新的与辐射诱导恶性转化相关全长基因Lcrp369克隆及生物信息学分析

Molecular cloning and bioinformatics analysis of radiation induced malignant transformation related novel gene Lcrp369

背景与目的:辐射诱导细胞恶性转化过程中涉及到众多基因,其中包括大量未知的新基因,克隆新的辐射诱导相关全长基因,从基因水平认识细胞恶性转化过程中生长、分化、再生和癌变的分子机理,对于研究辐射致癌的发生、发展规律具有重要意义。对此,本文旨在克隆新的与辐射致癌相关的全长基因。方法:应用SMARTRACE方法扩增T54EST片段的全长序列,测序后进行拼接及RT-PCR验证。并应用生物信息学方法对得到的新的mRNA序列进行生物信息学分析。结果:克隆了一条新的全长基因Lcrp369。经对其进行初步的生物信息学分析发现该基因是位于染色体14p22号臂上的一条功能尚不清楚的基因。其编码区域由7个外显子和6个内含子组成,mRNA全长1 038 bp,编码一244个氨基酸的蛋白,合成的蛋白位于细胞核内,具有DNA结合位点及N-糖基化位点、依赖于cAMP/cGMP的蛋白激酶磷酸化作用位点、酪蛋白激酶磷酸化位点。该蛋白二级结构推测为含有α螺旋结构为主的蛋白,其分子量为28 000,等电点为6.19。数字化组织表达谱系分析结果发现,该基因表达广泛,可在多个组织中表达及在多种肿瘤中表达。该基因全长已经登录到GENBANK上,ID号DQ525179。结论:成功地利用SMART RACE技术克隆了一条新的全长基因Lcrp369,生物信息学分析结果提示其与辐射致癌有关,其具体作用及功能尚有待进一步的实验学研究。

Background and purpose： Numerous genes are involved in the progress of cellular malignant transformation induced by radiation, lots of them are still unknown. It＇s meaningful to clone the full length of novel gene related to the radiation carcinogenesis, and also important to study the molecular mechanism of cell differentiation, regeneration and caneerizatioin from the gene level. Our aim was tn elucidate the development regularity of radiation careinogenesis. Methods： The technique of SMART RACE was adopted to clone the full length mRNA of the new EST sequence T54. The sequences were spliced after sequencing and then verified by RT-PCR. The function was preliminarily analyzed with bio-infor-maties methods. Results： Preliminary bio-informatics analysis found that the new cloned gene （Lcrp369） whose function was unknown located in the 14p22. Its coding region ine.luded 7 exons and 6 introns. The full length of mRNA was 1038 bp with a PolyA “tail” of 21 bp. The encoded protein included 244 amino acids and was located in the cell nucleus, It had DNA binding site, N-glycosylation site, eAMP/cGMP dependent protein kinase phosphorylation, site, easein kinase phos- phorylation site. The secondary, structure was presumed to be α-helix. Its mnleeular weight was 28000 and the isoeleetric point was 6.19. Digital tissue expression speetrum analysis indieated that Lcrp369 was widely expressed in many tissues and tumors. The full length of Lcrp369 had been registered into the GenBank with ID of DQ525179. Conclusions： A novel gene Lcrp369 was cloned successfully with SMART RACE. Lcrp369 played a role in radiation induced carcinogenesis and its function needs to be further studied.