[Construction of physical map and polymorphism analysis of mtDNA region R from Mo17CMS-J of maize]

Total DNA from twenty-six CMS lines of maize under Mo17, 77 and W23 nuclear background were used for PCR amplification, including N, T, C, S four groups of cytoplasms. The primers was prefabricated according the sequence of R region published by Zebala (1997). Through these amplifications, mitochondrial DNA fragments were obtained from maize total DNA. Generally the results in one group are identical. And they are different from the others. The amplified fragments were sequenced and also give us much more information about the structure of mitochondrial genes that may lead to CMS. In order to isolate and identify the CMS genes, we developed a new platform to construct physical map of chromosome DNA by means of restriction enzyme double-digestion. The elongation of contigs is based on Southern hybridization. Having retrieved DNA from agarose gel after electrophoresis by beta-agarase, we labelled it with 32P-dCTP as a probe. We detected the positive clones in the gene library. Two contigs were revealed. And a restriction map covering 40 kb was constructed, including R region.