Directional Transfer And Utilization Of Multiple-allele Male Sterile Line Of Brassica Campestris Ssp. Chinensis Var Rosularis Tsen Et L. And Brassica Campestris Ssp. Chinenesis L.

Wutacai(Brassica campestris ssp. chinensis var rosularis Tsen et Lee) and Baicaitai (Brassica campestris ssp. chinenesis L.) are two spetical types of Brassica crops (AA,2n=2x=20) which own significant hybrid vigor. In order to resolve the problem of breeding in utilizing hybrid vigor, a "directional transfer program" was designed to breed the multiple-allele male sterile lines of Wutacai and Baicaitai. The multiple-allele male sterile lines of Naibaicai(Brassica campestris ssp. chinensis L., GMS-1) and Chinese cabbage (GMS-2) were used as the male sterile resourse, and the inbred lines of Wutacai (WT01) and Baitaicai (BrO2) were used as the target lines. Crossing, recurrent backcrossing and selfing were employed to transfer the male sterility and other botanical traits simultaneously. Several SSR markers were adopted for molecular marker-assisted selection (MAS) to solve the problem in identifying genotypes during the transferring of the male sterile line in Baicaitai. Aming at studying the molecular mechanism of aborted bud which found in male sterile line of Wutacai, cDNA-AFLP technology was used to analyze gene expression difference between normal bud and aborted bud. The main results were as follows.1. The genotype of the inbred line’TW01’was cheked by using a male sterile line (Msms) of Naibaicai’GMS-1’. It indicated that genotype of’TW01’was MsfMsf. According to the genotype, a directional transfer program was designed to transfer male sterile line. After backcrossing for four generations, a new male sterile line,’GMS-3’, with similar botanical traits to’WT01’,100%sterility degree and100%sterility rate, was bred.2. Four hybrid combinations were generated with’GMS-3’as female parent and four excellent inbred lines as male parents. The’Vitamin’ was as a control. Evenness degree, quality and yield of hybrid combinations were analyzed through Variety comparison test. Four hybrid combinations showed high evenness drgree in botanical traits. The component analysis results show that the components contents contained soluble sugar, protein, vitamin C, organic acids, crude fiber and trace elements were also at a high level. Most quality index is significantly higher than that of control. Yield of hybrid combinations showed remarkable hybrid vigor and signify cant differences was found between combinations and control. An excellent one,’Cl’was developed from the new male sterile line.3. Normal bud and aborted bud of the male sterile line of Wutacai were collected from primary branches of five plants, respectively. RNA was extracted from mixed buds to set up gene pools. cDNA-AFLP technology and bioinformatics analysis were used to analyze gene expression difference between normal bud and aborted bud. A total of64differential expressed transcript-derived fragments (TDFs) which were between80bp and400bp were obtained. They were divided into four types according to expression of "normal budâ†'aborted bud":a:"presentâ†'absent" type, approximately accounting for43.0%; b:"absentâ†'present" type, approximately accounting for36.5%; c:"strongâ†'weak" type, approximately accounting for10.8%; d:"weakâ†'strong" type, approximately accounting for9.7%.42typical TDFs were chosen to sequence. The sequencing results were compared similarity by BRAD and NCBI.16TDFs similarity were more than90%. Gene functions involved eight types:â‘ 1TDF was related with energy and metabolism;â‘¡5TDFs was related with stress response;â‘¢1TDF was related with signal transduction;â‘£4TDFs was related with transcription and translation;⑤2TDFs was related with disease resistance and senescence;â‘¥2TDFs was related with amino acid synthesizing and processing;⑦3TDFs was related with transmembrane transport;⑧2TDFs were function unknown.4. The genotype of the inbred line’Br02’was cheked by using a male sterile line (Msms) of Chinese cabbage’GMS-2’. F1plants genotype identification results indicated that genotype of’Br02’was MsfMsf. According to the genotype, a directional transfer program was designed to transfer male sterile line. Recurrent backcrossing method was employed to transfer male sterility and botanical traits while the genotype was identified by MAS. The GSSR1, which is closely linked to the male sterility gene "Ms", was chosen from26pairs of SSR primers to apply in the selection. GSSR1also could select the fertile gene "ms" and the fertile restoration gene "Msf". After three times recurrent backcrossing, a new genetic male sterile line of Baicaitai with100%of male sterile plants and100%of male sterility,’GMS-4’, was bred. The new male sterile line was similar to’Br02’in most botanical traits.5. Six hybrid combinations were generated with’GMS-4’ as female parent and excellent inbred lines as male parents. The evenness degree and yield of hybrid combinations were analyzed through Variety comparison test. High evenness drgree of botanical traits were showed between blocks and that showed much difference between hybrid combinations. Hybrid vigor was in hybrid combinations and signi-ficant differences were found between combinations and control. C3was developed as the excellent hybrid combination.