[Objective] This study aimed to establish a high-efficiency and stable SSR amplification system and screen polymorphic primers in order to further compare the tri-group probability analysis method and traditional QTL ...[Objective] This study aimed to establish a high-efficiency and stable SSR amplification system and screen polymorphic primers in order to further compare the tri-group probability analysis method and traditional QTL mapping method. [Method] In this study, we preliminarily screened the 605 pairs of primers evenly distributed on the 12 chromosomes through investigating their polymorphism performance in amplification, and established an optimized SSR-PCR reaction system. [Result] A 10μL SSR-PCR reaction system suitable for rice was set up as fol ows： 2 μl of 10 × Buffer, 2.0 mmol/L Mg2＋ （final concentration）, 0.5 mmol/L dNTPs, 1.0 μmol/L primers （final concentration）, 1 μl of DNA template, 0.15 U Taq DNA polymerase. Among the SSR primers distributed over the genome, 142 pairs that were polymorphic upon the parents were screened. [Conclusion] This study lays a good foundation for sub-sequent QTL mapping studies.展开更多
Potato variety Kexin18 was used as testing materials in this research to study the influence on main components in multiplex PCR system, different primer ratios and annealing temperatures in SSR marker amplification. ...Potato variety Kexin18 was used as testing materials in this research to study the influence on main components in multiplex PCR system, different primer ratios and annealing temperatures in SSR marker amplification. Concentration and gradient experiments for four components （enzyme, MgCl2, DNA template and dNTPs） in PCR system were used in the research with the concentration of the other component remained the same; the orthogonal design L9 （34） was applied in the optimization of four sets of primers （STM0014, Pat, SSI, and UGP） in the reaction system at three levels; the temperature gradient selection was used to find out the optimum annealing temperature for the primer. The optimized multiplex PCR system of potato SSR marker with a total volume of 20 μL ： 2.5 μL 25 mmol.L-1 MgCl2, 0.6 μL 10 mmol·L-1 dNTPs, 0.8 U Taq, 80 ng DNA template was ultimately established through the comparison and analysis of test results; the ratio of four pairs of 4 mmol. L1 primers was 2 ： 1 ： 2 ： 3, and the annealing temperature was 54.7℃. The optimized reaction system could be repeated stably; and the stable and reliable amplification results were able to clearly distinguish different potato varieties. This research built the solid foundation for the further study of genetic diversity of potato germplasms and construction of DNA fingerprinting..展开更多
In this study, single-plant DNA sampling method, multi-plant leaf-mixing DNA sampling method, and single-plant DNA-mixing sampling method were adopted, to analyze the genetic variations of Meng A population and C popu...In this study, single-plant DNA sampling method, multi-plant leaf-mixing DNA sampling method, and single-plant DNA-mixing sampling method were adopted, to analyze the genetic variations of Meng A population and C population 1 using SSR markers and establish the optimal technological system for analyzing genetic diversity of maize populations. DNA samples in different treatments were amplified using 34 SSR primers which were uniformly distributed in ten chromosomes of maize. Polyacrylamidc gel electrophoresis was performed to analyze the polymorphism information content and genetic similarity coefficient of 60 individuals and compare the numbers of alleles amplified from DNA samples in different treatments. The results indicated that Meng A population and C population 1 both had relatively abundant genetic variations and the established technological system could be applied in researches of maize genetic diversity. Extracting DNA samples from mixed leaves of 12 individuals with five replications is the best sampling method, which could achieve similar results to mixed DNA samples of 12 individuals. This sampling method can be applied to analyze the genetic relationship among a large number of maize populations, which can not only reduce the workload, but also significantly improve the efficiency.展开更多
基金Supported by the Governor Special Fund for Excellent Talents for Education of Science and Technology of Guizhou Province(2012093025)~~
文摘[Objective] This study aimed to establish a high-efficiency and stable SSR amplification system and screen polymorphic primers in order to further compare the tri-group probability analysis method and traditional QTL mapping method. [Method] In this study, we preliminarily screened the 605 pairs of primers evenly distributed on the 12 chromosomes through investigating their polymorphism performance in amplification, and established an optimized SSR-PCR reaction system. [Result] A 10μL SSR-PCR reaction system suitable for rice was set up as fol ows： 2 μl of 10 × Buffer, 2.0 mmol/L Mg2＋ （final concentration）, 0.5 mmol/L dNTPs, 1.0 μmol/L primers （final concentration）, 1 μl of DNA template, 0.15 U Taq DNA polymerase. Among the SSR primers distributed over the genome, 142 pairs that were polymorphic upon the parents were screened. [Conclusion] This study lays a good foundation for sub-sequent QTL mapping studies.
基金Supported by the International Cooperation Project of Heilongjiang Science and Technology Department (WC05B08)
文摘Potato variety Kexin18 was used as testing materials in this research to study the influence on main components in multiplex PCR system, different primer ratios and annealing temperatures in SSR marker amplification. Concentration and gradient experiments for four components （enzyme, MgCl2, DNA template and dNTPs） in PCR system were used in the research with the concentration of the other component remained the same; the orthogonal design L9 （34） was applied in the optimization of four sets of primers （STM0014, Pat, SSI, and UGP） in the reaction system at three levels; the temperature gradient selection was used to find out the optimum annealing temperature for the primer. The optimized multiplex PCR system of potato SSR marker with a total volume of 20 μL ： 2.5 μL 25 mmol.L-1 MgCl2, 0.6 μL 10 mmol·L-1 dNTPs, 0.8 U Taq, 80 ng DNA template was ultimately established through the comparison and analysis of test results; the ratio of four pairs of 4 mmol. L1 primers was 2 ： 1 ： 2 ： 3, and the annealing temperature was 54.7℃. The optimized reaction system could be repeated stably; and the stable and reliable amplification results were able to clearly distinguish different potato varieties. This research built the solid foundation for the further study of genetic diversity of potato germplasms and construction of DNA fingerprinting..
基金Supported by Youth Innovation Fund from Inner Mongolia Academy of Agricultural and Animal Husbandry Sciences(2009QNJJN05)Key Project from Natural Science Foundation of Inner Mongolia Autonomous Region(20080404ZD03)
文摘In this study, single-plant DNA sampling method, multi-plant leaf-mixing DNA sampling method, and single-plant DNA-mixing sampling method were adopted, to analyze the genetic variations of Meng A population and C population 1 using SSR markers and establish the optimal technological system for analyzing genetic diversity of maize populations. DNA samples in different treatments were amplified using 34 SSR primers which were uniformly distributed in ten chromosomes of maize. Polyacrylamidc gel electrophoresis was performed to analyze the polymorphism information content and genetic similarity coefficient of 60 individuals and compare the numbers of alleles amplified from DNA samples in different treatments. The results indicated that Meng A population and C population 1 both had relatively abundant genetic variations and the established technological system could be applied in researches of maize genetic diversity. Extracting DNA samples from mixed leaves of 12 individuals with five replications is the best sampling method, which could achieve similar results to mixed DNA samples of 12 individuals. This sampling method can be applied to analyze the genetic relationship among a large number of maize populations, which can not only reduce the workload, but also significantly improve the efficiency.
