摘要
为明确柯巴基焦磷酸合酶基因(CPS2)的功能和调控机制,利用CRISPR/Cas9技术对高香气烤烟品系8306中的CPS2基因进行定向编辑,得到了7株靶位点单碱基A/T/C插入的转化植株。利用q-PCR技术分析转化植株二萜类关键基因的表达,并利用GC/MS分析叶面分泌物成分及香叶基香叶基焦磷酸(GGPP)含量(质量分数)。结果表明:①T0代转化植株CPS2及ABS表达量显著降低,冷杉醇和赖百当烯二醇含量也显著降低。②DXR表达量均显著升高,GGPP含量也大幅提高。③CYC-1和CYP71D16表达量差异较大,西柏三烯二醇出现小幅降低。④T0代转化植株的株高、茎围、叶间距、最大叶长、最大叶宽均增加。因此,敲除CPS2能够抑制赖百当类化合物合成,有助于上游途径中萜类合成前体GGPP的积累,可能抑制西柏三烯二醇积累,同时使植株表型性状发生变化。
In order to explore the function and regulatory mechanism of CPS2 gene,CRISPR/Cas9 system was used to edit the CPS2 gene in high aroma flue-cured tobacco line 8306.A total of 7 transformed plants with single base A/T/C inserted at the target sites were obtained.The expression of key diterpenoid genes in transformed plants were analyzed by q-PCR.The components in leaf surface exudates and the content(mass fraction)of GGPP were analyzed by GC-MS.The results showed that:1)The expression levels of CPS2 and ABS decreased significantly in T0 generation transformed plants,and their contents of cis-abienol and labdenediol decreased significantly.2)The expression level of DXR significantly increased,and the GGPP content also raised greatly.3)The expression of CYC-1 and CYP71D16 differed significantly,while the content of cembratrien-diol decreased slightly.4)The plant height,stalk circumference,leaf spacing,maximum leaf length and maximum leaf width of T0 generation transformed plants increased.Therefore,the knockout of CPS2 can inhibit the synthesis of labdanoids compounds and contribute to the accumulation of terpenoid synthetic precursor GGPP in the upstream pathway,which might inhibit the accumulation of cembtriene-diol and associate changes in the phenotypic traits.
作者
张思琦
何佳
贺凌霄
薛刚
孙聚涛
李晓辉
段杜薇
徐世晓
ZHANG Siqi;HE Jia;HE Lingxiao;XUE Gang;SUN Jutao;LI Xiaohui;DUAN Duwei;XU Shixiao(College of Tobacco Science,Henan Agricultural University,Zhengzhou 450002,China;China Tobacco Henan Industrial Co.,Ltd.,Zhengzhou 450000,China)
出处
《烟草科技》
EI
CAS
CSCD
北大核心
2020年第5期17-25,共9页
Tobacco Science & Technology
基金
国家烟草专卖局科技项目“河南豫浓香型优质烤烟品种筛选与配套技术的应用研究”(No201841000270035)。
