Sterility Analysis, Cloning And Bioinformatics Analysis Of Gene Related To CMS In Chinese Cabbage CMS Line RC7

CMS is a good parental line in the F1 breeding of Chinese cabbage. RC97-1, the new Brassica napus CMS material, was introduced by Ke Guilan, Zhao Limin, et al in 1997. The new Chinese cabbage CMS line RC7 was breeded throng hybriding between RC97-1, the CMS doner, and different types of Chinese cabbage and multi-generation backcrossing and severe screening from seedling, florescence to economic properties. The RC7 had a stable and complete male sterility with 100% of pollen abortion rate, no etiolated seedling, normal nectary, good combining ability. Throng breeding and selection of RC7, the new Chinese cabbage varieties Jinqiu70 and Jinqiu90 were breeded, which have obviously advantage and appeal to market, for applicate in production. In order to determine the mechanism of pollen abortion of CMS RC7 and provid a scientific basis for further application this excellent CMS line, this experiment tried to explore the sterile mechanism in anatomical and molecular biological study.1. Morphological anatomy of florescence bud and blooming flower of RC7 by paraffin sectioning indicated that RC7 buds appeared abortive and the abortion stage of RC7 was tetrad stage. Tapetum cells in RC7 were big abnormaly. Tetrad were subjected to sequeezing and then ruptured and degraded, thus couldn't format normal microspore.2. A 417bp special DNA fragment was amplificated in mtDNA of CMS line RC7 but its maintainer line B7. This 417bp special DNA fragment had 100% homology compared with orf138. This indicated that the infecundity of RC7 is similar to Ogu CMS line. Its cytoplasmic male sterility was derived from Ogu CMS line.3. A 588bp special DNA fragment was amplificated in mtDNA of CMS line RC7 rather than its maintainer line B7. This 588bp special DNA fragment could code 75 amino acids,and was designated as orf75. The amino acids sequence deduced from orf75 had an N-terminus, which had the same amino acids sequence of 28 amino acids with ORF138 of Ogu CMS radish, and a C-terminus, which had 54% homology compared with solute carrier family 24 (sodium/ potassium/ calcium exchanger) member 1. We concluded that orf75 maybe a new open reading frame, which originated from rearrangement of orf138 and the gene of solute carrier family 24 (sodium/potassium/calcium exchanger) member 1. This 588bp specific fragment could also code a protein of 67 amino acids. The coding sequence was named as orf67. This feasible amino acid was a soluble protein with a transmembrane helical regions and hydrophobic group.4. RT-PCR analysis indicated that the 588bp specific fragment expressed in both leaf and bud, and without post-transcriptional modification.