植物学报2023，Vol.58Issue(6)：905-916,12.DOI:10.11983/CBB22250
湖北地黄CRISPR/Cas9基因编辑体系的建立
Establishment of CRISPR/Cas9 Gene Editing System in Rehmannia henryi
摘要
Abstract
Rehmannia henryi is an important plant with great medicinal value,but no research of CRISPR/Cas9 has been done on this species.To establish the gene editing system of R.henryi,the gene encoding phytoene desaturase(PDS)in carotenoid biosynthesis was screened,and the CRISPR/Cas9 vector of RhPDS1 was constructed and transformed into R.henryi genome by Agrobacterium-mediated transformation method.The transcript of RhPDS1 with a 1 764 bp open reading frame(ORF)of RhPDS1 was obtained,the deduced amino acid sequence of RhPDS1 has the typical structural domains of phytoene desaturase.RhPDS1 showed higher expression levels in bud,flower and new leaf.Using CRISPR/Cas9 method,three regenerated shoots with albino phenotype were finally obtained,the differentiation rate of albino shoot was 3.7％.Sequencing analysis revealed that the three albino shoots belong to 2 editing events,in which deletion of 1 bp or(and)5 bp occurred,respectively,which caused frame shift mutations.The contents of chlorophylls and carotenoids were significantly decreased in the albino mutants as compared to wild type,and the expression levels of RhPDS1 were also decreased in the albino mutants.Taken together,the RhPDS1 gene was cloned and knocked out by using CRISPR/Cas9 method,which laid down the foundations for functional genomics studies and de novo domestication of R.henryi.关键词
湖北地黄/RhPDS1/CRISPR/Cas9/基因敲除Key words
Rehmannia henryi/RhPDS1/CRISPR/Cas9/gene knock out引用本文复制引用
苗春妍,李铭铭,左鑫,丁宁,杜家方,李娟,张重义,王丰青..湖北地黄CRISPR/Cas9基因编辑体系的建立[J].植物学报,2023,58(6):905-916,12.基金项目
国家自然科学基金(No.81473299)和河南省高等学校重点科研项目(No.22A360009) （No.81473299）
