南方农业学报2023，Vol.54Issue(8)：2310-2319,10.DOI:10.3969/j.issn.2095-1191.2023.08.012
甘蔗分子伴侣ShDnaJ基因克隆及在甘蔗线条花叶病毒胁迫下的表达分析
Cloning of molecular chaperone ShDnaJ gene in sugarcane and its expression analysis under stress of sugarcane streak mosaic virus
摘要
Abstract
[Objective]The purpose of the study was to clone the molecular chaperone gene ShDnaJ in sugarcane,and analyze its expression pattern under the stress of sugarcane streak mosaic virus(SCSMV),so as to provide a theoreti-cal reference for exploring the function and mechanism of action of ShDnaJ gene in sugarcane in response to SCSMV in-fection.[Method]ShDnaJ gene complete open reading frame(ORF)sequence was cloned by homologous cloning techno-logy and was analyzed by bioinformatics software.The subcellular localization of ShDnaJ protein was analyzed by fusion expression with green fluorescent protein(GFP).The expression levels of ShDnaJ gene in different tissues of sugarcane and under SCSMV stress were detected by real-time fluorescence quantitative PCR.[Result]The open reading frame(ORF)of ShDnaJ gene was1260 bp in length,and encoded 419 amino acid residues.ShDnaJ protein,with molecular weight of 45682.47 Da and theoretical isoelectric point(pI)of 8.78,was a stable hydrophilic protein,which contained a DnaJ structural domain and belonged to the member of DnaJ superfamily.ShDnaJ protein was located in the endoplasmic reticulum.The secondary structure of ShDnaJ protein consisted of 113 α-helixes,31 β-turns,78 extended strands and 197 random coils.The amino acid sequence of ShDnaJ protein had the highest amino acid sequence similarity with that of SbDnaJ in sorghum,which was 96.93%,indicating that the two proteins were most closely related.ShDnaJ gene was ex-pressed in roots,stems and leaves of sugarcane,and the expression levels in roots and stems were significantly higher than that in leaves(P<0.05,the same below).However,the expression level of ShDnaJ gene in immature stems was higher than that in mature stems.The expression level of ShDnaJ gene in mature leaves was higher than that in immature leaves.With the pass of treatment time,the relative expression of ShDnaJ gene in the SCSMV stress group and the control group(treated with phosphate buffer)showed an overall downward trend,but the expression levels of ShDnaJ genes were sig-nificantly higher than those in the control group at different time points after SCSMV inoculation.[Conclusion]ShDnaJ gene has tissue expression specificity,and the maturity of the tissue affects its expression level.The expression of this gene is up-regulated under SCSMV stress,suggesting that this gene is involved in regulating sugarcane plant resistance to SCSMV.关键词
甘蔗/分子伴侣DnaJ/基因克隆/甘蔗线条花叶病毒(SCSMV)/表达分析Key words
sugarcane/molecular chaperone DnaJ/gene cloning/sugarcane streak mosaic virus(SCSMV)/expres-sion analysis分类
农业科技引用本文复制引用
冯小艳,王文治,王俊刚,沈林波,赵婷婷,冯翠莲,张树珍..甘蔗分子伴侣ShDnaJ基因克隆及在甘蔗线条花叶病毒胁迫下的表达分析[J].南方农业学报,2023,54(8):2310-2319,10.基金项目
国家自然科学基金项目(32001604) （32001604）
海南省自然科学基金项目(320QN333) National Natural Science Foundation of China(32001604) （320QN333）
Hainan Natural Science Foundation(320QN333) （320QN333）
