Based on previous morphological classification and RAPD analysis, we recovered and sequenced genomic DNA of a specific fragment obtained from No. 45 Trichoderma harzianum strain in RAPD amplification. A pair of specif...Based on previous morphological classification and RAPD analysis, we recovered and sequenced genomic DNA of a specific fragment obtained from No. 45 Trichoderma harzianum strain in RAPD amplification. A pair of specific primers was synthesized and the RAPD markers were successfully converted into SCAR markers. The SCAR molecular marker exhibited the specificity that could differ T. harzianum from other Trichoderma strains. This pair of molecular primers was used for specific PCR amplification of 45 Trichoderma strains, and T. harzianum strains could amplify a 779 bp DNA band while other strains had no amplification products.展开更多
In this study, an F2 segregated population obtained by hybridization between the aphid-sensitive sorghum strain Qiansan and aphid-resistant cultivar Henong 16 was used to establish an aphid-resistant pool and an aphid...In this study, an F2 segregated population obtained by hybridization between the aphid-sensitive sorghum strain Qiansan and aphid-resistant cultivar Henong 16 was used to establish an aphid-resistant pool and an aphid-sensitive pool. 192 pairs of AFLP （amplified fragment length polymorphism） marker primers were screened in these pools using BSA （bulked segregant analysis）. Three pairs of EcoR I-CTG/Mse I-CCT, EcoR I-CTG/Mse I-CAT, and EcoR I-AGT/Mse I-CCC showed linkage with aphis resistance. EcoR I-CTG/Mse I-CCT-475, EcoR I-CTG/Mse I-CAT-390, and EcoR I-AGT/Mse I-CCC- 350 （E42/M52-350） were mapped within 6, 10, and 13 cM distances with the aphid-resistant gene by using Mapmaker 3.0 software. The bands amplified by EcoR I-CTG/Mse I-CCT-475 and EcoR I-CTG/Mse I-CAT-390 were extracted, cloned, and sequenced. Specific primers of SCAR （sequence characterized amplified regions） were then designed from these bands. A specific band of 300 bp was amplified by a pair of SCAR primers designed based on the sequence obtained from the EcoR I-CTG/Mse I-CAT-390 marker. The SCAR marker was named SCAS0. The marker was used to detect the F2, BC1, and F2：3 populations. The selective efficiency was 86.8, 91.1, and 86.3% in the BC1, F2, and F2：3 populations, respectively. The average selective efficiency was 88.2%.展开更多
The versatility of the PCR technique is that several kinds of primers can be explored for genome analysis depending on the purpose of study. The easy to access and low cost PCR-based markers include Random Amplified P...The versatility of the PCR technique is that several kinds of primers can be explored for genome analysis depending on the purpose of study. The easy to access and low cost PCR-based markers include Random Amplified Polymorphic DNA (RAPD). The RAPD markers are easy to develop but lack of reproducibility makes it less reliable and obstacles to their further use in authentication of traits. In addition, other PCR and non PCR based markers like Amplified Fragment Length Polymorphism (AFLP), Simple Sequence Repeat (SSR) and Restriction Fragment Length Polymorphism (RFLP) are also employed in authentication of traits with certain restrictions vis-à-vis use of radioactive materials, high cost and requirement of sequence information etc. Therefore, this problem can be overcome by converting RAPD markers to more robust sequence characterized amplified regions i.e. SCAR markers. SCARs are locus specific, co-dominant in nature and amplified by PCR using specific 15 - 30 bp DNA fragments. For developing SCAR markers, primers are designed from the nucleotide sequences of a cloned RAPD fragments linked to a trait of interest. SCAR markers are easy to develop and reliable tools for DNA fingerprinting. This mini review is an attempt to summarize efficacy of RAPD-SCAR interface in authentication of traits.展开更多
基金Supported by Natural Science Fund of Hainan Province in 2013"DNA Barcode Research of Medical Plants in Euphorbiaceae in Hainan Province"(813190)
文摘Based on previous morphological classification and RAPD analysis, we recovered and sequenced genomic DNA of a specific fragment obtained from No. 45 Trichoderma harzianum strain in RAPD amplification. A pair of specific primers was synthesized and the RAPD markers were successfully converted into SCAR markers. The SCAR molecular marker exhibited the specificity that could differ T. harzianum from other Trichoderma strains. This pair of molecular primers was used for specific PCR amplification of 45 Trichoderma strains, and T. harzianum strains could amplify a 779 bp DNA band while other strains had no amplification products.
基金the Natural Science Foundation, Hebei Province, China (C2010000758)the Science and Technology Department of Hebei Province,China (06547004D-2)
文摘In this study, an F2 segregated population obtained by hybridization between the aphid-sensitive sorghum strain Qiansan and aphid-resistant cultivar Henong 16 was used to establish an aphid-resistant pool and an aphid-sensitive pool. 192 pairs of AFLP （amplified fragment length polymorphism） marker primers were screened in these pools using BSA （bulked segregant analysis）. Three pairs of EcoR I-CTG/Mse I-CCT, EcoR I-CTG/Mse I-CAT, and EcoR I-AGT/Mse I-CCC showed linkage with aphis resistance. EcoR I-CTG/Mse I-CCT-475, EcoR I-CTG/Mse I-CAT-390, and EcoR I-AGT/Mse I-CCC- 350 （E42/M52-350） were mapped within 6, 10, and 13 cM distances with the aphid-resistant gene by using Mapmaker 3.0 software. The bands amplified by EcoR I-CTG/Mse I-CCT-475 and EcoR I-CTG/Mse I-CAT-390 were extracted, cloned, and sequenced. Specific primers of SCAR （sequence characterized amplified regions） were then designed from these bands. A specific band of 300 bp was amplified by a pair of SCAR primers designed based on the sequence obtained from the EcoR I-CTG/Mse I-CAT-390 marker. The SCAR marker was named SCAS0. The marker was used to detect the F2, BC1, and F2：3 populations. The selective efficiency was 86.8, 91.1, and 86.3% in the BC1, F2, and F2：3 populations, respectively. The average selective efficiency was 88.2%.
文摘The versatility of the PCR technique is that several kinds of primers can be explored for genome analysis depending on the purpose of study. The easy to access and low cost PCR-based markers include Random Amplified Polymorphic DNA (RAPD). The RAPD markers are easy to develop but lack of reproducibility makes it less reliable and obstacles to their further use in authentication of traits. In addition, other PCR and non PCR based markers like Amplified Fragment Length Polymorphism (AFLP), Simple Sequence Repeat (SSR) and Restriction Fragment Length Polymorphism (RFLP) are also employed in authentication of traits with certain restrictions vis-à-vis use of radioactive materials, high cost and requirement of sequence information etc. Therefore, this problem can be overcome by converting RAPD markers to more robust sequence characterized amplified regions i.e. SCAR markers. SCARs are locus specific, co-dominant in nature and amplified by PCR using specific 15 - 30 bp DNA fragments. For developing SCAR markers, primers are designed from the nucleotide sequences of a cloned RAPD fragments linked to a trait of interest. SCAR markers are easy to develop and reliable tools for DNA fingerprinting. This mini review is an attempt to summarize efficacy of RAPD-SCAR interface in authentication of traits.
