醫(yī)學(xué)論文范文:假型逆轉(zhuǎn)錄病毒載體高效介導(dǎo)兔平滑肌細胞基因轉(zhuǎn)染
【摘要】 目的 構(gòu)建假型逆轉(zhuǎn)錄病毒載體MuLV/VSVG,并用于轉(zhuǎn)染兔平滑肌細胞,為兔平滑肌細胞基因轉(zhuǎn)染尋找一種高效的載體。方法 構(gòu)建含有報道基因lacZ的假型逆轉(zhuǎn)錄病毒載體MuLV/VSVG,測定滴度,并轉(zhuǎn)染兔平滑肌細胞,觀察其轉(zhuǎn)導(dǎo)效率。并與MuLV的轉(zhuǎn)導(dǎo)效率進行比較。結(jié)果 構(gòu)建的MuLV/VSVG載體,病毒滴度為6~7.8×106CFU,轉(zhuǎn)染兔平滑肌細胞的效率是(92±12)%。而MuLV的轉(zhuǎn)導(dǎo)效率為(24±5)%。結(jié)論 成功構(gòu)建了假型逆轉(zhuǎn)錄病毒載體MuLV/VSVG載體,該載體作為一種高效的載體可用于兔平滑肌細胞基因轉(zhuǎn)染。
【關(guān)鍵詞】 假型逆轉(zhuǎn)錄病毒載體;平滑肌細胞;轉(zhuǎn)染
Highefficiency gene transfer into rabbit smooth muscle cellsby pseudotyped retroviral vector
PEI Fei, HE Rui, LI Junyan, YU Hong
Department of Cardiothoracic Surgery, the Second Affiliated Hospital,Medical School of Xian Jiaotong University, Xian 710004, China
ABSTRACT: Objective To construct pseudotyped retroviral vector MuLV/VSVG and transfer it into rabbit smooth muscle cells (SMC) in order to provide a highefficiency vector for SMC gene transfer. Methods We constructed pseudotyped retroviral vector MuLV/VSVG containing the previously reported gene lacZ, determined the titer, and determined the efficiency of gene transfer into SMC mediated by pseudotyped retroviral vector MuLV/VSVG. Finally the transfer efficiency was compared with that by MuLV. Results MuLV/VSVG vector was constructed. The titer of the vector was 6-7.8×106CFU, the transfer efficiency was (92±12)% by using MuLV/VSVG vector and (24±5)% by MuLV vector. Conclusion Pseudotyped retroviral vector MuLV/VSVG which was constructed successfully is a kind of highefficiency gene transfer vector in smooth muscle cells醫(yī).學(xué)全.在.線網(wǎng)站bhskgw.cn.
KEY WORDS: pseudotyped retroviral vector; smooth muscle cell; gene transfer
基因治療是指將具有防治潛能的外源基因,通過相應(yīng)的載體轉(zhuǎn)移到患者的有關(guān)組織器官和細胞內(nèi),獲得適當(dāng)?shù)谋磉_達到防治和減輕疾病的目的。其中目的基因載體的選擇是基因治療的關(guān)鍵之一。逆轉(zhuǎn)錄病毒是較常用的一種病毒載體。鼠白血病病毒MuLV衍生的逆轉(zhuǎn)錄病毒載體被廣泛應(yīng)用于將基因轉(zhuǎn)染到哺乳細胞中并長期表達[1]。但該載體穩(wěn)定性差,產(chǎn)生的病毒滴度低,轉(zhuǎn)染到人類細胞效率低,并且只能轉(zhuǎn)染分裂期細胞。VSVG,皰疹性口炎蛋白VSV的包膜蛋白,是一種單鏈蛋白,可以完全替代MuLV的env蛋白,產(chǎn)生基于MuLV的高轉(zhuǎn)染率的病毒載體MuLV/VSVG,已經(jīng)被用來構(gòu)建假型MuLV/VSVG以提高MuLV載體的穩(wěn)定性及轉(zhuǎn)染效率[2]。本研究用兔平滑肌細胞作為靶細胞,觀察MuLV/VSVG的轉(zhuǎn)導(dǎo)效率,并與MuLV的轉(zhuǎn)導(dǎo)效率進行比較,探討MuLV/VSVG用于平滑肌細胞基因轉(zhuǎn)染的可行性。
1 材料與方法