Biotechnology Bulletin ›› 2024, Vol. 40 ›› Issue (6): 152-160.doi: 10.13560/j.cnki.biotech.bull.1985.2023-1127
Previous Articles Next Articles
WANG Yu-shu1(), ZHAO Lin-lin1, ZHAO Shuang1, HU Qi1, BAI Hui-xia1, WANG Huan2, CAO Ye-ping1, FAN Zhen-yu1
Received:
2023-12-01
Online:
2024-06-26
Published:
2024-06-24
WANG Yu-shu, ZHAO Lin-lin, ZHAO Shuang, HU Qi, BAI Hui-xia, WANG Huan, CAO Ye-ping, FAN Zhen-yu. Cloning and Expression Analysis of BrCYP83B1 Gene in Chinese Cabbage[J]. Biotechnology Bulletin, 2024, 40(6): 152-160.
引物名称Primer name | 序列Sequence(5'-3') | 用途 Application |
---|---|---|
BrCYP83B1-F | GC GTCGACATGGATCTCTTCTTGATTATTGCCG | 片段扩增Fragment amplification |
BrCYP83B1-R | G GAATTCTCAAATGTGCGTCCTTGGTGC | 片段扩增Fragment amplification |
RT-CYP83B1-F | CGCAAGTTTCCGACCCGTTAGAG | 荧光定量PCR RT-qPCR |
RT-CYP83B1-R | ACAGTTGGTGAAGGACAAGAGAAGC | 荧光定量PCR RT-qPCR 荧光定量PCR RT-qPCR |
RT-BrActin-F | GGAGCTGAGAGATTCCGTTG | |
RT-BrActin-R | GAACCACCACTGAGGACGAT | 荧光定量PCR RT-qPCR |
Table 1 Primer used in this study
引物名称Primer name | 序列Sequence(5'-3') | 用途 Application |
---|---|---|
BrCYP83B1-F | GC GTCGACATGGATCTCTTCTTGATTATTGCCG | 片段扩增Fragment amplification |
BrCYP83B1-R | G GAATTCTCAAATGTGCGTCCTTGGTGC | 片段扩增Fragment amplification |
RT-CYP83B1-F | CGCAAGTTTCCGACCCGTTAGAG | 荧光定量PCR RT-qPCR |
RT-CYP83B1-R | ACAGTTGGTGAAGGACAAGAGAAGC | 荧光定量PCR RT-qPCR 荧光定量PCR RT-qPCR |
RT-BrActin-F | GGAGCTGAGAGATTCCGTTG | |
RT-BrActin-R | GAACCACCACTGAGGACGAT | 荧光定量PCR RT-qPCR |
Fig. 3 Prediction of secondary(A)and tertiary structure(B)of BrCYP83B1 protein Capital letters indicate amino acid sequence of BrCYP83B1 protein. Lowercase letters indicate different secondary structures, where h refers to α helix, c to random coil, e to extended strand, and t to β turn
Fig. 4 Multiple sequence alignment of BrCYP83B1 with its homologous protein from other plant species+ BnCYP83B1: Brassica napus, CDY35791.1; RsCYP83B1: Raphanus sativus, CDY35791.1; AtCYP83B1: Arabidopsis thaliana, NP_194878.1; BoCYP83B1: Brassica oleracea var. italica, APZ76511.1; ItCYP83B1: Isatis tinctoria, APY26702.1; EsCYP83B1: Eutrema salsugineum, XP_006396006.1; EvCYP83B1: Eruca vesicaria subsp. sativa, AGS49168.1; CrCYP83B1: Capsella rubella, XP_006285988.1; ThCYP83B1: Tarenaya hassleriana, XP_010530341.1; BcCYP83B1: Brassica rapa subsp. chinensis, HM347235.1. Amino acids highlighted in black, pink, and blue respectively represent residues completely conserved, partially conserved and similar
元件名称Name of element | 序列Sequence(5'-3') | 元件类型Type of element | 数量Amount |
---|---|---|---|
TGACG-motif | TGACG | 茉莉酸甲酯响应元件Cis-acting regulatory element involved in the MeJA-responsiveness | 3 |
CGTCA-motif | CGTCA | 茉莉酸甲酯响应元件Cis-acting regulatory element involved in the MeJA-responsiveness | 2 |
GT1-motif | GGTTAA | 光响应元件Light responsive element | 2 |
ABRE | AACCCGG | 脱落酸响应元件Cis-acting element involved in the abscisic acid responsiveness | 1 |
Box 4 | ATTAAT | 光响应元件Light responsive element | 1 |
G-Box | CACGTT | 光响应元件Light responsive element | 1 |
GARE-motif | TCTGTTG | 赤霉素响应元件Gibberellin-responsive element | 1 |
MRE | AACCTAA | 光响应元件Light responsive element | 1 |
TCA-element | CCATCTTTTT | 水杨酸响应元件Cis-acting element involved in salicylic acid responsiveness | 1 |
Table 2 Upstream elements in BrCYP83B1 promoter
元件名称Name of element | 序列Sequence(5'-3') | 元件类型Type of element | 数量Amount |
---|---|---|---|
TGACG-motif | TGACG | 茉莉酸甲酯响应元件Cis-acting regulatory element involved in the MeJA-responsiveness | 3 |
CGTCA-motif | CGTCA | 茉莉酸甲酯响应元件Cis-acting regulatory element involved in the MeJA-responsiveness | 2 |
GT1-motif | GGTTAA | 光响应元件Light responsive element | 2 |
ABRE | AACCCGG | 脱落酸响应元件Cis-acting element involved in the abscisic acid responsiveness | 1 |
Box 4 | ATTAAT | 光响应元件Light responsive element | 1 |
G-Box | CACGTT | 光响应元件Light responsive element | 1 |
GARE-motif | TCTGTTG | 赤霉素响应元件Gibberellin-responsive element | 1 |
MRE | AACCTAA | 光响应元件Light responsive element | 1 |
TCA-element | CCATCTTTTT | 水杨酸响应元件Cis-acting element involved in salicylic acid responsiveness | 1 |
Fig. 6 Expression pattern of BrCYP83B1 A: Expression of BrCYP83B1 in different tissues. B: Expression of BrCYP83B1 treated with MeJA, SA and ABA. Different lowercase letters indicate significant differences in relative expression at the P<0.05 level
Fig. 7 Electrophoretic detection on double enzyme digestion of plant expression vector and PCR of Agro-bacterium tumefaciens liquid M: DNA marker; 1: double digestion products; 2: PCR products of Agrobacterium tumefaciens liquid
[1] |
Kim YB, Chun JH, Kim HR, et al. Variation of glucosinolate accumulation and gene expression of transcription factors at different stages of Chinese cabbage seedlings under light and dark conditions[J]. Nat Prod Commun, 2014, 9(4): 533-537.
pmid: 24868877 |
[2] | Artemyeva AM, Solovyeva AE. Quality evaluation of some cultivar types of leafy Brassica rapa[J]. Acta Hortic, 2006(706): 121-128. |
[3] |
杜海, 冉凤, 刘静, 等. 拟南芥硫苷生物合成相关基因的组织和胁迫诱导表达谱的全基因组分析[J]. 中国农业科学, 2016, 49(15): 2879-2897.
doi: 10.3864/j.issn.0578-1752.2016.15.003 |
Du H, Ran F, Liu J, et al. Genome-wide expression analysis of glucosinolate biosynthetic genes in Arabidopsis across diverse tissues and stresses induction[J]. Sci Agric Sin, 2016, 49(15): 2879-2897. | |
[4] | 雷建军, 陈长明, 陈国菊, 等. 硫苷及其生物合成分子生物学机理研究进展[J]. 华南农业大学学报, 2019, 40(5): 59-70. |
Lei JJ, Chen CM, Chen GJ, et al. Progress in glucosinolates and its molecular mechanism of biosynthesis[J]. J South China Agric Univ, 2019, 40(5): 59-70. | |
[5] | 秦晗, 张文姗, 王猛, 等. 四个芸薹属物种硫苷含量与种类分析及特殊硫苷成分的种间导入[J]. 植物遗传资源学报, 2020, 21(1): 94-104. |
Qin H, Zhang WS, Wang M, et al. Characterizing glucosinolates of four Brassica species and interspecific transferring of specific glucosinolates[J]. J Plant Genet Resour, 2020, 21(1): 94-104. | |
[6] |
Nintemann SJ, Hunziker P, Andersen TG, et al. Localization of the glucosinolate biosynthetic enzymes reveals distinct spatial patterns for the biosynthesis of indole and aliphatic glucosinolates[J]. Physiol Plant, 2018, 163(2): 138-154.
doi: 10.1111/ppl.12672 pmid: 29194649 |
[7] | Zhu B, Wang ZZ, Yang J, et al. Isolation and expression of glucosinolate synthesis genes CYP83A1 and CYP83B1 in Pak Choi(Brassica rapa L. ssp. chinensis var. communis(N. Tsen & S.H. Lee)Hanelt)[J]. Int J Mol Sci, 2012, 13(5): 5832-5843. |
[8] |
Hull AK, Vij R, Celenza JL. Arabidopsis cytochrome P450s that catalyze the first step of tryptophan-dependent indole-3-acetic acid biosynthesis[J]. Proc Natl Acad Sci USA, 2000, 97(5): 2379-2384.
doi: 10.1073/pnas.040569997 pmid: 10681464 |
[9] |
Bak S, Tax FE, Feldmann KA, et al. CYP83B1, a cytochrome P450 at the metabolic branch point in auxin and indole glucosinolate biosynthesis in Arabidopsis[J]. Plant Cell, 2001, 13(1): 101-111.
doi: 10.1105/tpc.13.1.101 pmid: 11158532 |
[10] | 程文财, 丁聪聪, 赵云, 等. 油菜BnCYP83B1基因的克隆与生长素相互作用的研究[J]. 四川大学学报: 自然科学版, 2014, 51(5): 1092-1098. |
Cheng WC, Ding CC, Zhao Y, et al. Cloning and mRNA expression of BnCYP83B1 gene under 2, 4-D hormone in Brassica napus[J]. J Sichuan Univ Nat Sci Ed, 2014, 51(5): 1092-1098. | |
[11] | 赵桂红, 石宏, 张妮妮, 等. 菘蓝CYP83B1基因的克隆与表达分析[J]. 植物科学学报, 2017, 35(1): 64-72. |
Zhao GH, Shi H, Zhang NN, et al. Cloning and expression analysis of CYP83B1 from Isatis indigotica Fort[J]. Plant Sci J, 2017, 35(1): 64-72. | |
[12] | 徐蕊. 西兰花CYP83B1基因的克隆及功能鉴定[D]. 哈尔滨: 东北农业大学, 2018. |
Xu R. Cloning and functional identification of BoCYP83B1 from broccoli(Brassica oleracea var. italica)[D]. Harbin: Northeast Agricultural University, 2018. | |
[13] | Zang YX, Lim MH, Park BS, et al. Metabolic engineering of indole glucosinolates in Chinese cabbage plants by expression of Arabidopsis CYP79B2, CYP79B3, and CYP83B1[J]. Mol Cells, 2008, 25(2): 231-241. |
[14] |
Bridge LJ, Mirams GR, Kieffer ML, et al. Distinguishing possible mechanisms for auxin-mediated developmental control in Arabidopsis: models with two Aux/IAA and ARF proteins, and two target gene-sets[J]. Math Biosci, 2012, 235(1): 32-44.
doi: 10.1016/j.mbs.2011.10.005 pmid: 22067512 |
[15] |
González-Romero ME, Rivera C, Cancino K, et al. Bioengineering potato plants to produce benzylglucosinolate for improved broad-spectrum pest and disease resistance[J]. Transgenic Res, 2021, 30(5): 649-660.
doi: 10.1007/s11248-021-00255-w pmid: 33956271 |
[16] | 赵爽, 周羽琪, 杨旭妍, 等. 硒硫互作对白菜芽苗菜硫代葡萄糖苷含量及抗氧化性的影响[J]. 食品科学, 2023, 44(1): 22-29. |
Zhao S, Zhou YQ, Yang XY, et al. Effect of selenium and sulfur interaction on the glucosinolate content and antioxidant activity of sprouts of Chinese cabbage[J]. Food Sci, 2023, 44(1): 22-29. | |
[17] | Connolly EL, Sim M, Travica N, et al. Glucosinolates from cruciferous vegetables and their potential role in chronic disease: investigating the preclinical and clinical evidence[J]. Front Pharmacol, 2021, 12: 767975. |
[18] | Zang YX, Ge JL, Huang LH, et al. Leaf and root glucosinolate profiles of Chinese cabbage(Brassica rapa ssp. pekinensis)as a systemic response to methyl jasmonate and salicylic acid elicitation[J]. J Zhejiang Univ Sci B, 2015, 16(8): 696-708. |
[19] | Harun S, Abdullah-Zawawi MR, Goh HH, et al. A comprehensive gene inventory for glucosinolate biosynthetic pathway in Arabidopsis thaliana[J]. J Agric Food Chem, 2020, 68(28): 7281-7297. |
[20] | Sun B, Jiang M, Yuan Q, et al. Bioinformatics analysis of the gene CYP83B1 in cabbage(Brassica oleracea var. capitata)[C]// Proceedings of the 2018 3rd International Workshop on Materials Engineering and Computer Sciences(IWMECS 2018). January 27-28, 2018. Jinan, China. Paris, France: Atlantis Press, 2018: 328-331. |
[21] |
Hoecker U, Toledo-Ortiz G, Bender J, et al. The photomorphogenesis-related mutant red1 is defective in CYP83B1, a red light-induced gene encoding a cytochrome P450 required for normal auxin homeostasis[J]. Planta, 2004, 219(2): 195-200.
pmid: 14963708 |
[22] | Wang JW, Mao SX, Wu Q, et al. Effects of LED illumination spectra on glucosinolate and sulforaphane accumulation in broccoli seedlings[J]. Food Chem, 2021, 356: 129550. |
[23] | 费丹丹, 王军伟, 吴小媚, 等. 细胞色素P450基因家族参与植物硫代葡萄糖苷生物合成的研究进展[J]. 中国蔬菜, 2023(5): 30-41. |
Fei DD, Wang JW, Wu XM, et al. Research progress of cytochrome P450 gene family taking part in biosynthesis of plant glucosinolates[J]. China Veg, 2023(5): 30-41. | |
[24] | Xu R, Kong WW, Peng YF, et al. Identification and expression pattern analysis of the glucosinolate biosynthetic gene BoCYP83B1 from broccoli[J]. Biol Plant, 2018, 62(3): 521-533. |
[25] |
Mewis I, Appel HM, Hom A, et al. Major signaling pathways modulate Arabidopsis glucosinolate accumulation and response to both phloem-feeding and chewing insects[J]. Plant Physiol, 2005, 138(2): 1149-1162.
doi: 10.1104/pp.104.053389 pmid: 15923339 |
[26] | Fritz VA, Justen VL, Bode AM, et al. Glucosinolate enhancement in cabbage induced by jasmonic acid application[J]. HortScience, 2010, 45(8): 1188-1191. |
[27] | Pérez-Balibrea S, Moreno DA, García-Viguera C. Improving the phytochemical composition of broccoli sprouts by elicitation[J]. Food Chem, 2011, 129(1): 35-44. |
[28] | Sun B, Yan HZ, Zhang F, et al. Effects of plant hormones on main health-promoting compounds and antioxidant capacity of Chinese kale[J]. Food Res Int, 2012, 48(2): 359-366. |
[29] |
Kliebenstein DJ, Figuth A, Mitchell-Olds T. Genetic architecture of plastic methyl jasmonate responses in Arabidopsis thaliana[J]. Genetics, 2002, 161(4): 1685-1696.
doi: 10.1093/genetics/161.4.1685 pmid: 12196411 |
[30] | Wang ZY, Yang RQ, Guo LP, et al. Effects of abscisic acid on glucosinolate content, isothiocyanate formation and myrosinase activity in cabbage sprouts[J]. Int J Food Sci Tech, 2015, 50(8): 1839-1846. |
[31] | 陈微. 外源激素对萝卜肉质根硫苷和萝卜硫素含量的影响[D]. 南京: 南京农业大学, 2017. |
Chen W. Effect of exogenous hormone on the contents of glucosinolate and sulforaphane in radish(Raphanus sativus L.)taproot[D]. Nanjing: Nanjing Agricultural University, 2017. |
[1] | CHANG Xue-rui, WANG Tian-tian, WANG Jing. Identification and Analysis of E2 Gene Family in Pepper(Capsicum annuum L.) [J]. Biotechnology Bulletin, 2024, 40(6): 238-250. |
[2] | LIU Rong, TIAN Min-yu, LI Guang-ze, TAN Cheng-fang, RUAN Ying, LIU Chun-lin. Identification and Induced-expression Analysis of REVEILLE Family in Brassica napus L. [J]. Biotechnology Bulletin, 2024, 40(6): 161-171. |
[3] | LI Bo-jing, ZHENG La-mei, WU Wu-yun, GAO Fei, ZHOU Yi-jun. Evolution, Expression, and Functional Analysis of the HSP20 Gene Family from Simmondisa chinensis [J]. Biotechnology Bulletin, 2024, 40(6): 190-202. |
[4] | HAO Si-yi, ZHANG Jun-ke, WANG Bin, QU Peng-yan, LI Rui-de, CHENG Chun-zhen. Cloning and Expression Analysis of Banana EARLY FLOWERING 3(ELF3)Genes [J]. Biotechnology Bulletin, 2024, 40(5): 131-140. |
[5] | LI Jia-xin, LI Hong-yan, LIU Li-e, ZHANG Tian, ZHOU Wu. Identification and Expression Analysis of the NRAMP Family in Seabuckthorn Under Lead Stress [J]. Biotechnology Bulletin, 2024, 40(5): 191-202. |
[6] | LOU Yin, GAO Hao-jun, WANG Xi, NIU Jing-ping, WANG Min, DU Wei-jun, YUE Ai-qin. Identification and Expression Pattern Analysis of GmHMGS Gene in Soybean [J]. Biotechnology Bulletin, 2024, 40(4): 110-121. |
[7] | DU Ze-guang, REN Shao-wen, ZHANG Feng-qin, LI Mei-lan, LI Gai-zhen, QI Xian-hui. Cloning,Expression and Functional Identification of BrMLP328 Gene in Brassica rapa subsp. pekinensis [J]. Biotechnology Bulletin, 2024, 40(4): 122-129. |
[8] | CHEN Chun-lin, LI Bai-xue, LI Jin-ling, DU Qing-jie, LI Meng, XIAO Huai-juan. Identification and Expression Analysis of Epidermal Patterning Factor (EPF) Genes in Cucumis melo [J]. Biotechnology Bulletin, 2024, 40(4): 130-138. |
[9] | XIAO Ya-ru, JIA Ting-ting, LUO Dan, WU Zhe, LI Li-xia. Cloning and Expression Analysis of CsERF025L Transcription Factor in Cucumber [J]. Biotechnology Bulletin, 2024, 40(4): 159-166. |
[10] | LIU Huan-huan, YANG Li-chun, LI Huo-gen. Cloning and Functional Analysis of LtMYB305 in Liriodendron tulipifera [J]. Biotechnology Bulletin, 2024, 40(4): 179-188. |
[11] | ZHONG Yun, LIN Chun, LIU Zheng-jie, DONG Chen-wen-hua, MAO Zi-chao, LI Xing-yu. Cloning and Prokaryotic Expression Analysis of Asparagus Saponin Synthesis Related Glycosyltransferase Genes [J]. Biotechnology Bulletin, 2024, 40(4): 255-263. |
[12] | YANG Wei-cheng, SUN Yan, YANG Qian, WANG Zhuang-lin, MA Ju-hua, XUE Jin-ai, LI Run-zhi. Genome-wide Identification of the FAX family in Gossypium hirsutum and Functional Analysis of GhFAX1 [J]. Biotechnology Bulletin, 2024, 40(3): 155-169. |
[13] | YANG Yan, HU Yang, LIU Ni-ru, YIN Lu, YANG Rui, WANG Peng-fei, MU Xiao-peng, ZHANG Shuai, CHENG Chun-zhen, ZHANG Jian-cheng. Cloning and Functional Analysis of MbbZIP43 Gene in ‘Hongmantang’ Red-flesh Apple [J]. Biotechnology Bulletin, 2024, 40(2): 146-159. |
[14] | REN Yan-jing, ZHANG Lu-gang, ZHAO Meng-liang, LI Jiang, SHAO Deng-kui. cDNA Yeast Library Construction of Chinese Cabbage Seeds and Screening and Analysis of BrTTG1 Interacting Proteins [J]. Biotechnology Bulletin, 2024, 40(2): 223-232. |
[15] | ZHU Yi, LIU Tang-jing, GONG Guo-yi, ZHANG Jie, WANG Jin-fang, ZHANG Hai-ying. Cloning and Expression Analysis of ClPP2C3 in Citrullus lanatus [J]. Biotechnology Bulletin, 2024, 40(1): 243-249. |
Viewed | ||||||
Full text |
|
|||||
Abstract |
|
|||||