关键词: 芝麻, 抗裂蒴基因, 基因克隆, 生物信息学分析, 原核表达

Abstract: LOC105167765 belongs to KANADI (KAN) gene family,which is related with sesame capsule indehiscence.In this study,the capsule dehiscent variety Yuzhi No.11 and capsule indehiscent variety Zhengzhi InD01 were selected as experiment materials, and the SiIND1 full-length cDNAs of LOC105167765 were amplified. Bioinformatics analysis and prokaryotic expression were conducted.The results showed that the whole cDNA sequence of SiIND1 in Yuzhi No.11 was 1 320 bp and encoded 439 amino acids.The putative protein of the gene had an isoelectric point of 7.53 and a calculated molecular weight of 50.0 ku.Sequence analysis showed that the protein of SiIND1 in Yuzhi No.11 contained a SHAQKYF class MYB-like DNA binding domain similar to Arabidopsis KAN family members and shared a highly conserved GARP domain.The whole cDNA sequence of SiIND1 in Zhengzhi InD01 was 1 246 bp,which contained an ORF of 900 bp and encoded 299 amino acids.The putative protein of the gene had an isoelectric point of 8.37 and a calculated molecular weight of 32.9 ku.Compared to Yuzhi No.11,the SiIND1 gene in Zhengzhi InD01 had a deletion and generated a premature stop codon in GARP domain,which might lead to loss function of this gene.The SiIND1 gene was connected to the prokaryotic expression vector pET30a and transformed into Escherichia coli BL21 to induce expression.After the detection of SDS-PAGE,bands of target protein were identified.It was further confirmed that there was a difference in SiIND1 protein expression size between the capsule dehiscent variety and indehiscent variety,and there was a premature termination of translation in capsule indehiscent variety.

Key words: Sesame, Capsule indehiscent gene, Gene cloning, Bioinformatics analysis, Prokaryotic expression