miR-21靶向猪TSC1和PPP3CA基因的初步鉴定

doi:10.11843/j.issn.0366-6964.2013.06.023

畜牧兽医学报 ›› 2013, Vol. 44 ›› Issue (6): 985-992.doi: 10.11843/j.issn.0366-6964.2013.06.023

• 研究简报 • 上一篇

miR-21靶向猪TSC1和PPP3CA基因的初步鉴定

彭兴^1,2 , 谢炳坤³，唐中林^2*，周荣²，敖红²，黄生强^1*，李奎²

（1. 湖南农业大学动物科学技术学院，长沙 410128； 2. 中国农业科学院北京畜牧兽医研究所，北京 100193; 3. 广西畜牧研究所，南宁 530001）

收稿日期:2012-12-31 出版日期:2013-06-23 发布日期:2013-06-23
通讯作者: 唐中林，E-mail：zhonglinqy_99@sina.com；黄生强，E-mail：hsq07@126.com
作者简介:彭兴（1986 -），男，瑶族，湖南怀化人，硕士，主要从事动物遗传育种与繁殖方面的研究，E-mail：px_bio@126.com
基金资助:
转基因生物新品种培育重大专项（2011ZX08006-002）；广西家畜遗传改良重点实验室培育基地开放课题（桂牧研科2012-27）；国家自然科学基金（31171192）

Preliminary Identification of miR-21 Targeted TSC1 and PPP3CA Genes of Pigs

PENG Xing¹^，2, XIE Bing-kun³, TANG Zhong-lin^2*, ZHOU Rong², AO Hong², HUANG Sheng-qiang^1*, LI Kui²

(1. College of Animal Science and Technology, Hunan Agricultural University, Changsha 410128, China; 2. Institute of Animal Science, Chinese Academy of Agricultural Sciences, Beijing 100193, China; 3. Guangxi Institute of Animal Science, Nanning 530001, China)

Received:2012-12-31 Online:2013-06-23 Published:2013-06-23

摘要/Abstract

摘要：

旨在初步鉴定miR-21靶向肌肉发育相关的基因。通过靶标预测网站发现结节性硬化症蛋白1基因(TSC1)和蛋白磷酸酶3催化亚基-α-亚型基因（PPP3CA）基因的3′非编码区（3′UTR）均具有与miR-21结合的潜在靶位点；以通城猪为试验材料，扩增其TSC1和PPP3CA 基因的3′ UTR，经Not Ⅰ和Xho Ⅰ双酶切后回收目的片段，连接到双荧光素酶载体psiCHECK中；将重组载体pCHE-TSC1-3′UTR和pCHE-PPP3CA-3′UTR分别转染到猪髋动脉内皮细胞（PIEC）和猪肾细胞（PK15）中，检测其荧光素酶活性。结果显示，当2个靶标载体分别与miR-21共转时，miR-21过表达组在PIEC细胞和PK15细胞中的荧光素酶活性均显著低于对照组（P<0.05）。研究结果表明，猪miR-21可能靶向调控TSC1和PPP3CA的表达，本研究为深入研究miR-21的生物学功能和作用机制等提供了基础。

Abstract:

The study was to investigate the regulation mechanism of miR-21 in skeletal muscle development. Using computational target prediction programs, we identified TSC1 and PPP3CA genes as the potential targets of miR-21 which can specify bind to their 3′ UTR regions. The 3′UTR regions of these two genes were amplified by PCR from Tongcheng pigs genomic DNA. The PCR productions were digested by double digestion with Not I and Xho I. We got the pCHE-TSC1-3′UTR and pCHE-PPP3CA-3′UTR vectors after digested products were ligated into the vector psiCHECK with the dual-luciferase gene. These vectors were transfected into porcine iliac endothelial cell line (PIEC) and porcine kidney epithelial cell line (PK15), and then the luciferase activity was tested by a dual-luciferase reporter assay. The results showed that the luciferase activity of experimental group was significantly lower than that of the control（P<0.05）both in PIEC and PK15. This study demonstrated that TSC1 and PPP3CA were the possible target genes of miR-21 and provides basic for further research the function of miR-21.

中图分类号:

S828.2

彭兴,谢炳坤，唐中林，周荣，敖红，黄生强，李奎. miR-21靶向猪TSC1和PPP3CA基因的初步鉴定[J]. 畜牧兽医学报, 2013, 44(6): 985-992.

PENG Xing, XIE Bing-kun, TANG Zhong-lin, ZHOU Rong, AO Hong, HUANG Sheng-qiang, LI Kui. Preliminary Identification of miR-21 Targeted TSC1 and PPP3CA Genes of Pigs[J]. ACTA VETERINARIA ET ZOOTECHNICA SINICA, 2013, 44(6): 985-992.

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miR-21靶向猪TSC1和PPP3CA基因的初步鉴定

Preliminary Identification of miR-21 Targeted TSC1 and PPP3CA Genes of Pigs

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