PMC:7151644 / 9044-9898 JSONTXT

{"target":"http://pubannotation.org/docs/sourcedb/PMC/sourceid/7151644","sourcedb":"PMC","sourceid":"7151644","source_url":"https://www.ncbi.nlm.nih.gov/pmc/7151644","text":"2.3 PCR and sequencing\nTotal RNA was reverse transcribed using a one-step RT-PCR kit (TaKaRa). The viral RNA in ticks was detected by nested PCR targeting the conserved regions of the RNA-dependent RNA polymerase (RdRp) gene of both the pestivirus and coltivirus. To recover complete viral genomes, primers were designed based on the assembled pestivirus and coltivirus contigs obtained by meta-transcriptomics (Supplementary Table S3). The genome termini were determined by 5ʹ/3ʹ RACE kits (TaKaRa).\nThe QIAquick Gel Extraction kit (Qiagen) was used to purify the PCR products before sequencing. Purified DNA \u003c700 bp in length was sequenced directly, while those larger than 700 bp were first cloned into a pMD18-T vector (TaKaRa), and then transformed into JM109-143 competent cells. For each sample, at least three clones were selected for sequencing.","divisions":[{"label":"p","span":{"begin":23,"end":500}},{"label":"title","span":{"begin":0,"end":22}}],"tracks":[]}