Construction of adenovirus vectors encoding the lumican gene by gateway recombinant cloning technology

• 作者: Bing Qi Gui-Fang Wang Guo-Cheng Yu Jing-Xiang Zhong Lei-Lei Tu Lian Liu
• 来源期刊: 国际眼科杂志：英文版
• 学科: 医学
• 年，卷（期）: 2016,（9）
• 页码范围: 1271-1275
• 页数: 5页
• 分类号: R77

摘要

AIM:To construct adenovirus vectors of lumican gene by gateway recombinant cloning technology to further understand the role of lumican gene in myopia.METHODS:Gateway recombinant cloning technology was used to construct adenovirus vectors.The wild-type(wt) and mutant(mut) forms of the lumican gene were synthesized and amplified by polymerase chain reaction(PCR).The lumican cDNA fragments were purified and ligated into the adenovirus shuttle vector pDownmultiple cloning site(MCS)-/internal ribozyme entry site(IRES)/enhanced green fluorescent protein(EGFP).Then the desired DNA fragments were integrated into the destination vector pAV.Des1 d yielding the final expression constructs pAV.Ex1d-CMV>wt-lumican/IRES/EGFP and pAV.Ex1d-cytomegalovirus(CMV)>mutlumican/IRES/EGFP,respectively.RESULTS:The adenovirus plasmids pAV.Ex1d-CMV>wt-lumican/IRES/EGFP and pAV.Ex1d-CMV>mutlumican/IRES/EGFP were successfully constructed by gateway recombinant cloning technology.Positive clones identified by PCR and sequencing were selected and packaged into recombinant adenovirus in HEK293 cells.CONCLUSION:We construct adenovirus vectors containing the lumican gene by gateway recombinant cloning technology,which provides a basis for investigating the role of lumican gene in the pathogenesis of high myopia.