关键词: 甘蓝型油菜, 细胞质雄性不育, 差异表达, 花粉败育, cDNA–AFLP

Abstract:

Previous studies showed that NCa CMS was a novel CMS and would be of great importance for rapeseed hybrid breeding and production. In this article, differentially expressed genes during pollen abortion among NCa cms, maintainer line and fertile F1 were analyzed by means of cDNA-AFLP. 224 pair of AFLP primers were used to analyze the cDNA of the above 3 materials. The transcript-derived fragments (TDFs) were cloned, and then 41 TDFs were sequenced after identified as positive clones. Most of the TDFs were cloned firstly in Brassica napus. Gene function analysis revealed that 82% of TDFs were related with metabolism processes of (1) cell formation and degradation, (2) protein synthesis, processing and transporting, (3) energy metabolism and (4) signal transferring, involving all the key metabolism processes during pollen development. The differential expression of seven key genes among these TDFs in young leaves and floral buds of NCa cms and maintainer line were further analyzed by RT-PCR. The pectin esterase gene and amino acid permease gene abundantly expressed in floral buds of NCa cms while pectin esterase gene showed no transcript either in young leaves of NCa cms or in the two tissues of maintainer line, and amino acid permease gene slightly expressed in the corresponding tissues. However, kinesin gene and aldehyde dehydrogenase (ALDH) gene expressed abundantly in floral buds of maintainer line but just slightly detected in young leaves of maintainer line and both tissues of NCa cms. The transcriptions of ZEP gene and mono-dehydroascorbate reductase were reverse to that of amino acid permease gene. The transcription of pollen-special oleosin-like protein gene was detected almost twice in abundance in floral buds of maintainer line than in those of NCa cms, whereas not detected in young leaves of the two materials. As for the gene transcription in the floral buds, the differentially expressed genes cloned from the NCa cms were detected abundantly in its floral buds but almost no transcription in those of maintainer line, and vice versa, which demonstrated that the authenticity and efficiency of cDNA–AFLP applied to reveal and clone differentially expressed genes. Further studies were underway in order to investigate the relationship between these genes with NCa pollen abortion.

Key words: Brassica napus L., cytoplasmic male sterility (CMS), differential expression, pollen abortion, cDNA–AFLP