One-cell model for inhibiting Cav3.3 mRNA expression by RNA interference
2017
Li, Xiaohong | Peng, Changzheng | Wang, Xiaoping | Wen, Xianjie
To gain insights into the function of Cav3.3 T-type calcium channel in dorsal root ganglion neurons, we designed a one-cell model in dorsal root ganglion neurons that specifically inhibits the expression of Cav3.3 mRNA. We designed three different shRNA sequences and then connected them to pYr-1.1 plasmid and labelled them as pYr-1.1-Cav3.3 shRNA1, pYr-1.1-Cav3.3 shRNA2 and pYr-1.1-Cav3.3 shRNA3, respectively. The correct sequence vector (pYr-1.1-Cav3.3 shRNA3) was then recombined with the pAd/PL-DEST vector into pAd-Cav3.3 shRNA the adenovirus vector. The dorsal root ganglion neurons were infected with the pAd-Cav3.3 shRNA adenovirus vector. After detecting the expression of Cav3.3 mRNA and protein, the pAd-Cav3.3 shRNA adenovirus vector was verified by digestion and sequencing. These results showed that the constructed pAd-Cav3.3 shRNA adenovirus vector could infect dorsal root ganglion cells and inhibit the expression of Cav3.3 mRNA or protein. These results established a one-cell model for further functional study of Cav3.3.
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