Rapid competent cell preps kit

For the epidemiological survey of PSV, a total of 260 clinical samples, including feces, fecal swabs, and intestine samples, were collected between 2020 and 2022 from sows and suckling, post-weaning, and fattening pigs that were asymptomatic or with clinical diarrhea (25 with clinical diarrhea and 5 asymptomatic) from 30 farms in Fujian Province. A description of the studied pig population and the number of collected samples for each farm is shown in Supplementary Table S1. All samples were detected for PSV using real-time PCR. The PSV2020 strain was preserved by the Institute of Animal Husbandry and Veterinary Medicine, Fujian Academy of Agricultural Sciences. M-MuLV First Strand cDNA Synthesis Kit (Code No: B532435), Rapid Competent Cell Preps Kit (One-step, Code No: B529307), SanPrep Column Plasmid Mini-Preps Kit (Code No: B518191), SanPrep Spin Column & Collection Kit (Code No: B515103), and Taq Plus DNA polymerase, 2 × Taqman Fast qPCR Mix (High Rox) (Code No: B639276, BBI) were bought from Sangon Biotech (Shanghai, China). RNA extraction kit (Code No: 9767), PrimeScriptTM RT Master Mix Reverse Transcription Kit (Code No: RR036A), TaKaRa LA Taq^® (Code No: RR02MQ), and pMD 18-T Vector (Code No: 6011) were purchased from TaKaRa Bio Inc. (Daliang, China).

Total DNA of hippocampal tissues was extracted, respectively, by DNA extraction kit (Sangon Biotech, Shanghai, China). The genomic DNA were modified by bisulfite sodium by BisulFlash DNA Modification Kit (Epigentek, Farmingdale, NY, USA), and PCR amplification. PCR electrophoresis and recovery was by SanPrep Column Plasmid Mini-Preps Kit (Sangon Biotech, Shanghai, China). PCR-purified products are connected to pUC18-T carrier. Competent cells were prepared by Rapid Competent Cell Preps Kit (Sangon Biotech, Shanghai, China). Each transformed clone was placed onto LB plates which contained ampicillin/IPTG/X-gal exactly as described in the manufacturer´s instructions and subjected to overnight growth at 37 °C. Plasmid DNA extraction was by SanPrep Column Endotoxin-Free Plasmid Mini-Preps Kit (Sangon Biotech, Shanghai, China). Sequencing was completed by Sangon Biological Engineering Co., Ltd. (Shanghai, China). The specific gene primers were synthesized by the Sangon Biological Engineering Co., Ltd. (Shanghai, China). Moreover, the production lengths of the primer sequences used in this experiment are shown in Table 2.