过表达青绿苔草滞绿基因CbSGR加速拟南芥叶绿素降解和衰老  

Overexpression of CbSGR,A Stay-green Gene from Carex breviculmis Accelerated Chlorophyll Degradation and Senescence in Arabidopsis thaliana

作　　者：刘凌云 檀鹏辉 姜红岩 常智慧[2] 滕珂 LIU Ling-yun;TAN Peng-hui;JIANG Hong-yan;CHANG Zhi-hui;TENG Ke(Beijing Research and Development Center for Grass and Environment,Beijing Academy of Agriculture and Forestry Sciences,Beijing 100097,China;College of Grassland Science,Beijing Forestry University,Beijing 100083,China)

机构地区：[1]北京市农林科学院北京草业与环境研究发展中心,北京100097 [2]北京林业大学草业与草原学院,北京100083

出　　处：《中国草地学报》2020年第4期15-24,共10页Chinese Journal of Grassland

基　　金：国家自然科学青年基金项目(31901397);北京市自然科学青年基金项目(6204039);北京市科技创新计划(171100007217001)共同资助。

摘　　要：根据全长转录组数据,克隆得到青绿苔草滞绿基因CbSGR(GenBank登录号:MN746439),其完整编码区全长为771bp,编码256个氨基酸。NCBI Nr数据库Blast结果显示,CbSGR属于STAY-GREEN超家族,系统进化分析表明CbSGR与油棕和海枣SGR的相似度最高。为研究CbSGR的转录激活特性,构建pGBKT7-CbSGR载体并转化Y2HGold酵母感受态细胞,其在三缺培养基(SD-Trp-His-Ade)上不能生长,表明CbSGR不具有转录激活活性。利用本生烟草瞬时表达方法观察亚细胞定位情况,结果表明CbSGR定位于叶绿体。利用花序侵染法转化拟南芥,过表达株系表现出黄化,叶绿素含量显著低于对照且净光合速率明显小于对照。荧光定量分析结果表明,过表达CbSGR转基因株系中光合效率相关的AtCAB1、AtPSAF、AtRCA基因表达量均显著低于对照,而衰老相关的AtNYC1、AtNOL、AtSEN、AtSAG12和AtSAG14基因表达量均显著高于对照,表明CbSGR的过表达降低了光合效率并加速了植物衰老。CbSGR是一个调节植物叶绿素降解和衰老的关键基因,为后续通过基因编辑等措施改良青绿苔草绿期奠定了基础。Based on the full-length transcriptome data,a stay-green gene CbSGR(GenBank No.MN746439)from Carex breviculmis was isolated.Its open reading frame is 771bp in length,encoding 256 amino acids.NCBI Nr database Blast results showed that the CbSGR is a typical member of STAY-GREEN superfamily.Phylogenetic analysis revealed that CbSGR had the highest similarity with Elaeis guineensis or Phoenix dactylifera.To reveal the transcriptional activation characteristics of CbSGR,a pGBKT7-CbSGR vector was constructed and transformed into Y2HGold yeast competent cells,which could not grow on the SD-Trp-His-Ade medium,indicating that CbSGR did not have transcriptional activation activity.The transient expression method of Nicotiana benthamiana was used to abserve the subcellular localization,and the results showed that CbSGR was localized in the chloroplasts.Arabidopsis thaliana was transformed by inflorescence infestation,phenotype observation results showed that the CbSGR-overexpressing lines showed accelerated yellowing tendency.The chlorophyll content and net photosynthetic rate(Pn)of transgenic lines was lower than the control.qRT-PCR analysis revealed that the expression levels of AtCAB 1,AtPSAF and AtRCA genes related to photosynthetic efficiency in transgenic lines were significantly lower than those in the control,while the expression levels of AtNYC1,AtNOL,AtSEN,AtSAG12 and AtSAG 14 genes related to senescence process in transgenic lines were significantly higher than those in the control.This indicated that overexpression of GbSGR reduced photosynthetic efficiency and accelerated plant senescence.In conclusion,CbSGR was a key gene regulating chlorophyll degradation and senescence in plants,which laid a foundation for improving the green period of C.breviculmis through gene editing method and other measures.

关 键 词：青绿苔草 滞绿基因 转基因 叶绿素降解 衰老 

分 类 号：S543.9[农业科学—作物学]