CAJ | 학술논문

目的：清除人类免疫缺陷病毒（ human immunodeficiency virus ， HIV）前病毒是治愈艾滋病的关键。构建Tre酶真核表达载体，鉴定其特异性识别HIV前病毒中所含有的loxLTR序列功能。方法经基因合成、PCR、酶切、连接等基因重组技术将Tre基因插入pcDNA3．1真核表达载体中，将EGFPpA-LoxLTR序列插入pmCherry-N1载体中，并经酶切、PCR、测序鉴定；用电穿孔法将构建的载体转染到HeLa细胞，用荧光显微镜观察荧光强弱及变化。结果经PCR、酶切、电泳、测序分析鉴定，Tre酶真核表达载体构建正确，转染HeLa细胞后，能特异性识别并剪切loxLTR序列。结论构建的Tre酶真核表达载体可在HeLa细胞中表达，并特异性识别loxLTR序列。
목적：청제인류면역결함병독（ human immunodeficiency virus ， HIV）전병독시치유애자병적관건。구건Tre매진핵표체재체，감정기특이성식별HIV전병독중소함유적loxLTR서렬공능。방법경기인합성、PCR、매절、련접등기인중조기술장Tre기인삽입pcDNA3．1진핵표체재체중，장EGFPpA-LoxLTR서렬삽입pmCherry-N1재체중，병경매절、PCR、측서감정；용전천공법장구건적재체전염도HeLa세포，용형광현미경관찰형광강약급변화。결과경PCR、매절、전영、측서분석감정，Tre매진핵표체재체구건정학，전염HeLa세포후，능특이성식별병전절loxLTR서렬。결론구건적Tre매진핵표체재체가재HeLa세포중표체，병특이성식별loxLTR서렬。
Objective Clearing HIV provirus is the key to cure AIDS .The study was to construct the Tre enzyme eukaryotic expression vector and identify its function in specific recognition of loxLTR sequence in HIV provirus . Methods Tre gene was in-serted into eukaryotic expression vector pcDNA 3.1 gene recombination manipulation by genetic recombination techniques including gene synthesis , PCR, restriction enzyme digestion and ligation .EGFPpA-LoxLTR sequence was inserted into pmCherry-N1 vector and was tested by restriction enzyme digestion , PCR and sequencing .Constructed vectors were electroporated into HeLa cells , then using fluorescence microscopy to observe fluorescence intensity changes . Results PCR, restriction enzyme digestion , electrophoresis and sequencing confirmed that Tre enzyme eukaryotic expression vector had been constructed successfully , and it could specifically recog-nize and cut loxLTR sequence after being transfected into Hela cells . Conclusion Constructed Tre enzyme eukaryotic expression vector can be expressed in Hela cells and specifically recognize loxLTR sequence , which has prepared the experimental ground for fur-ther studies of clearing HIV provirus .