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
N-Methylene phosphonic chitosan （NMPCS）, an amphiphilic macromolecule with powerful chelating ability of Ca^2＋ ions, was synthesized and characterized. The physicochernical properties of NMPCS and the interactions ...N-Methylene phosphonic chitosan （NMPCS）, an amphiphilic macromolecule with powerful chelating ability of Ca^2＋ ions, was synthesized and characterized. The physicochernical properties of NMPCS and the interactions between NMPCS and plasmid DNA were investigated by FTIR, ^13C NMR, X-ray, agarose gel electrophoresis retardation assay, atomic force microscopy （AFM） and circular dichroism （CD）. The results suggest that at charge ratio 2：1 or above, DNA could be completely entrapped and spherical complexes with mean size of 80-210 nm were formed. Taking HeLa as host cell, luciferase expression mediated by NMPCS improved about 100 times compared to the expression mediated by chitosan.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.
基金supports from National Natural Science Foundation of China(No.50233020,30300086)the financial support from Natural Science Foundation of Tianjin(No.05YFJMJC10200).
文摘N-Methylene phosphonic chitosan （NMPCS）, an amphiphilic macromolecule with powerful chelating ability of Ca^2＋ ions, was synthesized and characterized. The physicochernical properties of NMPCS and the interactions between NMPCS and plasmid DNA were investigated by FTIR, ^13C NMR, X-ray, agarose gel electrophoresis retardation assay, atomic force microscopy （AFM） and circular dichroism （CD）. The results suggest that at charge ratio 2：1 or above, DNA could be completely entrapped and spherical complexes with mean size of 80-210 nm were formed. Taking HeLa as host cell, luciferase expression mediated by NMPCS improved about 100 times compared to the expression mediated by chitosan.
