كليدواژه :
PCR , طيور , MgCL2 , بهينه سازي , dNTPs , كلستريديوم
چكيده لاتين :
At this experiment, it was studied effects of affected factors on optimization of PCR for detection of Clostridium spp. bacteria in broiler gastrointestinal tract. After broiler slaughter, it is removed gut contents and extracted DNA. Primers are selected and their sequences and products confirmed using NCBI database. Then, different amounts of MgC12 (0.4, 0.8, 1.2 and 2.0 mM), dNTPs (0.016, 0.04, 0.08 and 0.12 mM), forward and reverse primers (0.08, 0.16, 0.24 and 0.40 pM), DNA (0.825, 2.5, 8.25, 25.0, 50.0, 75.0 and 100.0 ng) and different PCR cycles (20, 25, 30 and 35 cycles) were examined and the best option selected for each step. PCR reactions were conduct in 25 ?1 volume and their products loaded, electrophorzed and imaged. From obtained results, it is shown that 1.2 mM MgC12 is the best concentration for optimization of these PCRs. Furthermore, the best DNA concentration for optimization of these PCRs was 25.0 ng. When the best concentration dNTPs for optimization of these PCRs is 0.08 mM. Furthermore, 30 cycles was the best number of cycles for optimization of these PCRs. Furthermore, the best concentration forward and reverse primers for optimization of these PCRs were 0.24 pM. Therefore, PCRs were developed successfully and efficiently for detection of Clostridium spp. bacteria in broiler gastrointestinal tract. Vet. J.of Islamic. Azad. Univ., Garmsar Branch. 3,3:153-162,2007.
