Construction and titration of rat CGRP gene recombinant lentivirus

VernacularTitle:大鼠CGRP基因重组慢病毒载体的构建及滴度测定
Author: Panke CHEN ; Bei SHI ; Guanxue XU ; Zhijiang LIU ; Dongmei WANG
Publication Type:Journal Article
Keywords: calcitonin gene-related peptide; lentivirus; DNA,recombinant
From: Chongqing Medicine 2013;(34):4157-4159
CountryChina
Language:Chinese
Abstract: Objective To construct lentiviral vector carrying rat′s calcitonin gene-related peptide(CGRP) gene for the following-up study on the function of CGRP .Methods CGRP gene segment was subcloned into shuttle plasmid ,become Puc57-CGRP .The pLenO-DCE-CGRP expression vector was be constructed by double digests .The pLenO-DCE-CGRP and 4 auxiliary packaging plas-mids were co-transfected into 293T cells .Cells were cultured for 48 hours .The supernant was collected and concentrated ,and then the viral titers were tested by multiple proportions dilution method and flow cytometer .The expression levels of CGRP were detec-ted in CGRP-modified 293T cells by Real-time PCR .Results The results of digestion and sequencing show that the pLenO-DCE-CGRP vector was constructed successfully .The titer of the lentiviral particles was 5 .1 × 108 TU /mL .Conclusion The high-titer lentvirus vector containing CGRP gene is constructed successfully ,which lay a foundation for transfecting mesenchymal stem cell (MSC) and studying the function of CGRP .