Stem lignin content(SLC) in common wheat(Triticum aestivum L.) contributes to lodging resistance. Caffeic acid 3-O-methyltransferase(COMT) is a key enzyme involved in lignin biosynthesis. Characterization of TaCOMT ge...Stem lignin content(SLC) in common wheat(Triticum aestivum L.) contributes to lodging resistance. Caffeic acid 3-O-methyltransferase(COMT) is a key enzyme involved in lignin biosynthesis. Characterization of TaCOMT genes and development of gene-specific markers could enable marker-assisted selection in wheat breeding. In the present study, the full-length genomic DNA(gDNA) sequences of TaCOMT genes located on chromosomes 3 A, 3 B, and 3 D were cloned by homologous cloning. Two allelic variants, TaCOMT-3 Ba and TaCOMT-3 Bb, were identified and differed by a 222-bp insertion/deletion(InDel) in the 3′-untranslated region(3′-UTR). A co-dominant gene-specific marker based on this InDel was developed and designated as Ta COMT-3 BM. A total of 157 wheat cultivars and advanced lines grown in four environments were used to validate the associations between allelic patterns and SLC. The SLC of cultivars with TaCOMT-3 Ba was significantly(P<0.01) higher than that of those with TaCOMT-3 Bb, and the marker TaCOMT-3 BM could be effectively used in wheat breeding.展开更多
In marker-assisted breeding for bacterial blight(BB) resistance in rice, three major resistance genes, viz., Xa21, xa13 and xa5, are routinely deployed either singly or in combinations. As efficient and functional mar...In marker-assisted breeding for bacterial blight(BB) resistance in rice, three major resistance genes, viz., Xa21, xa13 and xa5, are routinely deployed either singly or in combinations. As efficient and functional markers are yet to be developed for xa13 and xa5, we have developed simple PCR-based functional markers for both the genes. For xa13, we designed a functional PCR-based marker, xa13-prom targeting the In Del polymorphism in the promoter of candidate gene Os8N3 located on chromosome 8 of rice. With respect to xa5, a multiplex-PCR based functional marker system, named xa5 FM, consisting of two sets of primer pairs targeting the 2-bp functional nucleotide polymorphism in the exon II of the gene TFIIA5(candidate for xa5), has been developed. Both xa13-prom and xa5 FM can differentiate the resistant and susceptible alleles for xa13 and xa5, respectively, in a co-dominant fashion. Using these two functional markers along with the already reported functional PCR-based marker for Xa21(p TA248), we designed a single-tube multiplex PCR based assay for simultaneous detection of all the three major resistance genes and demonstrated the utility of the multiplex marker system in a segregating population.展开更多
61 varieties of wheat collected in the gene fund of the Research Institute of Crop Husbandry were screened using SCAR-markers associated with the gene of resistance to brown leaf rust, Lr19. As a result of PCR analysi...61 varieties of wheat collected in the gene fund of the Research Institute of Crop Husbandry were screened using SCAR-markers associated with the gene of resistance to brown leaf rust, Lr19. As a result of PCR analysis using SCS123 marker the 737 bp locus was detected in 48 genotypes. The expected fragment of the 688 bp was detected in 53 genotypes using the SCS253 marker. The results obtained using both markers indicate that the Lr19 gene is present on 7D chromosomes of 45 genotypes. The existence of the Lr19 gene has not been proven only for 5 from the 61 analyzed wheat genotypes.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
The aim of this study was to detect the expression of 4 clinically-important efflux pumps in the Resistance-Nodulation-Cell Division (RND) family including MexAB-OprM, MexXY, MexCD-OprJ and MexEF-OprN in Pseudomonas a...The aim of this study was to detect the expression of 4 clinically-important efflux pumps in the Resistance-Nodulation-Cell Division (RND) family including MexAB-OprM, MexXY, MexCD-OprJ and MexEF-OprN in Pseudomonas aeruginosa using a combination of resistance-phenotypic markers and multiplex RT-PCR (mRT-PCR). The antibiotic substrates specific for each Mex systems were used as phenotypic markers including carbenicillin, MexAB-OprM, erythromycin, MexCD-OprJ, norfloxacin and imipenem, MexEF-OprN and gentamicin, MexXY-OprM. The methods were validated with reference strains with known genotypes of the Mex systems and the potential applicability in clinical practice was tested with clinical isolates. The results for the reference strains support that the combination of resistance phenotype and mRT-PCR is a potential-attractive method for diagnosis of efflux-mediated resistance in P. aeruginosa. Further development to make it more practical for clinical use and study in a larger number of clinical isolates is required.展开更多
In order to identify the molecular markers that can be widely used in the breeding of Brassica napus L.varieties with high seed oil content under different genetic backgrounds,we developed a Kompetitive Allele Specifi...In order to identify the molecular markers that can be widely used in the breeding of Brassica napus L.varieties with high seed oil content under different genetic backgrounds,we developed a Kompetitive Allele Specific PCR(KASP)marker for seed oil content on the basis of the results from available studies.The verification in the F_(2) population showed that the marker was closely linked to the quantitative trait locus(QTL)for oil content on chromosome A05.The findings helped to breed the‘Fengyou’varieties with high seed oil content in the middle reaches of the Yangtze River.展开更多
[Objective] This study was to screen the economic or stable PCR system of rice and detect the generality of the selected system in different molecular markers based on PCR.[Method] With DNA extracted from rice leaves ...[Objective] This study was to screen the economic or stable PCR system of rice and detect the generality of the selected system in different molecular markers based on PCR.[Method] With DNA extracted from rice leaves by CTAB method as the template,PCR system was optimized by L16(45)orthogonal design.[Result] Clear bands were amplified from 16 different combinations,but the amplification effects and yields had difference.The most economic and applicable system was as follows:20 ng DNA template,150 μmol/L dNT...展开更多
基金supported by the National Natural Science Foundation of China(31161140346 and 31461143021)the Beijing Municipal Science and Technology Project,China(D151100004415003)+1 种基金the National Key Technology R&D Program of China(2014BAD01B05)the earmarked fund for China Agriculture Research System(CARS-3-1-3)
文摘Stem lignin content(SLC) in common wheat(Triticum aestivum L.) contributes to lodging resistance. Caffeic acid 3-O-methyltransferase(COMT) is a key enzyme involved in lignin biosynthesis. Characterization of TaCOMT genes and development of gene-specific markers could enable marker-assisted selection in wheat breeding. In the present study, the full-length genomic DNA(gDNA) sequences of TaCOMT genes located on chromosomes 3 A, 3 B, and 3 D were cloned by homologous cloning. Two allelic variants, TaCOMT-3 Ba and TaCOMT-3 Bb, were identified and differed by a 222-bp insertion/deletion(InDel) in the 3′-untranslated region(3′-UTR). A co-dominant gene-specific marker based on this InDel was developed and designated as Ta COMT-3 BM. A total of 157 wheat cultivars and advanced lines grown in four environments were used to validate the associations between allelic patterns and SLC. The SLC of cultivars with TaCOMT-3 Ba was significantly(P<0.01) higher than that of those with TaCOMT-3 Bb, and the marker TaCOMT-3 BM could be effectively used in wheat breeding.
基金the funding support provided by the Department of Biotechnology(DBT),Government of India(Grant Nos.BT/AB/FG-2 (PH-II)/2009 and BT/PR11705/AGR/02/646/2008)
文摘In marker-assisted breeding for bacterial blight(BB) resistance in rice, three major resistance genes, viz., Xa21, xa13 and xa5, are routinely deployed either singly or in combinations. As efficient and functional markers are yet to be developed for xa13 and xa5, we have developed simple PCR-based functional markers for both the genes. For xa13, we designed a functional PCR-based marker, xa13-prom targeting the In Del polymorphism in the promoter of candidate gene Os8N3 located on chromosome 8 of rice. With respect to xa5, a multiplex-PCR based functional marker system, named xa5 FM, consisting of two sets of primer pairs targeting the 2-bp functional nucleotide polymorphism in the exon II of the gene TFIIA5(candidate for xa5), has been developed. Both xa13-prom and xa5 FM can differentiate the resistant and susceptible alleles for xa13 and xa5, respectively, in a co-dominant fashion. Using these two functional markers along with the already reported functional PCR-based marker for Xa21(p TA248), we designed a single-tube multiplex PCR based assay for simultaneous detection of all the three major resistance genes and demonstrated the utility of the multiplex marker system in a segregating population.
文摘61 varieties of wheat collected in the gene fund of the Research Institute of Crop Husbandry were screened using SCAR-markers associated with the gene of resistance to brown leaf rust, Lr19. As a result of PCR analysis using SCS123 marker the 737 bp locus was detected in 48 genotypes. The expected fragment of the 688 bp was detected in 53 genotypes using the SCS253 marker. The results obtained using both markers indicate that the Lr19 gene is present on 7D chromosomes of 45 genotypes. The existence of the Lr19 gene has not been proven only for 5 from the 61 analyzed wheat genotypes.
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..
文摘The aim of this study was to detect the expression of 4 clinically-important efflux pumps in the Resistance-Nodulation-Cell Division (RND) family including MexAB-OprM, MexXY, MexCD-OprJ and MexEF-OprN in Pseudomonas aeruginosa using a combination of resistance-phenotypic markers and multiplex RT-PCR (mRT-PCR). The antibiotic substrates specific for each Mex systems were used as phenotypic markers including carbenicillin, MexAB-OprM, erythromycin, MexCD-OprJ, norfloxacin and imipenem, MexEF-OprN and gentamicin, MexXY-OprM. The methods were validated with reference strains with known genotypes of the Mex systems and the potential applicability in clinical practice was tested with clinical isolates. The results for the reference strains support that the combination of resistance phenotype and mRT-PCR is a potential-attractive method for diagnosis of efflux-mediated resistance in P. aeruginosa. Further development to make it more practical for clinical use and study in a larger number of clinical isolates is required.
文摘In order to identify the molecular markers that can be widely used in the breeding of Brassica napus L.varieties with high seed oil content under different genetic backgrounds,we developed a Kompetitive Allele Specific PCR(KASP)marker for seed oil content on the basis of the results from available studies.The verification in the F_(2) population showed that the marker was closely linked to the quantitative trait locus(QTL)for oil content on chromosome A05.The findings helped to breed the‘Fengyou’varieties with high seed oil content in the middle reaches of the Yangtze River.
基金Supported by the Key Project from Department of Science and Technology of Heilongjiang Province(GA06B102-3-4)the Key Project from Heilongjiang Provincial Reclamation General Administration(HNKXIV-01-01-02)~~
文摘[Objective] This study was to screen the economic or stable PCR system of rice and detect the generality of the selected system in different molecular markers based on PCR.[Method] With DNA extracted from rice leaves by CTAB method as the template,PCR system was optimized by L16(45)orthogonal design.[Result] Clear bands were amplified from 16 different combinations,but the amplification effects and yields had difference.The most economic and applicable system was as follows:20 ng DNA template,150 μmol/L dNT...
