CAJ | 학술논문

利用慢病毒干扰系统，对东北梅花鹿角柄骨膜干细胞（PP细胞）P21基因进行干扰。结果表明：筛选出的2条针对梅花鹿P21基因的siRNA与载体质粒PLVTHM连接成功，并与pMD2．G、pCMV-dr8.9质粒共转染到293t细胞，获得重组慢病毒；通过感染PP细胞并利用流式细胞仪进行分选，获得了纯度90％以上的感染细胞；荧光定量RT-PCR检测表明P21基因的mRNA水平大幅度下调，干扰效率达到70％。表明成功干扰了P21基因在PP细胞中的表达，获得了低表达P21的PP细胞系。
이용만병독간우계통，대동북매화록각병골막간세포（PP세포）P21기인진행간우。결과표명：사선출적2조침대매화록P21기인적siRNA여재체질립PLVTHM련접성공，병여pMD2．G、pCMV-dr8.9질립공전염도293t세포，획득중조만병독；통과감염PP세포병이용류식세포의진행분선，획득료순도90％이상적감염세포；형광정량RT-PCR검측표명P21기인적mRNA수평대폭도하조，간우효솔체도70％。표명성공간우료P21기인재PP세포중적표체，획득료저표체P21적PP세포계。
P21 gene of the pedicle periosteal cells of sika deer was interfered using RNAi in lentiviral vector system .The results showed that:(1 )Two sequences of small interfering RNAs,targeting P21 gene of sika deer,were successfully reassembled into the lentiviral plasmids(Plvthm).Positive clones were identified based on the results of both PCR and sequencing .Recombinant lentivirus was acquired by each positive plasmid co-transfecting into 293 T cells with the plasmids pMD2 .G and pCMV-dr8.9;(2)Recombinant lentivirus was successfully interfered into the PP cells,and the GFP positive cell pro-portion obtained by flow cytometry (FCM)sorting was about 90%;(3)The result of RT-PCR showed that the expression level of P21 mRNA in cells infected with recombinant lentiviral was obviously de-creased,and the interferential efficiency was about 70%.Therefore,in this study,we successfully inter-fered the expression of P21 gene in the pp cells,and obtained pp cell line with decreased expression of P21 gene,which lays foundation for revealing the regulatory mechanism of P21 underlying antler regener-ation .