Whole mitochondial sequencing of 3P pig: MII oocyte
Four overlapping mtDNA amplicons, size 7262 bp, 3067 bp, 3315 bp, and 3673 bp, were amplified separately using primer Sus1 (Sus1F=GTTTAACGGCCGCGGTATTC, Sus4R= TGATTACGTGCAACCCGTGA), Sus2 (Sus11F=TCACGGGTTGCACGTAATCA, Sus11R= AATGTCTCCGATGCGGTTGT), Sus3 (Sus5F=CACTCTCAGCCTAGTCCCCT, Sus5R= GAACGGGTGTTCTACGGGTT), and Sus4 (Sus3F=TTGGCCCTAGTAGCCTCCAT, Sus10R=ATGCTACCTTTGCACGGTCA), respectively. The amplicons were pooled and library preparation using Nextera DNA Flex Library Preparation Kit (Illumina). Library were 150 bp paired-end sequenced using NovaSeq 6000 SP Reagent Kit (Illumina). Image analysis was performed in real-time by the NovaSeq Control Software (NCS) v1.7.5, and Real-Time Analysis (RTA) v3.4.4 performed real-time base calling on the NovaSeq instrument computer. Then the Illumina bcl2fastq2.20.0.422 pipeline was used to generate the sequence data.