Screening for resistance gene candidate from a genomic TAC library of Triticum aestivum-Haynaldia villosa translocation line 6VS/6AL by pooled PCR / 生物工程学报
Chinese Journal of Biotechnology
;
(12): 313-317, 2002.
Artigo
em Chinês
| WPRIM
| ID: wpr-231327
ABSTRACT
A pair of degenerate primers were designed based on NBS (nucleotide binding site, NBS) domain of resistance(R) gene and used to perform PCR with cDNA from the translocation line 6VS/6AL of Triticum aestivum-Haynaldia villosa. A clone (N7) characterized with NBS was obtained by sequencing analysis. Two specific primers were designed from the N7 sequence and used to screen a genomic TAC (transformation-competent artificial chromosome, TAC) library of 6VS/6AL consisting of ca. 2 x 10(6) clones. The library was stored as clone pools in twenty-two 96-well plates, each well containing approximately 1000 TAC clones. TAC plasmids were prepared from all the 2112 pools. Using a pooled PCR screening procedure, a positive TAC clone having a 40 kb insert was obtained. The positive clone was confirmed by Southern hybridization with the NBS fragment as a probe. The results indicate that the pooled PCR method is effective for screening of genomic libraries having large number of clones.
Texto completo:
DisponíveL
Índice:
WPRIM (Pacífico Ocidental)
Assunto principal:
Translocação Genética
/
Triticum
/
Sítios de Ligação
/
Dados de Sequência Molecular
/
Sequência de Bases
/
Biblioteca Genômica
/
Southern Blotting
/
Reação em Cadeia da Polimerase
/
Sequência de Aminoácidos
/
Cromossomos Artificiais
Tipo de estudo:
Estudo diagnóstico
/
Estudo de rastreamento
Idioma:
Chinês
Revista:
Chinese Journal of Biotechnology
Ano de publicação:
2002
Tipo de documento:
Artigo
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