Biotechnology Centre, Indian Agricultural Research Institute, 10012, New Delhi, India.
Plant Cell Rep. 1992 Jul;11(8):428-32. doi: 10.1007/BF00234376.
Brassica carinata (2n=34, BBCC), was synthesized by fusing dark grown etiolated hypocotyl protoplasts of B. nigra (2n=16, BB) with green mesophyll protoplasts of B. oleracea (2n=18,CC) using polyethylene glycol. Heterokaryons could be microscopically distinguished from the parental types by their dark green chloroplasts in the colourless hypocotyl protoplast background. The mean heterokaryotic fusion frequency estimated on the basis of this morphological distinction was about 16%. A total of 626 calli were obtained, of which 92 regenerated shoots after transfer to zeatin (2 mg/l) supplemented MS medium. Of these, 81 calli differentiated into plants morphologically similar to naturally occurring B. carinata and 11 calli yielded plants resembling parental types. Meiosis in seven hybrid plants showed the chromosome number to be 2n=34 the sum of B. nigra and B. oleracea chromosomes. Molecular confirmation of the amphidiploid nature of hybrids was obtained by probing with a B. juncea derived genomic clone. The use of chloroplast and mitochondrial specific gene probes, revealed that one plant was a cytoplasmic hybrid having cp DNA sequences of both B. oleracea and B. nigra and mt DNA sequences of B. nigra.
芸薹属黑芥（2n=16, BB）暗生长的下胚轴原生质体与芸薹属甘蓝（2n=18, CC）的绿色叶肉原生质体融合，通过聚乙二醇合成了芥菜型油菜（2n=34, BBCC）。杂种细胞可通过其在无色下胚轴原生质体背景中的深绿色叶绿体在显微镜下与亲本类型区分开来。基于这种形态差异估计的杂种异核融合频率约为 16%。共获得 626 个愈伤组织，其中 92 个在添加 zeatin（2 mg/l）的 MS 培养基上再生出芽。其中，81 个愈伤组织分化成形态上类似于自然芥菜型油菜的植株，11 个愈伤组织产生的植株类似于亲本类型。7 株杂种植物的减数分裂显示染色体数为 2n=34，即黑芥和甘蓝染色体的总和。通过用一个来自芥菜型油菜的基因组克隆进行探测，获得了杂种的双二倍体性质的分子证实。使用叶绿体和线粒体特异性基因探针，揭示了一株植物是质体杂种，具有甘蓝和黑芥的 cpDNA 序列和黑芥的 mtDNA 序列。
