水稻端粒酶RNA候选序列的克隆及特征解析  

Cloning and Characteristic Analysis of Rice Telomerase RNA Candidate Sequence

作　　者：马登旭[1] 杨力媛[1] 马国兴[1] 郑洁[1] 应畅[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）》2013年第4期579-584,共6页Journal of Zhejiang Sci-Tech University(Natural Sciences)

摘　　要：端粒酶是由端粒酶反转录酶和端粒酶RNA构成,端粒酶RNA中含有端粒合成所需的模板序列。现仅获得了拟南芥的端粒酶RNA序列。为了建立快速有效的富集植物端粒酶的方法并克隆水稻的端粒酶RNA序列,研究中使用不同浓度的PEG6000选择性的沉淀端粒酶萃取液中的杂蛋白。结果表明当PEG6000浓度为4%时,端粒酶活力最高,并可以有效降低核糖体28SrRNA和18SrRNA的干扰。从端粒酶蛋白液中提取RNA,并在其3′端连接接头,在模板区设计上游引物,PCR扩增获得一条292bp的未知片段,其中含有端粒模板序列5′-AAACCCTAA-3′。将该片段及拟南芥端粒酶RNA的转录调控区进行序列比对分析发现,两者具有类似的转录调控元件。因此,初步判定292bp的目的片段为水稻端粒酶RNA的模板下游序列。Telomerase is composed of telomerase reverse transcriptase and telomerase RNA. Telomerase RNA contains the template sequence required for telomere synthesis. Only telomerase RNA sequence of arabidopsis is obtained now. To establish a rapid and effective method of hyperaecumulator telomerase and clone telomerase RNA sequence of rice, this research uses heteropolymeric protein in different concentrations of PEG6000 selective sedimentary telomerase extract. The result shows that, when PEG6000 concentration is 4%0, the activity of telomerase is the highest and it can effectively reduce the interference of ribosome 28S rRNA and 18S rRNA. This paper extracts RNA from telomerase protein fluid, connects the joint at 3r end, designs forward primer in the template area and obtains a 292 bp unknown fragment through PCR amplification. It contains telomerase template sequence 5＇-AAACCCTAA-3＇. The sequence alignment analysis on this fragment and the transcription regulation domain of Arabidopsis telomerase RNA finds that both have similar transcriptional regulatory elements. Therefore, it is preliminarily judged that the target fragment of 292 bp is template downstream sequence of rice telomerase RNA.

关 键 词：水稻 PEG6000 端粒酶RNA特征解析 

分 类 号：Q781[生物学—分子生物学]