烟草科技2016，Vol.49Issue(6)：1-7,7.DOI:10.16135/j.issn1002-0861.20160601
基于CRISPR/Cas9技术的烟草NtDXR基因敲除及功能分析
CRISPR/Cas9-mediated targeted mutagenesis and function analysis of DXR in Nicotiana tabacum
摘要
Abstract
In order to study the expression pattern and function of 1-deoxy-D-xylulose-5-phosphate reductoisomerase (DXR) in tobacco, NtDXR gene was cloned from tobacco cultivar Hongda (Nicotiana tabacum) and the expression level of NtDXR was also analysed in different tissues and organs. The CRISPR/Cas9 plant expression vector was constructed and the NtDXR knockout mutations were obtained. The results showed that NtDXR had the highest expression level in flower, higher in pistil and leaves. DNA insertion, deletion and displacement had been found in the target sites of NtDXR by targeted mutagenesis using the type II CRISPR/Cas9 system[Clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated 9]. Partial T0 mutant plants showed obviously photo-bleaching phenotypes, which implied that NtDXR played an important role in chlorophyll synthesis, and it suggested that the biosynthesis of plastid pigment in tobacco might be blocked by the disfunction of NtDXR.关键词
CRISPR/Cas9/烟草/5-磷酸脱氧木酮糖还原异构酶/荧光定量PCR/基因敲除Key words
CRISPR/Cas9/Tobacco/1-Deoxy-D-xylulose-5-phosphate reductoisomerase (DXR)/qRT-PCR/Gene knocking out分类
轻工纺织引用本文复制引用
聂梦云,高军平,罗培,陈晓乐,易小慧,王根洪,夏庆友..基于CRISPR/Cas9技术的烟草NtDXR基因敲除及功能分析[J].烟草科技,2016,49(6):1-7,7.基金项目
中国烟草总公司科技重大专项“不同香型烟草代谢组研究及其代谢途径构建”［110201301003B（JY-03B）］；中国烟草总公司重大专项“烟草去甲基尼古丁合成相关基因的克隆及低NNN育种材料创制”［110201401005（JY-05）］。 （）
