A simple method to create a chromosome-specific DNA library of rice, including microdissection, amplification,characterization and cloning, is described. Rice chromosome 4 from a metaphase cell has been isolated and a...A simple method to create a chromosome-specific DNA library of rice, including microdissection, amplification,characterization and cloning, is described. Rice chromosome 4 from a metaphase cell has been isolated and amplified by the Linker Adapter PCR (LA-PCR). The PCR products were labeled as probes with DIG-11-dUTP using the random priming method.Southern blot analysis with rice genomic DNA and specific RFLP markers demonstrated that the PCR products were derived from rice chromosome 4. A large library comprising over 100,000recombinant plasmid microclones from rice chromosome 4was constructed. Colony hybridization showed that 58% of the clones contained single or low-copy sequences and 42%contained repetitive sequences. The size of inserts generated by PCR ranged from 140bp to 500bp.This method will facilitate cloning of the specific chromosome DNA markers and important genes of rice.展开更多
文摘A simple method to create a chromosome-specific DNA library of rice, including microdissection, amplification,characterization and cloning, is described. Rice chromosome 4 from a metaphase cell has been isolated and amplified by the Linker Adapter PCR (LA-PCR). The PCR products were labeled as probes with DIG-11-dUTP using the random priming method.Southern blot analysis with rice genomic DNA and specific RFLP markers demonstrated that the PCR products were derived from rice chromosome 4. A large library comprising over 100,000recombinant plasmid microclones from rice chromosome 4was constructed. Colony hybridization showed that 58% of the clones contained single or low-copy sequences and 42%contained repetitive sequences. The size of inserts generated by PCR ranged from 140bp to 500bp.This method will facilitate cloning of the specific chromosome DNA markers and important genes of rice.
