One-cell model for inhibiting Cav3.3 mRNA expression by RNA interference

• Main Authors: Xiaohong Li, Changzheng Peng, Xiaoping Wang, Xianjie Wen
• Format: Article
• Language: English
• Published: Taylor & Francis Group 2017-03-01
• Series: Biotechnology & Biotechnological Equipment
• Subjects: Dorsal root ganglion neurons; T-type calcium channel; Cav3.3; RNA interfere; cell culture
• Online Access: http://dx.doi.org/10.1080/13102818.2017.1282838

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.