PMC:3091644 / 34767-35802 JSONTXT

{"target":"https://pubannotation.org/docs/sourcedb/PMC/sourceid/3091644","sourcedb":"PMC","sourceid":"3091644","source_url":"https://www.ncbi.nlm.nih.gov/pmc/3091644","text":"Amplification of mtDNA fragments\nThe total length of the mt genome was amplified in 9 overlapping fragments using EX TaqTM polymerases with 3'-5' exonuclease proofreading activity (Takara Biotechnology Co. Ltd, Dalian, China) using total genomic DNA purified from a single cyst or worm as the template. The overlapping fragments of T. multiceps, T. hydatigena and T. pisiformis mtDNAs were amplified using nine pairs of oligonucleotide primers (Additional file 5), designed according to the conserved regions from published complete mtDNA sequences of taeniid cestodes. All PCR reactions comprised ~20-40 ng of the genomic DNA in a 50 μl reaction containing 1.5 U Taq polymerase, 10 mM Tris-HCl pH9, 50 mM KCl, 2 mM MgCl2, 200 μM of each dNTP. PCR amplifications each proceeded with 35 cycles of 94°C for 1 min, 52°C for 45 s, 72°C for 2 to 4 min depending on product length. The amplicons were then cloned into the pGEM-T Easy vector (Promega Co., Winsconsin, USA). At least 3 clones from each amplicon were double-stranded sequenced.","divisions":[{"label":"title","span":{"begin":0,"end":32}}],"tracks":[]}