水稻端粒酶RNA候选序列的鉴定  

Identification of Telomerase RNA Candidate Sequence in Rice(Oryza Sativa L.)

作　　者：朱莲莲[1] 林高强[1] 叶聪莹 黄倩[1] 刘小川[1] 

机构地区：[1]浙江理工大学生命科学学院生物工程研究所,杭州310018

出　　处：《浙江理工大学学报（自然科学版）》2016年第1期116-121,149,共7页Journal of Zhejiang Sci-Tech University(Natural Sciences)

摘　　要：端粒酶由端粒酶反转录蛋白和端粒酶RNA构成,端粒酶RNA中除模板序列保守外,其余区域的保守度极低,难以基于同源序列实施克隆。目前,植物中仅拟南芥端粒酶RNA得到了克隆。为了鉴定一条具水稻端粒酶RNA特征的候选序列,研究中进行了对模板序列单碱基T/A突变(5′-AAACCCTAA-3′)后的转基因愈伤组织端粒酶活性的比较,结果显示在缺乏dATP的端粒重复序列扩增法反应中,突变型的端粒酶活性明显高于未突变型;对体外延伸得到的端粒ssDNA进行测序分析,结果发现所测10条片段均含有突变特征序列;再对转基因植株进行端粒序列的检测,结果所测8个端粒序列中,有2个含有突变特征序列。因此,初步判断该候选序列为水稻端粒酶RNA基因。Telomerase is composed of telomerase inverse transcription and telomerase RNA.Except template sequence which is conservative in telomerase RNA,conservation degree in olther zones is very low,so it is very hard to clone based on homologous sequence.Currently,only Arabidopsis thaliana RNA has been cloned in plants.To identify a candidate telomerase with the characteristic of telomerase RNA,the activity of the transgenic callus telomerase after site-directed mutagenesis T/A of the template sequence（5-AAACCCTAA-3）was compared.The result shows the activity of mutant telomerase is significantly higher than that of non-mutant telomerase in the absence of dATP during the TRAP approach.Furthermore,a sequencing result from the ssDNA extended in vitro by the transgenic telomerase shows that all of the 10 fragments contain mutant feature sequence.Then,telomere detection of transgenic plants was conducted.The results show that among 8telomere sequences,2contain mutant feature sequence.Therefore,we can preliminarily judge the candidate sequence is telomerase RNA gene in rice.

关 键 词：水稻 端粒酶 端粒酶RNA TRAP 定点突变 

分 类 号：Q785[生物学—分子生物学]