Favorprep blood genomic dna extraction mini kit

Genomic DNA was extracted from 200 µL of the blood samples using FavorPrep™ Blood Genomic DNA Extraction Mini Kit (Favorgen, Pingtung, Taiwan) according to the manufacturer's instruction and stored at -20 ºC until further analysis.
The conventional PCR (cPCR) assays were performed by universal Mycoplasma spp. primers targeting the partial sequence of the 16 S rRNA gene [51] . Subsequently, identification of CMhm, Mhf, and CMt in PCRpositive samples was performed using a panel of three species-specific primers in cPCR [22] . PCR reactions were performed in a 25 µL volume reaction mixture containing 12.5 µL of Taq DNA Polymerase 2X Mastermix (Ampliqon, Odense, Denmark), 2 µl of the template DNA, 1 µL of 10 pmol of each forward and reverse primer (synthesized by metabion international AG, Planegg, Germany), and 8.5 µL distilled deionized water. In each run, positive DNA controls that were kindly provided by Professor Dr. Roberta Iatta (University of Bari, Italy), and distilled deionized water were used as positive and negative controls. PCR amplification were run in a SimpliAmp™ thermal cycler (Thermo Fisher Scientific, Waltham, MA, USA) with conditions as described in Table 2. The PCR amplification products were documented using UV Imager (Transilluminator, Vilber Lourmat, France) after electrophoresis in a 1% agarose gel (SinaClon, Tehran, Iran) at 100 V for 60 min.