The semidwarf gene sd-g which has been used in indica rice breeding in southern China is a new one, non-allelic to sd-1. To map sd-g, an F2 population derived from the cross between Xinguiaishuangai and 02428 was con-...The semidwarf gene sd-g which has been used in indica rice breeding in southern China is a new one, non-allelic to sd-1. To map sd-g, an F2 population derived from the cross between Xinguiaishuangai and 02428 was con-structed. The sd-g was roughly mapped between two mi-crosatellite markers RM440 and RM163, with genetic dis-tances of 0.5 and 2.5 cM, respectively. Then nine new poly-morphic microsatellite markers were developed in this region. The sd-g was further mapped between two microsatellite markers SSR5-1 and SSR5-51, with genetic distances of 0.1 and 0.3 cM, respectively, while cosegregated with SSR418. A BAC contig was found to span the sd-g locus, the region be-ing delimited to 85 kb. This result was very useful for cloning of the sd-g gene.展开更多
Application and functional study of dwarf and semi-dwarf genes are of great importance to both crop breeding and molecular biology. A new semi-dwarf gene, sd-t(t), non-allelic to sd-1,had been identified in an indica ...Application and functional study of dwarf and semi-dwarf genes are of great importance to both crop breeding and molecular biology. A new semi-dwarf gene, sd-t(t), non-allelic to sd-1,had been identified in an indica rice variety, Aitaiyin 2. In this study the gene was genetically mapped by using an F2 population, which consisted of 474 individuals developed from a cross between Aitaiyin 2 and B30. The sd-t(t) gene was located between the RFLP markers R514 and R1408B with a distance of 1.1 cM to R514, and 4.5 cM to R1408B on chromosome 4. A physical contig covering the sd-t(t) mapping region was further constructed by screening a BAC library with R514 and R1408B as probes, and the physical distance between R514 and R1408B was estimated at approximately 147 kb. This result will facilitate map-based cloning of the sd-t(t) gene.展开更多
With the wide application of DNA sequencing technology, DNA sequences are still increasingly generated through the Sanger sequencing platform. SeqMan （in the LaserGene package） is an excellent program with an easy-t...With the wide application of DNA sequencing technology, DNA sequences are still increasingly generated through the Sanger sequencing platform. SeqMan （in the LaserGene package） is an excellent program with an easy-to-use graphical user interface （GUI） employed to assemble Sanger sequences into contigs. However, with increasing data size, larger sample sets and more sequenced loci make contig assemble complicated due to the considerable number of manual operations required to run SeqMan. Here, we present the ＇autoSeqMan＇ software program, which can automatedly assemble contigs using SeqMan scripting language. There are two main modules available, namely, ＇Classification＇ and ＇Assembly＇. Classification first undertakes preprocessing work, whereas Assembly generates a SeqMan script to consecutively assemble contigs for the classified files. Through comparison with manual operation, we showed that autoSeqMan saved substantial time in the preprocessing and assembly of Sanger sequences. We hope this tool will be useful for those with large sample sets to analyze, but with little programming experience. It is freely available at https：//github.com/ Sun-Yanbo/autoSeqMan.展开更多
As a part of the Multinational Genome Sequencing Project of Brassica rapa, linkage group R9 and R3 were sequenced using a bacterial artificial chromosome （BAC） by BAC strategy. The current physical contigs are expec...As a part of the Multinational Genome Sequencing Project of Brassica rapa, linkage group R9 and R3 were sequenced using a bacterial artificial chromosome （BAC） by BAC strategy. The current physical contigs are expected to cover approximately 90% euchromatins of both chromosomes. As the project progresses, BAC selection for sequence extension becomes more limited because BAC libraries are restriction enzyme-specific. To support the project, a random sheared fosmid library was constructed. The library consists of 97536 clones with average insert size of approximately 40 kb corresponding to seven genome equivalents, assuming a Chinese cabbage genome size of 550 Mb. The library was screened with primers designed at the end of sequences of nine points of scaffold gaps where BAC clones cannot be selected to extend the physical contigs. The selected positive clones were end-sequenced to check the overlap between the fosmid clones and the adjacent BAC clones. Nine fosmid clones were selected and fully sequenced. The sequences revealed two completed gap filling and seven sequence extensions, which can be used for further selection of BAC clones confirming that the fosmid library will facilitate the sequence completion of B. rapa.展开更多
A major gene for heading date in rice, Ef(t), was mapped on the same position as one RFLP marker C1369 on chromosome 10, which was located between two RFLP markers C234 and G37 at 1.0 cM interval. Initially, these thr...A major gene for heading date in rice, Ef(t), was mapped on the same position as one RFLP marker C1369 on chromosome 10, which was located between two RFLP markers C234 and G37 at 1.0 cM interval. Initially, these three RFLP markers were used to screen a rice BAC library and seven independent clones with the size ranged from 70kb to 180kb were identified. By the comparisons of Hind III restriction fragment in each clone, the relative location of these clones were determined and two primary contigs, contig C1369 and contig G37, were obtained. Chromosome walking was performed with one outmost BAC end of the primary BAC contig C1369, then two contigs were integrated into one. The resultant contig encompassing Ef(t) gene locus which consisted of 7 BAC clones was developed. It will facilitate the isolation of Ef(t) gene using map based cloning approach.展开更多
The YAC contig construction has been done for the Human X chromosome short armXp21.3—p11.3, a region which contains several genetic disease gene loci and is of highlybiomedical importance. Using known probes(OTC, DXS...The YAC contig construction has been done for the Human X chromosome short armXp21.3—p11.3, a region which contains several genetic disease gene loci and is of highlybiomedical importance. Using known probes(OTC, DXS166, DMDcDNA) and STS markersof this region, YAC screenings are performed by both YAC colony in situ hybridization andPCR methods. Totally 55 YACs are obtained from the YAC libraries of CEPH, ICRF andthe Institute. The size determination, the analysis of 26 pairs of microsatelite STS, the single copy probe hybridization and the Alu-PCR fingerprinting are performed for these YACs.The mapping of these YACs is performed, and finally, 6 YAC contigs in Xp21.3—11.3 are obtained, covering about 15 Mb. This work will greatly facilitate the positional cloning of disease genes or the genome sequencing in this important region.展开更多
Pepper （Capsicum annuum. L.） is a widely cultivated vegetable crop worldwide and has the second largest planting area and the first largest vegetable output and value in China. Pepper root-knot nematode （Meloidogyn...Pepper （Capsicum annuum. L.） is a widely cultivated vegetable crop worldwide and has the second largest planting area and the first largest vegetable output and value in China. Pepper root-knot nematode （Meloidogyne spp.） is one of the most serious pests of pepper, which caused huge losses every year. Previous studies showed that the Me3 gene is resistant to a wide range of Meloidogyne species, including M. arenaria, M. javanica, and M. incognita. HDA149, a double haploid pepper genotype, harboring the root-knot nematode resistance gene Me3, was used to construct bacterial artificial chro- mosome library （BAC） via the vector of CopyControFM pCC1 in this study. The library consists of 210 200 BAC clones and is equivalent to 5.3 pepper genomes. The average insert size is 95 kb, and most of them are 90-120 kb; but the empty clones are less than 3%. In order to screen the BAC library easily, 550 super pools with 384 BAC clones of each pool were further developed in this study. Specific primers from Me3 gene locus were used for BAC library screening, and more than 20 positive BAC clones were obtained. Then the selected positive BAC clones were analyzed by restriction enzyme digestion, BAC-end sequencing, marker development, and new positive BAC clones exploration, respectively. Finally, the contig with total length of about 300 kb linked to the Me3 locus was constructed based on chromosome walking strategy, which made a solid foundation for the cloning of the important root-knot nematode resistance gene Me3.展开更多
文摘The semidwarf gene sd-g which has been used in indica rice breeding in southern China is a new one, non-allelic to sd-1. To map sd-g, an F2 population derived from the cross between Xinguiaishuangai and 02428 was con-structed. The sd-g was roughly mapped between two mi-crosatellite markers RM440 and RM163, with genetic dis-tances of 0.5 and 2.5 cM, respectively. Then nine new poly-morphic microsatellite markers were developed in this region. The sd-g was further mapped between two microsatellite markers SSR5-1 and SSR5-51, with genetic distances of 0.1 and 0.3 cM, respectively, while cosegregated with SSR418. A BAC contig was found to span the sd-g locus, the region be-ing delimited to 85 kb. This result was very useful for cloning of the sd-g gene.
基金This work was supported by the Chinese 973 Program (Grant Nos. G1999011606 & G1999011604).
文摘Application and functional study of dwarf and semi-dwarf genes are of great importance to both crop breeding and molecular biology. A new semi-dwarf gene, sd-t(t), non-allelic to sd-1,had been identified in an indica rice variety, Aitaiyin 2. In this study the gene was genetically mapped by using an F2 population, which consisted of 474 individuals developed from a cross between Aitaiyin 2 and B30. The sd-t(t) gene was located between the RFLP markers R514 and R1408B with a distance of 1.1 cM to R514, and 4.5 cM to R1408B on chromosome 4. A physical contig covering the sd-t(t) mapping region was further constructed by screening a BAC library with R514 and R1408B as probes, and the physical distance between R514 and R1408B was estimated at approximately 147 kb. This result will facilitate map-based cloning of the sd-t(t) gene.
基金supported by the National Natural Science Foundation of China(31671326)the Youth Innovation Promotion Association,Chinese Academy of Sciences
文摘With the wide application of DNA sequencing technology, DNA sequences are still increasingly generated through the Sanger sequencing platform. SeqMan （in the LaserGene package） is an excellent program with an easy-to-use graphical user interface （GUI） employed to assemble Sanger sequences into contigs. However, with increasing data size, larger sample sets and more sequenced loci make contig assemble complicated due to the considerable number of manual operations required to run SeqMan. Here, we present the ＇autoSeqMan＇ software program, which can automatedly assemble contigs using SeqMan scripting language. There are two main modules available, namely, ＇Classification＇ and ＇Assembly＇. Classification first undertakes preprocessing work, whereas Assembly generates a SeqMan script to consecutively assemble contigs for the classified files. Through comparison with manual operation, we showed that autoSeqMan saved substantial time in the preprocessing and assembly of Sanger sequences. We hope this tool will be useful for those with large sample sets to analyze, but with little programming experience. It is freely available at https：//github.com/ Sun-Yanbo/autoSeqMan.
基金This work was supported by grants from the National Academy of Agricultural Science(Code #200901FHT020508369)the BioGreen21 Program(Code #20050301034438 and Code #20070301034037),Rural Development Administration, Republic of Korea
文摘As a part of the Multinational Genome Sequencing Project of Brassica rapa, linkage group R9 and R3 were sequenced using a bacterial artificial chromosome （BAC） by BAC strategy. The current physical contigs are expected to cover approximately 90% euchromatins of both chromosomes. As the project progresses, BAC selection for sequence extension becomes more limited because BAC libraries are restriction enzyme-specific. To support the project, a random sheared fosmid library was constructed. The library consists of 97536 clones with average insert size of approximately 40 kb corresponding to seven genome equivalents, assuming a Chinese cabbage genome size of 550 Mb. The library was screened with primers designed at the end of sequences of nine points of scaffold gaps where BAC clones cannot be selected to extend the physical contigs. The selected positive clones were end-sequenced to check the overlap between the fosmid clones and the adjacent BAC clones. Nine fosmid clones were selected and fully sequenced. The sequences revealed two completed gap filling and seven sequence extensions, which can be used for further selection of BAC clones confirming that the fosmid library will facilitate the sequence completion of B. rapa.
文摘A major gene for heading date in rice, Ef(t), was mapped on the same position as one RFLP marker C1369 on chromosome 10, which was located between two RFLP markers C234 and G37 at 1.0 cM interval. Initially, these three RFLP markers were used to screen a rice BAC library and seven independent clones with the size ranged from 70kb to 180kb were identified. By the comparisons of Hind III restriction fragment in each clone, the relative location of these clones were determined and two primary contigs, contig C1369 and contig G37, were obtained. Chromosome walking was performed with one outmost BAC end of the primary BAC contig C1369, then two contigs were integrated into one. The resultant contig encompassing Ef(t) gene locus which consisted of 7 BAC clones was developed. It will facilitate the isolation of Ef(t) gene using map based cloning approach.
基金Supported by the High Technology Research and Development Programme of Chinathe National Natural Science Foundation of China
文摘The YAC contig construction has been done for the Human X chromosome short armXp21.3—p11.3, a region which contains several genetic disease gene loci and is of highlybiomedical importance. Using known probes(OTC, DXS166, DMDcDNA) and STS markersof this region, YAC screenings are performed by both YAC colony in situ hybridization andPCR methods. Totally 55 YACs are obtained from the YAC libraries of CEPH, ICRF andthe Institute. The size determination, the analysis of 26 pairs of microsatelite STS, the single copy probe hybridization and the Alu-PCR fingerprinting are performed for these YACs.The mapping of these YACs is performed, and finally, 6 YAC contigs in Xp21.3—11.3 are obtained, covering about 15 Mb. This work will greatly facilitate the positional cloning of disease genes or the genome sequencing in this important region.
基金supported by the National High-Tech R&D Program in China (2013AA102603)the Natural Science Foundation of Shandong Province,China (ZR2014YL014)+3 种基金the Youth Scientific Research Foundation of Shandong Academy of Agricultural Sciences,China (2014QNZ03)the Taishan Scholars Program of Shandong Province,China (2016-2020)the National Natural Science Foundation of China (31101425)Prof. Alain Palloxin,French National Institute for Agricultural Research (INRA),for kindly providing the pepper genotype HDA149
文摘Pepper （Capsicum annuum. L.） is a widely cultivated vegetable crop worldwide and has the second largest planting area and the first largest vegetable output and value in China. Pepper root-knot nematode （Meloidogyne spp.） is one of the most serious pests of pepper, which caused huge losses every year. Previous studies showed that the Me3 gene is resistant to a wide range of Meloidogyne species, including M. arenaria, M. javanica, and M. incognita. HDA149, a double haploid pepper genotype, harboring the root-knot nematode resistance gene Me3, was used to construct bacterial artificial chro- mosome library （BAC） via the vector of CopyControFM pCC1 in this study. The library consists of 210 200 BAC clones and is equivalent to 5.3 pepper genomes. The average insert size is 95 kb, and most of them are 90-120 kb; but the empty clones are less than 3%. In order to screen the BAC library easily, 550 super pools with 384 BAC clones of each pool were further developed in this study. Specific primers from Me3 gene locus were used for BAC library screening, and more than 20 positive BAC clones were obtained. Then the selected positive BAC clones were analyzed by restriction enzyme digestion, BAC-end sequencing, marker development, and new positive BAC clones exploration, respectively. Finally, the contig with total length of about 300 kb linked to the Me3 locus was constructed based on chromosome walking strategy, which made a solid foundation for the cloning of the important root-knot nematode resistance gene Me3.
