Genome-wide Identification of Potato WOX Gene Family and Its Expression Analysis in in vitro Regeneration and Abiotic Stress

doi:10.13560/j.cnki.biotech.bull.1985.2023-0947

Abstract

Abstract:

【Objective】 The WUSCHEL-related homeobox（WOX）gene family is plant-specific, which plays important roles in plant growth and development, regulation of stem cell differentiation, and response to adverse stress. Identification of potato WOX gene family may provide a theoretical basis and gene resources for potato genetic improvement.【Method】 Based on the WOX protein sequences of Arabidopsis thaliana, tomato, tobacco and rice, all members of potato WOX gene family were identified by HMMER 3.0 and BLASTP, MCScanX was used for collinear analysis, and the phylogenetic tree was constructed using neighbor-joining method. Physical and chemical properties, gene structure, protein motif, and transcription factor binding sites of potato WOX members were analyzed by ExPASy, GSDS and other software. Expression patterns of StWOXs were analyzed based on potato transcriptome data from PGSC database. RT-qPCR assay was carried out to analyze relative expression of StWOX5 in four potato varieties（lines）with different in vitro regeneration abilities.【Result】 11 members of potato WOX gene family were identified, distributed on five chromosomes, and classified into three clades, namely WUS, intermediate and ancient, with differences in gene structure, motif composition, transcription factor binding sites. The results showed that members of potato WOX gene family were involved in in vitro regeneration and abiotic stress responses. StWOX4/5/11/13 were specifically expressed in callus, StWUS and StWOX1/3c/4/5/13 were up-regulated under mannitol treatment, StWOX3a/11 were up-regulated under salt treatment, and StWOX4/5/13 were up-regulated under heat stress. The relative expression of StWOX5 was positively correlated with the differentiation rate.【Conclusion】 Different potato WOX genes had potential functional diversity. StWOX5 may promote potato regeneration in vitro and participate in abiotic stress responses.

Key words: Solanum tuberosum L., WOX gene family, callus, gene expression

ZHANG Yu, SHI Lei, GONG Lei, NIE Feng-jie, YANG Jiang-wei, LIU Xuan, YANG Wen-jing, ZHANG Guo-hui, XIE Rui-xia, ZHANG Li. Genome-wide Identification of Potato WOX Gene Family and Its Expression Analysis in in vitro Regeneration and Abiotic Stress[J]. Biotechnology Bulletin, 2024, 40(3): 170-180.

Figures/Tables 9

Table 1 Primers used in real-time quantitative PCR

基因 Gene	引物序列 Primer sequence（5'-3'）
StWOX5	F: TGCTGAAACTGAAAAGCTTAGACT
StWOX5	R: GGATGATCCATCTCAGCTCCA
EF1α	F: CAAGGATGACCCAGCCAAG
EF1α	R: TTCCTTACCTGAACGCCTGT

Table 2 Information of WOX gene family members in S. tuberosum

序号 No.	基因 Gene	基因ID Gene ID	染色体上位置 Position in chromosome	氨基酸数量Number of amino acids	相对分子质量Molecular weight/kD	等电点Isoelectric points	亚细胞定位Subcellular localization
1	StWUS	Soltu.DM.02G023940	chr.02:37 323 370-37 324 685	263	29.8	6.55	细胞核Nuclear
2	StWOX1	Soltu.DM.03G033140	chr.03:56 859 969-56 862 746	391	44.1	6.71	细胞核Nuclear
3	StWOX2	Soltu.DM.06G031390	chr.06:56 294 501-56 295 977	236	26.7	8.61	细胞核Nuclear
4	StWOX3a	Soltu.DM.11G026210	chr.11:46 280 883-46 282 433	213	24.5	9.07	细胞核Nuclear
5	StWOX3b	Soltu.DM.11G026200	chr.11:46 246 409-46 248 855	200	23.1	9.28	细胞核Nuclear
6	StWOX3c	Soltu.DM.06G031020	chr.06:55 970 577-55 972 200	173	20.0	9.97	细胞核Nuclear
7	StWOX4	Soltu.DM.04G033400	chr.04:64 994 427-64 996 252	243	28.0	8.91	细胞核Nuclear
8	StWOX5	Soltu.DM.03G012800	chr.03:34 493 065-34 493 856	157	18.5	5.95	细胞核Nuclear
9	StWOX9	Soltu.DM.02G016680	chr.02:31 173 532-31 175 481	333	37.1	7.33	细胞核Nuclear
10	StWOX11	Soltu.DM.06G028250	chr.06:53 617 232-53 619 413	323	35.6	5.78	细胞核Nuclear
11	StWOX13	Soltu.DM.02G022590	chr.02:36 352 960-36 356 110	275	30.9	5.53	细胞核Nuclear

Fig. 1 Collinear analysis of potato WOX members A: The collinear analysis of WOX members in S. tuberosum. B: The collinear analysis in different WOX members between Arabidopsis thaliana, S. lycopersicum, Oryza sativa, and S. tuberosum

Fig. 2 Phylogenetic analysis of WOX proteins among S. tuberosum, A. thaliana, S. lycopersicum, N. tabacum, and O. sativa

Fig. 3 Structure of StWOX genes

Fig. 4 Distribution and sequence logos of motifs in StWOX proteins

Fig. 5 Prediction and numerical statistics of TFBS in the promoter region of StWOXs A: The prediction of TFBS. B: The statistical analysis of TFBS

Fig. 6 Relative expression patterns of StWOXs in different tissues（A）, under drought and salt stresses（B）, and under heat stress（C）

Fig. 7 Callus induction and differentiation of different potato varieties（lines）, and RT-qPCR verification of StWOX5 A: Phenotypes of different potato varieties（lines）at the stage of callus induction. B: Callus induction rates in different potato varieties（lines）. C: Phenotypes of different potato varieties（lines）at the stage of callus differentiation. D: Callus differentiation rates in different potato varieties（lines）. E: Relative expressions of StWOX5 in different potato varieties（lines）at the stage of callus differentiation. The lowercase letters indicate that the difference reached a significant level（P<0.05）

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