title: Optimization of SCoT-PCR reaction system in Dactylis glomerata by orthogonal design.
abstract: The effects of 5 factors (template DNA, Mg(2+), dNTPs, Taq DNA polymerase, and primer) on the polymerase chain reaction (PCR) were investigated to optimize the start codon targeted polymor-phism (SCoT)-PCR system of Dactylis glomerata L., using an orthogo-nal design L16 (4(5)). A suitable SCoT-PCR system for D. glomerata was established; the 20 μL reaction volume contained 3.0 mM Mg(2+), 0.2 mM dNTPs, 1.0 U Taq DNA polymerase, 0.2 μM primer, 20 ng tem-plate DNA, and 2 μL 10X buffer. Each factor had a different effect on the amplification reaction, and the concentration of dNTPs had the larg-est effect on the SCoT-PCR system. We tested 10 orchardgrass samples to determine and verify the stability of the reaction system. The results showed that amplified bands from diverse materials were clear, stable, and rich in polymorphisms, indicating that the optimized system was very stable.
authors.affiliation: Department of Animal Science, Southwest University, Rongchang, Chongqing Province, China.
authors.forename: B
authors.initials: B
authors.lastname: Zeng
authors.affiliation: Department of Grassland Science, College of Animal Science and Technology College, Sichuan Agricultural University, Ya'an, Sichuan Province, China.
authors.forename: X
authors.initials: X
authors.lastname: Huang
authors.affiliation: Department of Grassland Science, College of Animal Science and Technology College, Sichuan Agricultural University, Ya'an, Sichuan Province, China huanglinkai@sicau.edu.cn.
authors.forename: L K
authors.initials: LK
authors.lastname: Huang
authors.affiliation: Chongqing Academy of Animal Sciences, Yuzhong, Chongqing Province, China.
authors.forename: J
authors.initials: J
authors.lastname: Zhang
authors.affiliation: Department of Animal Science, Southwest University, Rongchang, Chongqing Province, China.
authors.forename: H D
authors.initials: HD
authors.lastname: Yan
authors.affiliation: Department of Animal Science, Southwest University, Rongchang, Chongqing Province, China.
authors.forename: D
authors.initials: D
authors.lastname: Luo
authors.affiliation: Department of Animal Science, Southwest University, Rongchang, Chongqing Province, China.
authors.forename: H
authors.initials: H
authors.lastname: Liang
authors.affiliation: Department of Animal Science, Southwest University, Rongchang, Chongqing Province, China.
authors.forename: Y
authors.initials: Y
authors.lastname: Yuan
mesh_terms: D018387:Codon, Initiator; D017931:DNA Primers; D018744:DNA, Plant / Q000032:analysis / Q000235:genetics; D031745:Dactylis / Q000235:genetics*; D016133:Polymerase Chain Reaction / Q000379:methods*
chemical_list: D018387:Codon, Initiator; D017931:DNA Primers; D018744:DNA, Plant