       Save

    【Objective】 To study the growth-promoting characteristics of a strain of Bacillus amyloliquefaciens SQ-2 isolated and screened in the laboratory, and to determine the concentration range, mechanism of action, and changes in soil microbiota induced by this strain to promote rice growth.【Method】 The phosphorus solubility and nitrogenase activity of strain SQ-2 were detected using the molybdenum blue colorimetric method and ELISA nitrogenase kit. Inoculating102, 104, 106, 108, and 3×108 CFU/mL bacterial suspension into hydroponic and soil cultured rice, the dry and fresh weight, length, and thickness of the roots and stems of hydroponic and soil cultured rice were measured, respectively after 14 and 20 d of cultivation. Meanwhile, the activities of urease, protease, and sucrase in soil cultured rice were determined using the phenol sodium hypochlorite colorimetric method, the ninhydrin detection method, and the 3,5-dinitrosalicylic acid colorimetric method. Besides, the soil pH value was determined using the pH meter potential method and the content of nitrogen, phosphorus, and potassium available in soil and the activity of acid phosphatase were detected by utilizing the corresponding reagent kit. Analysis of rhizosphere bacterial community structure in soil cultured rice soil was inoculated with 3×108 CFU/mL components.【Result】 The dissolution of tricalcium phosphate by strain SQ-2 was 229.63 mg/L, and the nitrogenase activity was 55.07 U/L. Compared with the control, the dry and fresh weight of hydroponic rice roots showed the most significant increase at an inoculum suspension concentration of 104 CFU/mL. At a vaccination concentration of 3×108 CFU/mL, the dry and fresh weight of soil cultivated rice rhizomes increased most significantly. As the concentration of the inoculum suspension increased, the activities of various enzymes and the concentration of available nitrogen, phosphorus, and potassium in the soil increased to varying degrees, the soil pH value while inoculating strain 3×108 CFU/mL decreased from 7.83 to 7.26. The inoculant strain SQ-2 changed the colony composition in the rice rhizosphere soil and significantly increased Ace, Chao, Sobs and Shannon indices of soil α-diversity.【Conclusion】 B. amyloliquefaciens SQ-2 had different growth promotion effects on soil culture and hydroculture rice. In soil culture experiments, the growth-promoting effect is notably pronounced through the augmentation of soil enzyme activity, rapid increases in available nitrogen, phosphorus, and potassium levels, and alterations in the soil microbial community structure. Overall, these experimental findings offer valuable insights into new strain resources for the development of bacterial fertilizer.

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    Effects of Arbuscular Mycorrhizal Fungi on Plant Stress Resistance and Secondary Metabolite Synthesis
    YUAN Hai-peng YE Yun-shu SI Hao JI Qiu-yan ZHANG Yu-hong
    Biotechnology Bulletin    2024, 40 (6): 45-56.   DOI: 10.13560/j.cnki.biotech.bull.1985.2023-1117
    Abstract2033)   HTML23)    PDF(pc) (3102KB)(771)       Save

    Arbuscular mycorrhizal(AM)is a symbiotic system composed of the fungi of the phylum Glomeromycota and plant roots. Seventy-one percent of vascular plants have AM. Glomeromycota is now under a single class, four orders, eleven families and twenty-seven genera, with about 300 species of arbuscular mycorrhizal fungi(AMF). AMF improves plant resistance to abiotic stresses such as drought, salinity, heavy metals, and biotic stresses induced by other organisms as well as regulates the synthesis of plant secondary metabolites.This article reviews the research progress on the effects of AMF on plant adversity stress and secondary metabolites. The effects of AMF on life activities such as plant morphology and structure, physiology and biochemistry, gene expression and secondary metabolism are described. The mechanism of action of AMF in enhancing plant resistance to adversity stress and regulating the synthesis of plant secondary metabolites is summarized. It aims to provide a reference for the in-depth study of the action mechanism of AMF in response to plant adversity stress and regulation of plant secondary metabolite synthesis.

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    High-throughput Screening System for Functional R-loop Loci Based on R-loop Targeted Editing Technology
    PI Yi-fei, SONG Xin-hui, WANG Xi-lin, LI Jin-jin, SUN Chang-bin, XU Wei
    Biotechnology Bulletin    2024, 40 (10): 181-190.   DOI: 10.13560/j.cnki.biotech.bull.1985.2024-0613
    Abstract2016)   HTML17)    PDF(pc) (4638KB)(53)       Save

    【Objective】 To develope R-loop targeted editing technology for mammalian cells and explore its applications in drug resistance of tumor cells. 【Method】 An expression vector was constructed to express a dCas9-RNaseH1 chimaera that combines catalytically dead Cas9 lacking endonuclease activity with RNase H1 possessing R-loop hydrolysis activity for R-loop targeted editing. This dCas9-RNaseH1 chimaera was transfected to HeLa cells and stably expressed to construct a model cell line. To create a cell library for R-loop screening, a genome-wide gRNA library covering transcription start sites was transfected to the model cell line, through which R-loop functional sites affecting resistance to paclitaxel and cisplatin were identified. 【Result】 Total 744 R-loop functional sites affecting HeLa cell drug resistance were identified, which cover key biological pathways such as cell cycle, apoptosis, and signal transduction. Among them, 26 sites confered resistance to two drugs, while 8 sites rendered sensitivity to both drugs, suggesting potential shared biological pathways. Functional validation revealed that certain R-loop sites modulated the expressions of relevant genes(e.g., ZBTB20, SPON2, ACTRT1), significantly impacting HeLa cell sensitivity to anticancer drugs. 【Conclusion】 A R-loop targeted editing system is successfully developed and a high-throughput screening platform is established for mammalian cells.

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    Advancing Biological Breeding for Tuber Crops to Ensure National Food Security
    XU Jian-fei, SHANG Yi
    Biotechnology Bulletin    2024, 40 (9): 1-3.  
    Abstract1953)   HTML232)    PDF(pc) (1116KB)(216)       Save
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    Effects of Endophytic Fungal Inoculation on the Seedling Growth of Silage Maize
    WANG Jia-wei, LI Chen, LIU Jian-li, ZHOU Shi-jie, YI Jia-min, YANG Jin-yuan, KANG Peng
    Biotechnology Bulletin    2024, 40 (4): 189-202.   DOI: 10.13560/j.cnki.biotech.bull.1985.2023-1081
    Abstract1844)   HTML10)    PDF(pc) (5170KB)(185)       Save

    Objective】Endophytic fungi represent significant microbial assets that exert a beneficial influence on plant growth. The method of inoculation plays a pivotal role in determining the growth-promoting efficacy of endophytic fungi.This study aims to investigate the impact of various inoculation methods on the growth-promoting potential of endophytic fungi, thereby offering valuable insights for the practical utilization of endophytic fungi. 【Method】Four strains of endophytic fungi and Piriformospora indica of different species that could promote the growth of silage maize were screened from the endophytic fungi of desert plant roots in the early stage of the research group. Five strains of endophytic fungi were inoculated into the potted seedlings of silage maize by means of root irrigation with mycelial fragment suspension and root wrapping with mycelial mass. After 30 d, the biological characteristics of the seedlings inoculated with silage maize were determined, and the effects of different inoculation methods on the growth of silage maize were compared.【Result】The inoculation method had no significant effect on the mycelial infection rate and basal stem diameter of the five endophytic fungi in the roots of silage maize seedlings, but the other 17 traits related to the growth of silage maize were significantly affected by the inoculation method. The results of principal component analysis showed that the growth-promoting effect of the five endophytic fungi using mycelial fragment suspension root irrigation was significantly better than that of mycelial pellet root, and the difference between the two inoculation methods of P. indica and strain Tm36 was the largest. Ten traits of silage maize seedlings inoculated with P. indica and strain Tm36 were significantly different between the two inoculation methods, they were plant height, aboveground fresh weight, aboveground dry weight, average leaf area, leaf relative water content, root fresh weight, root surface area, root volume, whole plant fresh weight and whole plant dry weight. Among them, the five indexes of plant height, aboveground fresh weight, average leaf area, root fresh weight and whole plant fresh weight were the most different. The growth rate of P. indica inoculated with mycelial fragment suspension was 7.78, 3.74, 15.97, 7.93 and 5.36 times that of mycelial mass inoculation, and was 8.65 times, 4.33 times, 11.18 times, 16.58 times and 7.53 times that of inoculation strain Tm36.【Conclusion】The inoculation method significantly affects the growth-promoting effect of endophytic fungi. The inoculation method of mycelial fragment suspension root irrigation has better effect on the growth of silage maize seedlings than that of mycelial pellet root inoculation. It is recommended to use mycelial fragment suspension root irrigation method.

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    Construction and Transcriptomic Analysis of Rice Histone H1 Triple Mutant
    YANG Qi, WEI Zi-di, SONG Juan, TONG Kun, YANG Liu, WANG Jia-han, LIU Hai-yan, LUAN Wei-jiang, MA Xuan
    Biotechnology Bulletin    2024, 40 (4): 85-96.   DOI: 10.13560/j.cnki.biotech.bull.1985.2023-1066
    Abstract1830)   HTML35)    PDF(pc) (6835KB)(225)       Save

    Objective】Histone H1 plays an important role in the maintenance and stabilization of higher chromatin structure. Elucidating the effect of rice histone H1 on gene expressions will help to better understand the regulatory functions of H1 in rice.【Method】Semi-quantitative RT-PCR and real-time quantitative PCR were used to detect the expressions of four H1 genes in rice. CRISPR technology was used to generate h1 mutant plants; subsequently, phenotypic and transcriptomic analysis of the h1 mutant was performed. 【Result】The four rice H1 genes were broadly expressed, and are weakly expressed in root. In T0 generation of CRISPR-based gene editing plants, multiple mutations in H1.1-H1.4 were identified. In T1 generation, one Osh1.1 Osh1.3 Osh1.4 homozygous triple mutant was identified, which had various developmental defects and became the material for further transcriptomic sequencing analysis. In T2 generation, quadruple and triple mutants were acquired, approximate 25% of which were albino seedlings, slow in plant development and defect in drought responses. By transcriptome sequencing of the h1 mutant, 1 055 differentially expressed genes were identified, of which the significantly up-regulated genes were approximately 2.5 times of the down-regulated genes, implying that H1 may inhibit gene expressions at genome-wide scale. 【Conclusion】In the mutant, pathways of photosynthesis, stress responses, amino acid and RNA metabolism were disrupted. The ribosome biogenesis and photosynthesis pathway genes were significantly up-regulated; while the stress-associated dehydrogenase genes were down-regulated. It is presumed that overexpression of ribosome pathway genes resulted in the disorder of protein homostasis, which ultimately caused plant developmental defects.

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    Advances in Methylation and Acetylation Modification of RNA in Plant Growth and Development
    ZHANG Yan-yan, LU Du-xian, ZUO Xin-xiu, LI Yan-jun, LIN Jin-xing, CUI Ya-ning
    Biotechnology Bulletin    2024, 40 (10): 108-121.   DOI: 10.13560/j.cnki.biotech.bull.1985.2024-0327
    Abstract1754)   HTML17)    PDF(pc) (2602KB)(113)       Save

    RNA modification is an important type of epigenetic modification, which refers to the addition of chemical modifying groups to the bases or ribose of RNA molecules, and can occur in a variety of RNAs, such as messenger RNAs(mRNAs), transfer RNAs(tRNAs), ribosomal RNAs(rRNAs), and cyclic RNAs(circRNAs), etc. RNA methylation and acetylation are two of the key types of modifications that perform important biological functions by regulating the genetic information of organisms. Both modifications are dynamically and reversibly regulated by three types of regulators, namely writers, erasers and readers, which have a crucial impact on different RNA metabolic processes such as RNA splicing, translocation, translation, transport and degradation. In recent years, with the development of RNA modification detection technologies, numerous studies have identified new RNA modification sites in plants and confirmed that methylation and acetylation modifications of RNAs play key roles in plant growth and development as well as in response to biotic and abiotic stresses. This paper firstly outlines the types of RNA modifications and their biological properties, and on this basis focuses on reviewing the research progress on RNA methylation and acetylation regulators in recent years, and finally summarizes the functions of RNA methylation and acetylation in plant growth and development and in response to stresses, and looks forward to some new research directions for RNA modifications in plants, with a view to providing theoretical basis and new ideas for further research related to RNA methylation and acetylation modification in plants.

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    Effects of Compound Microbial Agent on the Growth, Quality and Rhizosphere Environment of Grape
    CHE Jian-mei, LAI Gong-ti, LI Si-yu, GUO Ao-lin, CHEN Bing-xing, CHEN Xing, LIU Bo, LAI Cheng-chun
    Biotechnology Bulletin    2024, 40 (8): 264-274.   DOI: 10.13560/j.cnki.biotech.bull.1985.2024-0230
    Abstract1706)   HTML13)    PDF(pc) (3921KB)(404)       Save

    【Objective】Grape is one of the five varieties of fruit trees widely cultivated in China. Long-term excessive application of chemical fertilizer will destroy the microbial ecology, thus affecting the yield and quality of grapes. Compound microbial agent of different strains may be a more active and effective measure to improve the quality of grapes.【Method】The compound microbial agent(Brevibacillus brevis FJAT-0809-GLX, Bacillus velezensis FJAT-55034 and B. brevis FJAT-10623)were used to irrigate the roots, and the effects of the compound microbial agent on grape growth, fruit quality, rhizosphere soil enzymatic activity and diversity of culturable Bacillus were analyzed.【Result】The compound microbial agent promoted the growth of grape leaves, increased the contents of chlorophyll a and carotenoid, and the activities of POD and PPO. The chlorophyll b content, SOD and PPO activities of young grape fruits in the treatment group was 100%, 1.77% and 66.67% higher than that in the control group, respectively. The application of compound microbial agent promoted early ripening, fruit color transformation and quality of grapes. The single fruit weight was 29.06% higher than that of control group, and the soluble solid and soluble protein increased by 7.31% and 94.74%, respectively, compared with the control group. The PPO enzyme activity of ripe grape fruit in the treatment group was significantly higher than that in control group. In addition, the compound microbial agent significantly improved the activities of amylase, catalase, cellulase, sucrase, and the number and species of Bacillus in grape rhizosphere soil. Correlation analysis showed that Bacillus zanthoxyli and Bacillus pseudomycoides promoted grape leaf growth and fruit quality.【Conclusion】The compound microbial agent may improve soil, promote grape growth and increase fruit quality.

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    Screening and Growth-promoting Characteristics of Multifunctional Exopolysaccharides-producing Bacteria
    CHANG Hai-xia, LI Ming-yuan, MAIRIYANGU·Yasheng , ZHOU Qian, WANG Ji-lian
    Biotechnology Bulletin    2024, 40 (3): 273-285.   DOI: 10.13560/j.cnki.biotech.bull.1985.2023-0942
    Abstract1704)   HTML14)    PDF(pc) (4414KB)(255)       Save

    【Objective】 To explore the effect of exopolysaccharides-producing strains in promoting plant growth and improving soil properties in saline soils, which may lay a foundation for the development of microbial bacterial agents applicable to saline soil remediation.【Method】 Strains with high-yield exopolysaccharide were isolated from the rhizosphere soil of saline-alkali grassland in Kashi, Xinjiang, and species identification was performed based on 16S rRNA gene sequence comparison and phylogenetic analysis. Moreover, the single factor and response surface method were used to optimize the medium components of the excellent strains to increase the yield of exopolysaccharides, and its effects on plant seedlings cultivated in saline-alkali soil and soil aggregate formation were verified by inoculation test.【Result】 A total of 19 exopolysaccharides-producing strains were screened, which were divided into 7 genera, of which Pseudomonas were the dominant. The exopolysaccharides yield was 236-1 544 mg/L, especially Pantoea sp. MQ A0. In addition, MQ A0 was capable of nitrogen fixation, indole-3-acetic acid(IAA)secretion, siderophores production and phosphorus solubilization. The optimal fermentation medium of MQ A0 was 12.5 mL/L glycerol, 9.0 g/L peptone, 5.5 g/L yeast exact and 5.1 g/L CaCO3. Compared with the initial fermentation process, the exopolysaccharides yield reached 2 436 mg/L, which increased by 57.8%. Pot experiments showed that the inoculation of Zea mays with the fermentation broth of MQ A0 had a significant promoting effect on seedling in saline-alkali soil, including the development of plant height, fresh weight, stem diameter, root number and total root length, which increased by 41.2%, 203.0%, 42.7%, 30.4%, 99.7%, respectively. And the formation of soil aggregate was significantly promoted by the EPS(P<0.05).【Conclusion】 The high-yield exopolysaccharide strain MQ A0 had various growth-promoting properties, and showed positive effects on plant growth and soil improvement in saline-alkali soil.

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    Regulation of Nitrogen Application on Peanut Seed Germination and Spermosphere Bacterial Community Structure Under Salt Stress
    XU Yang, ZHANG Rui-ying, DAI Liang-xiang, ZHANG Guan-chu, DING Hong, ZHANG Zhi-meng
    Biotechnology Bulletin    2024, 40 (2): 253-265.   DOI: 10.13560/j.cnki.biotech.bull.1985.2023-0753
    Abstract1627)   HTML10)    PDF(pc) (5168KB)(202)       Save

    【Objective】 Salt stress affects seed germination and peanut growth, and the internal regulatory mechanism of increasing seed germination rate and peanut yield by appropriate fertilization under salt stress was elucidated, and the relationship between the process and the bacterial structure of peanut spermosphere soil was analyzed, so as to provide theoretical and technical basis for improving peanut emergence and health rate, salt tolerance and stress resistance and peanut production capacity by regulating the microenvironment in spermosphere soil.【Method】 We set up three nitrogen levels of 0, 90, and 180 kg/hm2 with a salt-tolerant peanut variety(Huayu 25, HY25)as experimental materials. Potted experiments and high-throughput sequencing techniques were used to study the effects of nitrogen fertilizer application on the spermosphere bacterial community structure, germination and yield of peanut under salt stress.【Result】 The application of nitrogen fertilizer increased the germination rate and peanut yield, and the optimal nitrogen application was 90 kg/hm2. Proteobacteria, Actinobacteria, Firmicutes, Acidobacteria, Bacteroidetes, Chloroflexi, and Gemmatimonadetes were the dominant phyla in peanut spermosphere soil of different groups via 16S rRNA sequence. At the genus level, salt stress increased the relative abundance of beneficial bacteria Bacteroides, but induced a large number of harmful Streptococcus, whereas the relative abundance of the beneficial bacteria genera Bacillus, Sphingomonas, and Lysobacter decreased. Nitrogen application under salt stress may improve the spermosphere soil microenvironment and increase the relative abundance of beneficial soil bacteria Bacteroides, Bacillus, and Sphingomonas, which is helpful to soil restoration and fertility improvement, and can also enhance the stress resistance of peanuts.【Conclusion】 Appropriate nitrogen application under salt stress increases the relative abundances of beneficial bacteria in the spermosphere soil, thus which may improve the germination rate and salt tolerance, and finally increase the peanut yield under salt stress.

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    Genome-wide Identification and Expression Analysis of bHLH Transcription Factor Family in Dendrobium nobile
    WANG Jian, YANG Sha, SUN Qing-wen, CHEN Hong-yu, YANG Tao, HUANG Yuan
    Biotechnology Bulletin    2024, 40 (6): 203-218.   DOI: 10.13560/j.cnki.biotech.bull.1985.2023-0907
    Abstract1554)   HTML13)    PDF(pc) (13087KB)(64)       Save

    【Objective】 bHLH transcription factors exist widely in plants and are one of the important transcription factor families that regulate plant growth and development and respond to stress. The DnbHLHs transcription factor family was identified from the whole genome of Dendrobium nobile and its expression characteristics were analyzed, which laid the foundation for further study on the biological function of DnbHLHs transcription factor. 【Method】 The bHLH transcription factor family members were identified from the whole genome of D. nobile by bioinformatics method, and their physical and chemical characteristics, phylogenetic relationship, gene structure, chromosome localization and promoter cis-acting elements were analyzed. Transcriptome data and RT-qPCR were used to analyze the tissue expression patterns of DnbHLHs transcription factors and their expression patterns under different abiotic stresses. 【Result】 A total of 137 bHLH transcription factors were identified in D. nobile. The amino acid number of DnbHLHs protein was 85-1 260, the molecular weight was 9 855.23-140 024.19 Da, the isoelectric point was 4.52-10.66, the instability coefficient was 31.94-87.42, the fat solubility index was 56.66-106.06, and the hydrophilic index was -0.815-0.185. DnbHLHs were distributed on 19 chromosomes of D. nobile, all of which had a bHLH domain and were homologous to the bHLH of Arabidopsis thaliana, and there are collinear relationships between groups and species. Analysis of gene expression patterns showed that DnbHLHs family members had tissue specificity, and DnbHLH26 and DnbHLH33 were responsive to various stresses. 【Conclusion】 A total of 137 DnbHLHs transcription factor family members are identified from the whole genome of D. nobile. The physicochemical properties and conserved mods of the family members showe specificity and diversity. There are tissue expression differences among different DnbHLHs transcription factors, and DnbHLH26 is mainly induced by high salt and high temperature stress. DnbHLH33 is mainly induced by high salt, high temperature and low temperature stress.

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    Regulation of PIN-FORMED-mediated Polar Auxin Transport in Plant Gravitropism
    WANG Xian, PENG Ya-kun, CHEN Meng, KONG Meng-juan, TAN Shu-tang
    Biotechnology Bulletin    2024, 40 (3): 25-40.   DOI: 10.13560/j.cnki.biotech.bull.1985.2023-0882
    Abstract1535)   HTML22)    PDF(pc) (3051KB)(602)       Save

    Plant tropisms are the directional growth response of plants to environmental stimuli such as light or gravity. In gravitropism, plant organs sense directional environmental cues to control growth orientation to promote their survival. The phytohormone auxin and its polar transport serve as a major coordinative signal in plant gravitropism. The PIN family auxin exporters, through their dynamic polar subcellular localizations at the plasma membrane(PM), redirect polar auxin transport in response to environmental stimuli, thus the auxin gradients across tissues or organs underlie differential cell elongation and bending, which regulates the morphological formation and growth process of plants. In this review, we focus on recent advances in the mechanisms underlying the perception and signal transduction of gravity that take place within the sensing cells in the early process of gravitropism, the regulatory mechanisms of PIN-mediated auxin polar transport, PIN polar localization, and PM protein abundance.

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    Research Progress in DNA Methylation Sequencing Technology
    YUAN Ming-bo, YE Guo-hua, YANG Dan, SONG Dong-xue
    Biotechnology Bulletin    2024, 40 (5): 58-65.   DOI: 10.13560/j.cnki.biotech.bull.1985.2023-1166
    Abstract1509)   HTML18)    PDF(pc) (3195KB)(306)       Save

    DNA methylation is an important epigenetic modification that plays a key role in the growth and development of animals and plants, as well as in the occurrence of diseases and the regulation of gene expression. Clinically, DNA methylation tumor markers can serve as biological markers for the diagnosis, prognosis, and treatment of tumors. Accurate detection of DNA methylation is of great significance for elucidating the life mechanisms of biological growth, development, and disease occurrence. DNA methylation sequencing technology is a powerful tool for studying DNA methylation and is widely used for mapping DNA methylation in the genome. In recent years, in order to better detect DNA methylation site information, scientists have proposed a series of high-throughput DNA methylation sequencing technologies, which have improved sequencing detection sensitivity, reduced sequencing costs and experimental expenses, and greatly promoted the development of epigenomics. This article reviews a series of DNA methylation sequencing technologies, focusing on the technical principles and application scenarios of three sequencing technologies: WGBS, TAPS, and EM-seq. It also introduces sequencing methods for locating DNA methylation at the single-cell level, and finally prospects the future development direction.

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    Transcriptome Sequencing of Male Sterile Buds at Different Developmental Stages in Sapindus mukorossi ‘Qirui’
    LIAO Yang-mei, ZHAO Guo-chun, WENG Xue-huang, JIA Li-ming, CHEN Zhong
    Biotechnology Bulletin    2024, 40 (7): 197-206.   DOI: 10.13560/j.cnki.biotech.bull.1985.2024-0039
    Abstract1474)   HTML8)    PDF(pc) (4159KB)(269)       Save

    【Objective】 The objective of this work is to observe the cytological characteristics of the anther development in the male sterile Sapindus mukorossi ‘Qirui’, identify differentially expressed genes(DEGs)during the critical period of anther development,which provides a theoretical basis for deeply analyzing the molecular mechanism of the male sterility. 【Method】 By examining the cytological features of male anther development stages, RNA-Seq technology was used to compare the transcriptome of male buds at different stages: microspore mother cell stage(T1), tetrad stage(T2)and mononuclear microspore stage(T3), to identify key DEGs. 【Result】 The study revealed that the continuous expansion and proliferation of ‘Qirui’ tapetum and endothecium led to a chaotic anthecium structure and insufficient anthecium space, resulting in abortive pollen. Additionally, a decrease in the level of jasmonic acid(JA)from endogenous hormone determination was observed during male bud development. A total of 2 990 DEGs were detected through transcriptome analysis at the three anther development stages, of which 722 in T2_vs_T1(516 up-regulated and 206 down-regulated), and 1 741 in T3_vs_T2(765 up-regulated, 976 down-regulated). GO and KEGG analysis showed that these DEGs were primarily enriched in metabolic processes, cell division, pollen wall synthesis, hormone signal transduction, etc. Specifically, 32 DEGs were associated with pollen development, and 27 DEGs were linked to hormone synthesis and signal transduction. Nine DEGs were randomly selected for RT-qPCR analysis, and the results were consistent with the trend of RNA-Seq data, which proved the accuracy of transcriptome data. 【Conclusion】 The continuous proliferation of tapetum, delayed degeneration of endothecium, and continuous extrusion of microspores are the key factors leading to male sterility in S. mukorossi ‘Qirui’. Overall, genes related to material metabolism, cell division, hormone level and pollen development play crucial roles in the mechanism of male sterility.

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    Identification of Salt-tolerant Plant Growth-promoting Bacterium W-1 and Its Effect on the Salt-tolerance of Sainfoin(Onobrychis viciaefolia
    GAO Zhi-wei, WEI Ming, YU Zu-long, WU Guo-qiang, WEI Jun-long
    Biotechnology Bulletin    2024, 40 (4): 217-227.   DOI: 10.13560/j.cnki.biotech.bull.1985.2023-0905
    Abstract1451)   HTML14)    PDF(pc) (5470KB)(423)       Save

    Objective】To obtain salt-tolerant plant growth-promoting bacteria, several strains of salt-tolerant bacteria were isolated from plant roots in a saline-alkali soil in Jingtai, Gansu province, and some of them had obvious plant growth-promoting characteristics.【Method】Based on the previous work, a strain of salt-tolerant bacterium W-1 and a salt-sensitive plant sainfoin(Onobrychis viciaefolia)were selected as experimental materials, and the species classification, salt tolerance, acid and alkali tolerance and plant growth-promoting characteristics of strain W-1 were tested. The effects of strain W-1 on the physiological status of sainfoin treated with different concentrations of NaCl were investigated. 【Result】The salt-tolerant bacterium W-1 was Gram-positive, spore-producing, without capsule and flagellated. The 16S rDNA sequencing and comparison results showed that the bacterium W-1 was Bacillus siamensis. The maximum NaCl tolerance of strain W-1 was 13%, and the pH tolerance range was 4.5-8.5. W-1 has various plant growth-promoting functions, such as phosphate and potassium solubilization, nitrogen fixation, siderophores production, indole-3-acetic acid(IAA)production and 1-aminocyclopropane-1-carboxylic acid(ACC)deaminase activity. Strain W-1 significantly increased dry weight and fresh weight and promoted root growth of sainfoin seedlings. Under 100 and 150 mmol/L NaCl stresses, strain W-1 significantly increased the contents of soluble sugar, soluble protein, proline, chlorophyll, and catalase activity, decreased the content of hydrogen peroxide, leaf yellowing rate and mortality, and alleviated the adverse effects of salt stress on the growth of sainfoin seedlings.【Conclusion】The W-1 strain was an excellent plant growth- promoting bacterium with salt tolerance, acid and alkali tolerance, and enhanced the salt tolerance of legume forage.

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    Application and Risk of RNAi Technology in Potato Insect Pest Management
    WANG Ke-ran, YAN Jun-jie, LIU Jian-feng, GAO Yu-lin
    Biotechnology Bulletin    2024, 40 (9): 4-10.   DOI: 10.13560/j.cnki.biotech.bull.1985.2024-0472
    Abstract1435)   HTML16)    PDF(pc) (1813KB)(257)       Save

    Insect pest damages to crops is a threat to global food security, farmers urgently need safe and effective crop protection tools to sustainably have yields that meet ever-increasing global demand. RNAi(RNA interference)technology is a molecular biology technique for post-transcriptional regulation of gene silencing. Its principle is based on the binding of small double-stranded RNA molecules composed of 19-25 nucleotide pairs to the target mRNA, which triggers the degradation of the mRNA and thus leads to the silencing of the target gene. RNAi technology has been widely applied in crop pest management, focusing mainly on the prevention and control of Coleoptera, Hemiptera, and Lepidoptera pests targeting potatoes. On December 22, 2023, the world's first RNAi biopesticide was officially approved for commercialization. It is used to control the potato beetle, an internationally recognized important and devastating quarantine pest of potatoes, which is becoming increasingly resistant to pesticides. It is the world's first sprayable RNA biopesticide allowed for commercial use on crops, and it is a landmark green control of potato pests. While products based on RNAi technology are used for agricultural pest control, their resistance, off-target effects and potential risks to environmental safety still need to be considered. This article provides a comprehensive review on the feasibility of applying RNAi technology in potato insect pest management, its applications, and potential risks, aiming to elaborate on the status and prospects of RNAi technology in potato pest management, and provide theoretical references for integrating RNAi technology into the comprehensive management for potato pest management.

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    Establishment of CRISPR/Cas9-mediated Highly Efficient Gene Editing System in Microtubers of Potatoes
    SONG Qian-na, DUAN Yong-hong, FENG Rui-yun
    Biotechnology Bulletin    2024, 40 (9): 33-41.   DOI: 10.13560/j.cnki.biotech.bull.1985.2024-0536
    Abstract1393)   HTML10)    PDF(pc) (2707KB)(223)       Save

    【Objective】 The development of gene editing technology has made it possible to achieve precise molecular breeding in potato. Microtubers are ideal materials for genetic transformation, but its induction and genetic transformation are of genotype-dependence. Thus, establishing an efficient and widely used gene editing technology may provide technical support for precise molecular breeding for potatoes in future. 【Method】Cultivated tetraploid potato variety Qingshu No. 9 and Bingshu No. 6 were used to explore microtuber induction and genetic transformation. Concurrently, the CRISPR/Cas9 gene editing vector was transformed for genome editing. In addition, the screening system was tested in another three potato varieties. 【Result】 The efficient tuberizations of five potato varieties were obtained using stem segment with two leaves on solid media including 10% sucrose and 5 mg/L kinetin under dark condition, while induction effect varied. The optimal ratio of hormones for the regeneration of Qingshu No.9 was 0.5 mg/L 6-benzylaminopurine, 0.2 mg/L indole-3-acetic acid, 0.2 mg/L gibberellin, and 2 mg/L zeatin, under which the regeneration rate, the transformation frequency, genome editing rate was 41.5%, 51.9%, and 82.1%, respectively. Using the above screened the formula of hormones, genetic transformation and regeneration was efficiently achieved in another four potato varieties, and the genome editing rate was 63.2%, 33.3%, and 10% for Bingshu No.6, Desiree, and Jingshu No. 16, respectively. 【Conclusion】We established the highly efficient genetic transformation and regeneration system for five potato varieties, and genome editing can be completed for four potato varieties.

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    Genome-wide Identification and Expression Pattern Analysis of BZR Transcription Factor Gene Family of Melon
    ZANG Wen-rui, MA Ming, CHE Gen, HASI Agula
    Biotechnology Bulletin    2024, 40 (7): 163-171.   DOI: 10.13560/j.cnki.biotech.bull.1985.2024-0162
    Abstract1361)   HTML14)    PDF(pc) (4213KB)(351)       Save

    【Objective】 Brassinazole-resistant(BZR)is a plant specific transcription factor that plays a crucial role in plant growth and development. Identifying members of the CmBZR gene family across the whole melon genome, and analyzing the expression patterns of related gene, may provide a basis for further exploring the biological functions of the melon BZR gene family. 【Method】 Based on the whole melon genome data, the BZR family genes in melon were identified using BLAST. Bioinformatics methods were used to analyze the physicochemical properties, chromosomal distribution, gene structure, protein domains, phylogenetic evolution, and cis acting elements of the family proteins. Fluorescence quantitative PCR was used to verify the expression of BZR family genes in different tissues and fruits of melon at different growth and development stages. 【Result】 A total of 6 BZR genes were identified in cantaloupe, which can be divided into 5 subfamilies through phylogenetic analysis. CmBZR genes are distributed on chromosomes 3, 7, 8, 11, and 12. All BZR proteins have conserved domains. The promoter region of the CmBZR gene is significantly enriched with cis-acting elements related to growth and development, hormone signal transduction, and abiotic stress. CmBEH1-4 is highly expressed in stems, female flowers, and ovaries, and is also expressed in growth, maturity, respiratory climacteric, and late respiratory climacteric stages. 【Conclusion】 Six members of the melon CmBZR gene family have been systematically identified across the whole genome, and different genes are expressed in various tissues and growth stages of the melon,and their expression patterns are various.

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    Molecular Epigenetic Understanding of Seasonal Regulation of Flowering Time in Wheat
    NIU De, HE Yue-hui
    Biotechnology Bulletin    2024, 40 (10): 30-40.   DOI: 10.13560/j.cnki.biotech.bull.1985.2024-0959
    Abstract1359)   HTML27)    PDF(pc) (2521KB)(118)       Save

    Wheat(Triticum aestivum, AABBDD)is one of the most important food crops in the world, and is also the second staple food crop in China. The wide adaptability and ecological diversity of wheat varieties is partly determined by their flowering time. Furthermore, flowering time is an important agronomic trait that partly determines the yield and quality of wheat grain. Prolonged cold exposure in winter(winter cold)and increasing daylengths in spring and early summer are two key seasonal factors that ensure winter wheat flowering and bearing fruit at the proper time. The vernalization and photoperiodic pathways have been evolved in winter wheat to integrate the signals of winter cold and daylength changes with developmental state, leading to the floral transition at a proper season for seed production. In this review, we summarize molecular epigenetic progresses on wheat flowering-time regulation, discuss unsolved issues as well as prospects future directions in this field.

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    Analytical Approaches and Functional Insights for Microbiome Studies
    GAO Yun-yun, YANG Hai-fei, LYU Hu-jie, LIU Yong-xin
    Biotechnology Bulletin    2024, 40 (10): 98-107.   DOI: 10.13560/j.cnki.biotech.bull.1985.2024-0788
    Abstract1355)   HTML51)    PDF(pc) (8714KB)(397)       Save

    Microbiomes are crucial resources in life science, playing vital roles in advancing scientific knowledge, promoting human health, and improving environmental quality. The rapid evolution of second- and third-generation metagenomic technologies has greatly enhanced our understanding of microbial world. With the proliferation of microbiome data, the need to select appropriate analytical methods for efficiently extracting information has become increasingly essential. Here, we provide a comprehensive review of recent progress in microbiome studies, focusing on updated analytical tools for short-read second-generation sequencing data, including amplicon, culture-based methods, and metagenomic data. In addition, we offer the processing strategies for long-read third-generation sequencing data. We highlight the necessity for standardized data analysis workflows and present case studies that demonstrate the application of these methods in exploring microbiome interactions, particularly in plant and root-associated microbial systems. We discuss the strengths and limitations of various methods in analyzing microbiome composition, structure, and functionality, highlighting the potential of metagenomic data mining for practical applications. Finally, we address current limitations and challenges in microbiome study, and we discuss future trends toward the standardization and streamlining of microbiome study methodologies to accelerate progress in understanding microbiome functions and applications.

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    Banana Rhizosphere Microecology and Its Relationship with Fusarium Wilt Control
    QI Yan-xiang, XIE Yi-xian, PENG Jun, ZENG Fan-yun, ZHANG Xin
    Biotechnology Bulletin    2024, 40 (6): 57-67.   DOI: 10.13560/j.cnki.biotech.bull.1985.2023-1222
    Abstract1315)   HTML15)    PDF(pc) (2027KB)(420)       Save

    Fusarium wilt seriously endangers banana production, it causes yield loss and even no yield. It is spread by soil and transportation of seedlings, and chlamydospores of its pathogen survive in soil for 30 years. It is very difficult for the disease to be controlled, and it is always the focus of the government, academic field and banana industry. The experts, scholars, and related practitioners of domestic and abroad have done extensive and deep related research for solving this hard issue. This paper reviewed the previous research progress from the occurrence and harm of banana Fusarium wilt, the relationship between plant rhizosphere microecology and soil-borne diseases, banana rhizosphere microecology and their effects and contributions in Fusarium wilt control. The paper proposed the research trends of the interaction mechanism between Fusarium wilt and banana rhizosphere microecology, strain screening scope and isolating incubation methods, development and application of biocontrol agents with high efficiency and strong stability, aiming to provide references for subsequent in-depth study of banana rhizosphere microecology, the excavation and innovation utilization of excellent biological control resources and the efficient control of banana Fusarium wilt.

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    Role of Acetylation in the Pathogenic Process of Plant Pathogens
    WANG Li-chao, LI Huan, SHENG Ruo-cheng, LI Min, CHEN Feng-mao
    Biotechnology Bulletin    2024, 40 (5): 1-12.   DOI: 10.13560/j.cnki.biotech.bull.1985.2023-1179
    Abstract1297)   HTML104)    PDF(pc) (1188KB)(464)       Save

    Lysine acetylation, as a conservative post-translational modification, participates in a variety of biological processes. Numerous medical and plant pathological studies have revealed that acetylation plays an important regulatory role in the development of animal and plant diseases. Based on previous research progress, this article elaborates on the role of acetylation in plant pathogens from three perspectives involved in the pathogenesis of plant pathogens: Regulation of pathogen growth and pathogenicity, host plant resistance, and pathogen-host plant interaction. The aim is to understand the role of acetylation in the pathogenic process and to provide new ideas and theoretical support for the prevention and control of the pathogens.

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    CRISPR-Cas9 Gene Editing Technology and Its Research Progress in Poultry
    XIAO Yi-meng, YANG Wen, CHENG Yi-yi, LUO Gang
    Biotechnology Bulletin    2024, 40 (5): 38-47.   DOI: 10.13560/j.cnki.biotech.bull.1985.2023-1135
    Abstract1256)   HTML15)    PDF(pc) (3591KB)(742)       Save

    The CRISPR-Cas system, found in bacteria or archaea, serves as an adaptive immune system to counteract viral re-invasion by effectively cleaving nucleic acids. By leveraging the characteristics of Cas9 protein and its variants, researchers have devised a range of gene editing techniques that enable gene operations like gene knock-out or knock-in within cells, thereby facilitating genetic diversity in organisms. Additionally, these tools allow the manipulation of DNA or RNA through epigenetic modification to regulate gene expression, rendering them extensively employed in life science research. Poultry, as a significant agricultural species, plays a crucial role in supplying individuals with protein of superior quality. The utilization of CRISPR-Cas9 technology for gene editing and modification in avian genome is presently confined to the research phase, with remarkable advancements observed solely in chicken and quail. This paper presents an overview of CRISPR-Cas9 gene editing technology, encompassing its components, delivery methods, and optimization strategies, while also exploring its potential applications in poultry production and prevention and control of diseases. These insights lay the groundwork for the future development of CRISPR-Cas9 technology in the context of poultry system.

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    CRISPR/Cas9 Editing MeHNL Gene to Generate Cassava Plants with Low Cyanogenic Glycoside
    TONG Wei-jing, LUO Shu, LU Xin-lu, SHEN Jian-fu, LU Bai-yi, LI Kai-mian, MA Qiu-xiang, ZHANG Peng
    Biotechnology Bulletin    2024, 40 (9): 11-19.   DOI: 10.13560/j.cnki.biotech.bull.1985.2024-0520
    Abstract1240)   HTML18)    PDF(pc) (7011KB)(141)       Save

    【Objective】 Because cassava(Manihot esculenta Crantz)contains potentially toxic cyanogenic glucosides, its food safety is affected, which leads to the increase of processing costs. It is of great significance to cultivate cassava with low cyanogenic glycosides by biotechnology.【Method】 CRISPR/Cas9 technology was used to edit the α-hydroxynitrile lyase gene MeHNL. This gene encodes α-hydroxynitrile lyase that catalyzes the decomposition of cyanogenic glycosides. The editing target is located on its first exon, and 27 positive plants were obtained through Agrobacterium-mediated stable transformation.【Result】Sequencing analysis indicated successful editing in 26 out of 27 transgenic plants, and a high editing efficiency was achieved by 96.3%. The editing types predominantly comprised base insertions and deletions, alongside minor base substitutions and large fragment deletions. HCN colorimetric kit and HPLC analysis confirmed a significant reduction in hydrocyanic acid and cyanogenic glycosides in the mutant lines. Additionally, transgenic plants demonstrated slender leaves compared to non-transgenic counterparts, implicating MeHNL's impact on plant growth and development.【Conclusion】The cassava germplasm with low cyanide is obtained by CRISPR/Cas9 technology, which provides materials for exploring the study of cyanogenic glycoside metabolism on cassava growth and development.

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    Crop Breeding Based on Promoter Editing
    ZHANG Jie-ping, GUAN Yue-feng
    Biotechnology Bulletin    2024, 40 (10): 53-61.   DOI: 10.13560/j.cnki.biotech.bull.1985.2024-0787
    Abstract1236)   HTML19)    PDF(pc) (2451KB)(154)       Save

    To meet the growing demand for food from human being, the study on editing important gene-coding regions to produce beneficial traits is countless. Transcriptional regulation is crucial for controlling gene expression, including the control of important agronomic traits in crops. The interaction between cis-acting elements and trans-acting factors determines the spatiotemporal expression patterns and levels of genes, and transcription factor binding sites and enhancers etc. play an important role in those pathways. What's more, the promoter, as the largest and most important cis-acting element, can be modified to alter the expression patterns of target genes, and create beneficial traits in crops, which is an effective strategy different from coding region editing breeding. There are two applications of promoter editing strategies. One is directed modification to generate specific beneficial traits, and the other is to randomly generate new alleles within specific promoter regions, followed by phenotypic selection, also producing new genetic variation resources. In this review, we mainly discuss the application of promoter editing in crops, including increasing yield, improving quality, and enhancing tolerance to biotic and abiotic stresses. It aims to focus on the cutting-edge precision breeding of crops, providing research ideas and theoretical references for the development and application of promoter gene editing technology.

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    Research Progress in the Auxin Signaling Pathway Involved in the Regulation of Female Gametophyte Development in Arabidopsis
    YANG Jia-hong, LI Jing-yi, WU Jia-hao, HUANG You-mei, LIU Yan-fen, QIN Yuan, CAI Han-yang
    Biotechnology Bulletin    2024, 40 (7): 19-27.   DOI: 10.13560/j.cnki.biotech.bull.1985.2024-0109
    Abstract1214)   HTML24)    PDF(pc) (2828KB)(315)       Save

    Female gametophyte development is an important prerequisite for sexual reproduction in flowering plants, including two key events, macrosporogenesis and female gametophyte development. The development of the female gametophyte determines whether the plant can be properly fertilized and complete the reproduction of generations. Auxin is a widespread phytohormone in plants, and the polar distribution of auxin in the ovule ensures the normal development of the ovule. However, due to the complexity of the auxin signaling pathway and the fact that female germ cells are located inside the tissues, the regulatory network and molecular mechanism of the auxin signaling pathway in the female germline are still unclear. In this paper, we summarize the studies related to auxin synthesis, dynamic equilibrium and signaling, and review the studies on auxin regulation of macrosporogenesis and female gametophyte genesis. The aim of this paper is to provide a reference for the further study of the auxin signaling pathway on female germline establishment.

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    Regulatory Genes Mining Related to Transcriptome Sequencing and Phenolic Metabolism Pathway of Canarium album Fruit with Different Fresh Food Quality
    XIE Qian, JIANG Lai, HE Jin, LIU Ling-ling, DING Ming-yue, CHEN Qing-xi
    Biotechnology Bulletin    2024, 40 (3): 215-228.   DOI: 10.13560/j.cnki.biotech.bull.1985.2023-0747
    Abstract1198)   HTML11)    PDF(pc) (10116KB)(137)       Save

    【Objective】 The category and content of phenolic compounds in fruits are important quality traits closely related to the nutrition and flavor of Chinese olives(Canarium album). This work is to explore the molecular regulation mechanism of phenolic biosynthesis in Chinese olives.【Method】 The fruits of Chinese olives(low-phenol/high-phenol)with significant differences in total phenol content were taken as test materials, and the transcriptome analysis was carried out 80-160 d after flowering, and the differential genes of shikimic acid-hydrolyzed tannin/phenylpropane-flavonoid biosynthesis pathway were characterized, and the differentially expressed genes were analyzed by WGCNA, from which the transcription factors related to phenolic metabolism pathway were mined.【Result】 The 296 314 Unigenes were obtained, of which 73% were annotated to the database. A total of 1 628 differentially expressed genes(DEGs)were identified in the mature fruits of four varieties(lines)of Chinese olive. KEGG analysis showed that DEGs was significantly enriched in the “flavonoid biosynthesis” pathway of phenolic metabolism pathway. Furthermore, DEGs of shikimic acid-hydrolyzed tannins/phenylpropane-flavonoids biosynthetic pathway in fruit ripening process was characterized. Combining R2 and P values of each module and character in WGCNA analysis, four key modules were screened out. According to the regulation relationship of genes in modules, MCC topological analysis was used to mine key transcription factors in modules with degree value ≥1. A total of 137 transcription factors Unigenes were found to be co-expressed with 30 phenolic synthetic structural genes Unigenes. The functional annotations of the transcription factors Unigenes were from 35 gene families, the most of which were zinc finger proteins(C2C2, C3H, C2H2 and PHD), followed by B3, HB, MYB and NAC gene families.【Conclusion】 This study initially analyzed the differences in phenolic metabolic pathways in Chinese olive fresh food quality from a transcriptomic perspective, and concurrently excavated the differential genes regulating phenolic metabolism, which may provide an important basis for further investigation of the molecular mechanisms underlying the differences in Chinese olive fresh food quality.

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    Biotechnology Bulletin    2024, 40 (11): 1-2.  
    Abstract1190)   HTML1269)    PDF(pc) (1110KB)(219)       Save
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    Genome-wide Identification and Expression Analysis of the SWEET Gene Family in Camellia oleifera
    DU Bing-shuai, ZOU Xin-hui, WANG Zi-hao, ZHANG Xin-yuan, CAO Yi-bo, ZHANG Ling-yun
    Biotechnology Bulletin    2024, 40 (5): 179-190.   DOI: 10.13560/j.cnki.biotech.bull.1985.2024-0021
    Abstract1180)   HTML19)    PDF(pc) (6328KB)(395)       Save

    Objective】To explore the involvement of SWEET(sugars will eventually be exported transporters)in sugar metabolism and response to abiotic stress in Camellia oleifera.【Method】Bioinformatics methods were used to analyze the gene structure, protein motif, chromosome localization, collinearity, cis-acting elements of promoter region and upstream regulators in CoSWEETs family, and RT-qPCR was used to analyze the expression pattern of CoSWEETs in different periods, tissues and stress responses.【Result】Total 14 CoSWEET genes were identified in Camellia oleifera for the first time, they were localized on 10 chromosomes, and there were differences in intron-exon numbers between different members. With 1-2 MtN3 conserved domain, the 14 CoSWEETs were divided into four subgroups by phylogenetic tree, and there were similar gene structures and motif in the same subgroup. There were differences in number of introns and exons among different members. Based on the analysis of promoter cis-acting elements and upstream transcription factor predictions, multiple cis-elements related to development, plant hormones and environmental stresses were found in the promoter region, and their expressions were regulated by transcription factors, like ERF, DOF, BBR-BPC and MYB. RT-qPCR results showed that CoSWEETs highly expressed in the fruit and root, and the expression in the seeds was related to the developmental stages. Moreover, several genes responding to low-temperature, drought or salt stress, such as CoSWEET1, CoSWEET2 and CoSWEET17 were mined based the expression profile of CoSWEETs members under abiotic stresses of low temperature, drought and high salt.【Conclusion】The expression of CoSWEETs is regulated by various hormones and transcription factors, and plays a crucial role in the development of seeds and the response to stress in Camellia oleifera.

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    Construction of Pmepa1 Knockout TCMK1 Mouse Renal Tubular Epithelial Cell Line Using CRISPR/Cas9 Technology
    ZHANG Hong-min, LONG Wen, LAO Xiao-qing, CHEN Wen-yan, SHANG Xue-mei, WANG Hong-lian, WANG Li, SU Hong-wei, SHEN Hong-ping, SHEN Hong-chun
    Biotechnology Bulletin    2024, 40 (2): 73-79.   DOI: 10.13560/j.cnki.biotech.bull.1985.2023-0500
    Abstract1179)   HTML11)    PDF(pc) (3540KB)(150)       Save

    【Objective】 CRISPR/Cas9 technology was used to construct Pmepa1 knockdown TCMK1 mouse renal tubular epithelial cell lines and to explore the effect of Pmepa1 knockdown on TGF-β-stimulated TCMK1 cell fibrosis, which provides a cell model for studying the role of Pmepa1 in fibrosis models. 【Method】 The pX333 vector was created and transfected into TCMK1 cells in accordance with the CRISPR/Cas9 design principle. Flow sorting of mCherry-positive cells, monoclonal cell expansion, and sequencing were used to identify the Pmepa1 knockout cells, and Western blot was used to confirm the knockout status of the Pmepa1 gene. Pmepa1 knockdown was confirmed using a Western blot. By using RT-PCR and Western blot, the effects of Pmepa1 knockdown on TGF-1-induced R-Smad activation and fibrosis were examined.【Result】 The sgRNA was designed in exon 2 of Pmepa1 48 h after transfection of TCMK1 cells with pX333-Pmepa1 plasmid, the expression of mCherry fluorescent protein was observed, and flow cytometric analysis of the transfection efficiency was about 17%. pX333-Pmepa1 plasmid transfection-positive cells were cultured by flow sorting and monoclonal amplification to obtain 19 cell clones. PCR amplification sequencing analysis of the sgRNA target sequence at the PMEAP1 gene locus revealed 2 double allele shift mutant cells, and Western blot verified the absence of Pmepa1 protein expression, indicating that the Pmepa1 knockdown TCMK1 cell line was successfully constructed, compared with the non-knockdown cells, the Pmepa1 knockdown cells were stimulated by TGF-β, p-Smad2 and p-Smad3 expression was significantly elevated in Pmepa1 knockout cells compared with non-knockout cells, and the expressions of the fibrogenic genes Fibronectin and Collagen I were promoted. 【Conclusion】 CRISPR/Cas9 technique was used to construct the renal tubular epithelial cell line of PMEPA1 knockout TCMK1 mice successfully, which established the cell model for the functional study of PMEPA1 protein and the important role of PMEPA1 in fibrosis.

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    Development and Prospects of Rice Pan-genomics: Important Tools and Applications
    HE Wen-chuang, XU Qiang, QIAN Qian, SHANG Lian-guang
    Biotechnology Bulletin    2024, 40 (10): 9-18.   DOI: 10.13560/j.cnki.biotech.bull.1985.2024-0669
    Abstract1179)   HTML32)    PDF(pc) (3171KB)(130)       Save

    Unlike a single genome, a pan-genome generally refers to a data set that contains all genomic information in a species or population. Over the past decade, pan-genomics has gradually become a research hotspot in rice, and relevant pan-genomes and tools have been widely applied in many downstream research fields such as population genetics, evolutionary biology and biological breeding practices. This paper focuses on the development process and application prospect of rice pan-genomics, reviews the development of connotation of rice pan-genomics and the timeline of its research results, summarizes the existing representative and important achievement tools of rice pan-genomics and their main applications in different fields, and looks forward to the challenges and development prospects for rice pan-genomics.

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    RNA Silencing Efficiency Affected by RNA Structure
    ZHAO Jian-hua, GAO Feng, LIU Qing-yan, GUO Hui-shan
    Biotechnology Bulletin    2024, 40 (10): 19-29.   DOI: 10.13560/j.cnki.biotech.bull.1985.2024-0857
    Abstract1145)   HTML25)    PDF(pc) (2367KB)(117)       Save

    Small RNA(sRNA)-mediated RNA silencing(or RNA interference)is a conserved mechanism for regulating gene expression in eukaryotes. Endogenous or exogenous double-stranded RNAs(dsRNAs)are processed into sRNAs, which recognize complementary mRNAs or DNAs in a sequence-specific manner, and regulate gene expression at either the posttranscriptional level or the transcriptional level by degrading mRNAs, inhibiting translation and DNA methylation. Owing to the target-specific actions of dsRNAs, RNAi-based strategies have become ubiquitous tools in the study of gene function, the development of RNA drugs, the design of molecular breeding methods, and the exploitation of biopesticides. The discovery of the bidirectional transfer among species and function of sRNAs provides a theoretical basis for exploiting RNAi-based technologies. Previous studies have shown that several factors affect the efficiency of dsRNA-induced RNAi, such as the size and dose of dsRNAs, as well as the application method. The complex structure of RNA leads to its functional diversity in vivo. Here, we described the principles of RNAi-based strategies for crop protection, including host-induced gene silencing(HIGS), spray-induced gene silencing(SIGS)and microbe-induced gene silencing(MIGS). We summarized the experimental evidence that the structures of target mRNAs and sRNAs affect RNAi efficiency, aiming to deepen the understanding of the RNA structure affecting RNAi efficiency, to provide experience for target screening and dsRNA design, and provide reference for the development of efficient RNAi technology. We also reviewed the representative methods and computational tools for detecting and predicting RNA structures, thereby providing approaches in establishing efficient RNAi-based techniques.

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    Studies on the Growth-promoting Effect of Bacillus Strain from Rhizosphere in Ionic Rare Earth Ores
    LI Xi, BIAN Zi-jun, NING Zhou-shen, LIU Hong-yu, ZENG Bing, DONG Wei
    Biotechnology Bulletin    2024, 40 (11): 259-268.   DOI: 10.13560/j.cnki.biotech.bull.1985.2024-0292
    Abstract1130)   HTML1)    PDF(pc) (3972KB)(19)       Save

    【Objective】 To explore innovative ways for the development of microbial fertilizers, rhizosphere microbial resources of plants are being mined from ionic rare earth mines, of which growth-promoting bacteria with multiple functions were isolated and identified, and their biological characteristics and promoting effects were also investigated. 【Method】 The strain DW019 isolated from the rhizosphere in an ionic rare earth mining area in Gannan of Jiangxi province was identified based on 16S rRNA gene sequence analysis and culture characteristics, classifying its genus and species. Its growth-promoting characteristics was preliminarily determined by the detection of production of IAA, iron carriers, and ammonia. The antagonistic effect of the strain against typical soil-borne disease causative organisms was measured by using the plate-confrontation method. Finally, the growth indexes, chlorophyll, antioxidant enzyme activities, and nutritional quality of lettuce(Lactuca sativa)were measured by potting experiments to determine the growth-promoting effect of strain DW019 on lettuce. 【Result】 The strain was identified as Bacillus cereus DW019, with the ability of producing IAA, iron carrier and ammonia. The strain showed strong inhibition effects against Thanatephorus cucumeris, Fusarium moniliforme Sheldon and Alternaria alternata(Fries)Keissler. Compared to the control group, the above-ground part biomass, biological indices of potted lettuce were significantly improved by the addition of different concentrations of DW019, respectively, and its chlorophyll content, water content, vitamin C, soluble sugars, malondialdehyde(MDA)and peroxidase(POD)were also significantly promoted. 【Conclusion】 Bacillus cereus DW019 is a multifunctional plant growth-promoting rhizobacterium with phytohormone-producing and biocontrol ability, which has strong inhibitory effect on a variety of soil-borne disease-causing bacteria, and significantly promotes plant growth and improves plant quality.

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