Padtrack cmv shuttle vector

The process for adenovirus generation and proliferation was performed as previously described by our (Shi et al., 2017) (link). Briefly, the cDNA sequence of dairy goat PTEN (GenBank accession no. MK158074.1; https: / / www .ncbi .nlm .nih .gov/ nuccore/ MK158074 .1) was subcloned into the pAdTrack-CMV shuttle vector between the KpnI and HindIII (New England BioLabs Inc.) restriction sites to generate the pAdTrack-CMV-PTEN vector. Then, the vector was inserted into an adenoviral vector (pAdEasy-1) in BJ5183 bacterial cells to generate adenoviral plasmids. The linearized adenoviral plasmid by PacI (New England BioLabs Inc.) was transfected into 293A cells to generate the adenovirus pAd-PTEN. The adenovirus containing green fluorescent protein (Ad-GFP) was used as a negative control.

The procedure for adenovirus generation was previously described (Shi et al., 2013) . Briefly, the cDNA sequence of dairy goat THRSP (Gene ID: KU051565) was subcloned into the pAdTrack-CMV shuttle vector between the BglII and SalI (New England BioLabs Inc., Ipswich, MA) restriction sites to generate the pAdTrack-CMV-THRSP vector. Then, this vector was linearized by PmeI (New England BioLabs Inc.) and inserted into an adenoviral vector (pAdEasy-1) to generate adenoviral plasmids in BJ5183 cells. The linearized adenoviral plasmid by PacI (New England BioLabs Inc.) was transfected into 293A cells to generate the adenovirus with THRSP (Ad-THRSP) using a commercial system (AdEasy, Stratagene, La Jolla, CA). The adenovirus containing green fluorescent protein (Ad-GFP) was used as a negative control and was a gift from Zhijie Chang (Tsinghua University, Beijing, China).