拟南芥At2g23470基因T-DNA插入突变体的鉴定

摘要

At2g23470是拟南芥功能未知结构域DUF647蛋白家族的一个成员.为了研究At2g23470基因的功能,需要获得At2g23470功能缺失的突变体材料.根据拟南芥信息资源网站(The Arabidopsis Information Resource,TAIR)上公布的At2g23470基因可利用的T-DNA标签品系,从美国拟南芥生物资源中心(Arabidopsis Biological Resource Center,ABRC)购买了2套独立的At2g23470基因的T-DNA插入GABI-Kat株系种子,运用PCR法对这些T-DNA插入突变体进行了基因型分析和鉴定,结果获得了3个纯合的T-DNA插入位于RUS4启动子上的株系和1个T-DNA插入位于第2外显子的株系.RT-PCR分析表明,T-DNA插入位于第2外显子的株系中,At2g23470基因的表达完全缺失.该突变材料的获得为深入研究At2g23470基因的功能奠定了良好的基础.%At2g23470 is a member of the Domain of Unknown Function 647 protein family that is conserved in diverse eukaryotic organisms.To identify Arabidopsis T-DNA insertion mutants at the At2g23470 locus,we screened the ABRC collections.Two independent T-DNA lines carrying a T-DNA insertion either at the promoter or in the second exon,respectively,were genotyped using PCR-based analysis of genomic DNA.RT-PCR of homozygous T-DNA insertion mutants,using At2g23470-specifc primers,exhibited increased At2g23470 transcript levels in homozygous mutants with insertion at the promoter;whilst At2g23470 transcripts could not be detected in the homozygous mutant with T-DNA insertion at the exon,indicating that it contains a null mutation in the At2g23470 gene.These homozygous mutants have laid a foundation for investigating the function of At2g23470 gene in Arabidopsis growth and development.