利用CRISPR/Cas9技术靶向敲除番茄SlMYBL基因

Targeted knockout of tomato SlMYBL gene by CRISPR/Cas9 technology

MYB类蛋白家族是所有真核生物中存在的庞大且功能丰富的一类转录因子,MYB蛋白主要参与形态建成、次生代谢等进而参与植物的非生物胁迫。文章通过比对美国国家生物技术信息中心(National Center for Biotechnology Information,NCBI)数据库,在番茄中发现了一个具有EAR转录抑制子的R1-MYB类转录因子,即SlMYBL,通过同源克隆获得SlMYBL基因的全长cDNA。SlMYBL基因在番茄的所有组织中均有表达,在果实中的表达量相对较高,其次是根和叶片。有研究表明R1-MYB类转录因子可参与植株的逆境应答,为了解SlMYBL是否响应逆境应答,文章定量分析了番茄在不同胁迫处理下SlMYBL基因的转录水平,发现其转录水平在盐胁迫下明显降低,而对甘露醇胁迫并没有明显的差异,表明SlMYBL基因可能参与了番茄植株响应盐胁迫的过程。为研究SlMYBL基因作用的分子机理,进一步构建了该基因的CRISPR表达载体,并获得SlMYBL基因的CRISPR敲除转基因番茄。该研究为发现SlMYBL基因在植物生长发育中的作用奠定了一定的理论基础。

The MYB family of proteins is a large and functionally abundant class of transcription factors present in all eukaryotes,and MYB proteins are mainly involved in morphogenesis,metabolism,and abiotic stress in plants.In this study,an R1-MYB transcription factor with EAR transcription inhibitor,SlMYBL,was found in tomato through National Center for Biotechnology Information(NCBI)database.The full-length cDNA of the SlMYBL gene was obtained by homologous cloning.SlMYBL gene is expressed in all tissues of tomato,and its expression in fruit is relatively high,followed by root and leaf.Studies have shown that R1-MYB transcription factors can participate in adversity response of plants.In order to understand whether SlMYBL responds to adversity response,the transcription level of SlMYBL gene in tomato materials under different stress treatment was quantitatively analyzed.It was found that the transcription level of SlMYBL gene decreased significantly under salt stress,but there was no significant difference in mannitol stress,suggesting that SlMYBL gene may be involved in the response of tomato plants to salt stress.In order to study the molecular mechanism of SlMYBL gene,the CRISPR expression vector of SlMYBL gene was constructed,and the CRISPR knockout transgenic tomato of SlMYBL gene was obtained.This study lays a theoretical foundation for discovering the role of SlMYBL gene in plant growth and development.