甘蓝型油菜与紫罗兰体细胞杂交研究  

Studies on Somatic Hybridization between Brassica napus and Malthiola incana B. Br

作　　者：林良斌[1] 马占强[2] 张传利[1] 刘雅婷[1] 毛孝强[1] 厦国银[1] 马忠琴[1] 

机构地区：[1]云南农业大学农学与生物技术学院,云南昆明650201 [2]河南科技大学生命科学学院,河南洛阳471000

出　　处：《云南农业大学学报（自然科学版）》2008年第3期295-301,共7页Journal of Yunnan Agricultural University:Natural Science

基　　金：云南省自然科学基金(2004C0035M)

摘　　要：以甘蓝型油菜湘油15号和紫罗兰的下胚轴为材料,分离制备原生质体。采用PEG-高Ca2+-高pH法进行原生质体融合,在PEG浓度为30%,原生质体融合密度为5.0×105个/mL下融合25min,融合率可达17%。在附加1.5mg/L2,4-D,0.5mg/LNAA,0.5mL/LBA的改良的KM8p培养基中,以0.3mol/L蔗糖和0.1mol/L葡萄糖作渗透稳定剂进行液体浅层培养融合体,培养3～6d细胞发生第1次分裂,7d时统计分裂频率最高为13.8%,35d后形成大量的细胞团和肉眼可见的小愈伤组织,其植板率最高为6.5%。然后转移到含有0.1mg/L2,4-D和0.2mg/LBA的MS固体培养基上使其增殖。当愈伤组织长至5mm左右时,将其转到含有0.5mg/LBA,0.1mg/LNAA和0.2mg/LGA3的MS分化培养基上诱导芽分化。当芽长1～2cm时,将其切下插入附加0.5mg/LIBA的1/2MS生根培养基上,两周后即可获得完整植株。细胞学鉴定表明获得了杂种植株。Protoplast isolated from hypocotyls of cultivar Brassica napus. Xiangyou 15 and Malthiola incana B. Br were fused by the method of PEG-High Ca2＋ -High pH and the highest fusion frequency （ 17% ） was obtained while fusion solution contained 30% PEG and fusion was carried out at the density of 5.0×10^5 protoplasts/mL for 25 min. The fusion-protoplasts were then cultured by ＂thin liquid layer＂ method in improved KMSp medium supplemented with 1.5 mg/L 2, 4-D, 0. 5 mg/L NAA and 0.5 ml/L BA , 0. 3 mol/L sucrose and 0. 1 mol/L glucose that both were used as osmotic regulator. The fusion-protoplasts started to divide after 3 - 6 days, and the division frequency was 13.8% after being cultured for 7 days. The cell-masses and small calluses could be obtained after 35 days, and the plating efficiency was 6.5 %, then transferred to the proliferation MS medium supplemented with 0. 1 mg/L 2, 4-D, and 0. 2 mg/L BA. Calluses of 5mm in diameter were transferred on MS differentiation medium supplemented with O. 5 mg/L BA, 0. 1 mg/L NAA and 0. 2 mg/L GA3, and whole plants were obtained after transferring 1 - 2 em shoots to 1/2 MS medium with 0. 5 mg/L IBA for 2 weeks. Cytology identifications shown that hybrid plants were obtained.

关 键 词：甘蓝型油菜 紫罗兰 原生质体 体细胞杂交 植株再生 

分 类 号：S565.403.53[农业科学—作物学]