Scientia Agricultura Sinica ›› 2012, Vol. 45 ›› Issue (2): 359-368.doi: 10.3864/j.issn.0578-1752.2012.02.019

• ANIMAL SCIENCE·RESOURCE INSECT • Previous Articles     Next Articles

Construction of Knock-Out Vectors for Bovine MSTN

 ZHAO  Li-Hua, LIANG  Hao, YUN  Ting, LI  Rong-Feng   

  1. 1.内蒙古大学哺乳动物生殖生物学与生物技术教育部重点实验室, 呼和浩特 010021
  • Received:2011-02-26 Online:2012-01-15 Published:2011-05-13

Abstract: 【Objective】The objective of this study is to construct two replacement targeting vectors for knocking-out bovine MSTN gene. 【Method】 Primers with restricted endonucleases locus were designed and synthesized. The short arms and long arms for both vectors were isolated from the genomic DNA of cattle by polymerase chain reaction (PCR). Then, these fragments were cloned into basic targeting vectors pMCS-PLP and PⅢ to construct two replacement targeting vectors (pPLP-MSTN and PⅢ-MSTN), respectively.【Result】Two constructed gene targeting vectors were identified by PCR, T-vector ligation and DNA sequencing. The 2.8 kb short arm and the 4.0 kb long arm were detected in the targeting vector pPLP-MSTN, and the 1.3 kb short arm and the 6.8 kb long arm were detected in the targeting vector PⅢ-MSTN.【Conclusion】Two replacement targeting vectors for knocking-out bovine MSTN gene were successfully constructed. The targeting vector pPLP-MSTN is a traditional knock-out vector without a negative-selecting gene, and the targeting vector PⅢ-MSTN is a promoter trap vector with EGFP without a negative-selecting gene.

Key words: bovine, MSTN gene, knock-out vector, fetal fibroblast

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