Rice blast is one of the important diseases in major rice producing areas of China. The main blast resistance genes Pi-ta and Pi-b showed broad-spectrum and durable resistance to rice blast in many rice growing areas ...Rice blast is one of the important diseases in major rice producing areas of China. The main blast resistance genes Pi-ta and Pi-b showed broad-spectrum and durable resistance to rice blast in many rice growing areas of China, which have been widely utilized in rice breeding and commercial production. In this study, on the basis of detection and verification of the genotypes of 22 rice varieties har- boring known blast resistance genes （Pi-ta and Pi-b） and blast susceptibility genes （pi-ta and pi-b）, two multiple PCR systems for these genes were established by us- ing the functional markers of blast resistance genes Pi-ta and Pi-b as well as blast susceptibility genes pi-ta and pi-b, respectively. Specifically, multiple PCR system I could simultaneously detect blast resistance genes Pi-ta and Pi-b, while system II could detect simultaneously blast susceptibility genes pi-ta and pi-b. In addition, the genotypes of 336 high generation breeding materials were detected with these two multiple PCR systems. The results were highly consistent with those of conventional single mark detection, indicating that these two multiplex PCR systems were stable, reliable and time-saving. The established multiplex PCR systems may serve as a rapid and efficient method to identify and screen rice germplasm resources and can be applied in marker-assisted selection to polymerize multiple genes for blast resis- tance in rice breeding.展开更多
[Objective] This research aimed to search a best method for extracting the genomic DNA of Cymbidium ensifolium and establish the optimized ISSR-PCR reaction system.[Method] Genomic DNA was extracted from C.ensifolium ...[Objective] This research aimed to search a best method for extracting the genomic DNA of Cymbidium ensifolium and establish the optimized ISSR-PCR reaction system.[Method] Genomic DNA was extracted from C.ensifolium leaves by modified CTAB method.ISSR-PCR reaction system for C.ensifolium was optimized.[Result] High-quality genomic DNA was obtained from C.ensifolium.The 25 μl optimized ISSR-PCR reaction system for C.ensifolium contained 2.5 μl 10× PCR buffer,2.5 mmol/L MgCl2,240 ng template DNA,160 μmol/L dNTPs,1.25 U Taq DNA polymerase,0.4 μmol/L primer and 15.78 μl ddH2O.The optimal PCR procedures were：94 ℃ pre-denaturation for 5 min and then 40 cycles,94 ℃ denaturation for 30 s,50-60 ℃ annealing for 30 s （annealing temperature according to different primers）,72 ℃ extension for 50 s and a 72 ℃ extension for 7 min.[Conclusion] An optimized ISSR-PCR reaction system for C.ensifolium was established,which provides a basis for further study on genetic diversity of C.ensifolium by using ISSR molecular marker technique.展开更多
Two silkworm strains viz, B20 A (high cocoon shell ratio) and C.Nichi (low cocoon shell ratio) were sib mated for 10 generations to determine the homozygosis. Both bulked segregant analysis(BSA) and near isogenic line...Two silkworm strains viz, B20 A (high cocoon shell ratio) and C.Nichi (low cocoon shell ratio) were sib mated for 10 generations to determine the homozygosis. Both bulked segregant analysis(BSA) and near isogenic lines (NIL) studies were done to identify the RFLP markers closely linked to cocoon shell parameters. Three hundred and fifty two random clones were identified as the low copy number sequence and used for identification of Restriction Fragment Length Polymorphic (RFLP) marker linked to cocoon weight and cocoon shell character. In the bulk segregant analysis, DNA from the parents (B20 A, C.Nichi), F 1 and F 2 progeny of high shell ratio (HSR) and low shell ratio (LSR) were screened for hybridization with the random clones. Polymorphic banding pattern achieved through southern hybridization with different probes indicated the probable correlation of polymorphism with high and low cocoon shell character which are possible landmarks in identifying the putative marker(s) for the cocoon shell character. Out of the 100 probes tried with parents, F 1, F 2 and their bulks, 10 probes were found to be closely linked to cocoon shell characters.展开更多
基金Supported by Agricultural Science and Technology Independent Innovation Fund of Jiangsu Province[CX(12)1003]Science and Technology Support Program of Jiangsu Province(BE2013301)Special Fund for the Construction of Modern Agriculture Industry System of China(CARS-01-47)~~
文摘Rice blast is one of the important diseases in major rice producing areas of China. The main blast resistance genes Pi-ta and Pi-b showed broad-spectrum and durable resistance to rice blast in many rice growing areas of China, which have been widely utilized in rice breeding and commercial production. In this study, on the basis of detection and verification of the genotypes of 22 rice varieties har- boring known blast resistance genes （Pi-ta and Pi-b） and blast susceptibility genes （pi-ta and pi-b）, two multiple PCR systems for these genes were established by us- ing the functional markers of blast resistance genes Pi-ta and Pi-b as well as blast susceptibility genes pi-ta and pi-b, respectively. Specifically, multiple PCR system I could simultaneously detect blast resistance genes Pi-ta and Pi-b, while system II could detect simultaneously blast susceptibility genes pi-ta and pi-b. In addition, the genotypes of 336 high generation breeding materials were detected with these two multiple PCR systems. The results were highly consistent with those of conventional single mark detection, indicating that these two multiplex PCR systems were stable, reliable and time-saving. The established multiplex PCR systems may serve as a rapid and efficient method to identify and screen rice germplasm resources and can be applied in marker-assisted selection to polymerize multiple genes for blast resis- tance in rice breeding.
文摘[Objective] This research aimed to search a best method for extracting the genomic DNA of Cymbidium ensifolium and establish the optimized ISSR-PCR reaction system.[Method] Genomic DNA was extracted from C.ensifolium leaves by modified CTAB method.ISSR-PCR reaction system for C.ensifolium was optimized.[Result] High-quality genomic DNA was obtained from C.ensifolium.The 25 μl optimized ISSR-PCR reaction system for C.ensifolium contained 2.5 μl 10× PCR buffer,2.5 mmol/L MgCl2,240 ng template DNA,160 μmol/L dNTPs,1.25 U Taq DNA polymerase,0.4 μmol/L primer and 15.78 μl ddH2O.The optimal PCR procedures were：94 ℃ pre-denaturation for 5 min and then 40 cycles,94 ℃ denaturation for 30 s,50-60 ℃ annealing for 30 s （annealing temperature according to different primers）,72 ℃ extension for 50 s and a 72 ℃ extension for 7 min.[Conclusion] An optimized ISSR-PCR reaction system for C.ensifolium was established,which provides a basis for further study on genetic diversity of C.ensifolium by using ISSR molecular marker technique.
文摘Two silkworm strains viz, B20 A (high cocoon shell ratio) and C.Nichi (low cocoon shell ratio) were sib mated for 10 generations to determine the homozygosis. Both bulked segregant analysis(BSA) and near isogenic lines (NIL) studies were done to identify the RFLP markers closely linked to cocoon shell parameters. Three hundred and fifty two random clones were identified as the low copy number sequence and used for identification of Restriction Fragment Length Polymorphic (RFLP) marker linked to cocoon weight and cocoon shell character. In the bulk segregant analysis, DNA from the parents (B20 A, C.Nichi), F 1 and F 2 progeny of high shell ratio (HSR) and low shell ratio (LSR) were screened for hybridization with the random clones. Polymorphic banding pattern achieved through southern hybridization with different probes indicated the probable correlation of polymorphism with high and low cocoon shell character which are possible landmarks in identifying the putative marker(s) for the cocoon shell character. Out of the 100 probes tried with parents, F 1, F 2 and their bulks, 10 probes were found to be closely linked to cocoon shell characters.
