猪嵌合RNABCL2L2CR克隆与功能初步分析

Cloning and functional analysis of porcine chimeric RNA BCL2L2CR

研究前期鉴定一个嵌合子标签——BCL2L2CR,含有B细胞淋巴瘤/白血病-2样蛋白2(B-cell CLL/lymphoma 2 like protein 2,BCL2L2)重复序列,为分析BCL2L2CR作用,利用RT-PCR方法克隆其cDNA序列,采用流式细胞术、过表达、CCK-8和细胞划痕试验等分子生物学技术分析其对细胞凋亡、增殖和迁移的影响。发现BCL2L2CR是BCL2L2外显子3部分序列发生顺式重复形成,前141个氨基酸与BCL2L2完全相同,含有BCL2同源结构域4、3和1,缺少同源结构域2;BCL2L2CR促进PK-15和3T3L细胞增殖和迁移。研究结果确定嵌合RNA BCL2L2CR具有一定功能,将为深入揭示BCL2L2CR功能提供基础,有助于进一步认识嵌合RNAs及其在正常生理活动中的作用。

In this study,a chimeric tag,BCL2L2CR,containing tandem sequence of B-cell lymphoma/leukemia-2 like protein 2(BCL2L2)was identified.In order to analyze the role of BCL2L2CR,the cDNA sequence was cloned by RT-PCR in this study.The effects of BCL2L2CR on apoptosis,proliferation and migration were analyzed by flow cytometry,overexpression,CCK-8 and cell scratch test.The obtained BCL2L2CR cDNA was 998 bp long,containing 171 bp 5′UTR,573 bp open reading frame and 254 bp 3′UTR,and it was expected to encode 190 amino acid residues of protein polypeptide chain.It was found that BCL2L2CR was formed by cis-repeat in exon 3 of BCL2L2.The first 141 amino acids of BCL2L2CR were identical to BCL2L2.BCL2L2CR contained homologous domains 4,3 and 1,and lacked homologous domain 2.BCL2L2CR could promote the proliferation and migration of PK-15 and 3T3L cells.The results confirmed that chimeric RNA BCL2L2CR had a certain function,which would provide a basis for further revealing the function of BCL2L2CR,and help to further understand the chimeric RNA and its role in normal physiological activities.