Clonal analysis of heterogeneous crown gall tumor tissues induced by wild-type and shooter mutant strains ofAgrobacterium tumefaciens-expression of T-DNA genes-猫眼课题宝

根癌农杆菌野生型和射手突变株诱导异质冠瘿瘤组织的克隆分析-T-DNA基因表达

DOI：

发表时间：

1983

期刊：

Plant Molecular Biology

影响因子：

5.1

通讯作者：

中科院分区：

生物学2区

文献类型：

作者：

G. van Slogteren;J. H. C. Hoge;P. Hooykaas;R. Schilperoort

肿瘤是由一株含有野生型章鱼素钛的农杆菌菌株和在T区生长素基因座上插入转座子的突变体诱导的。分离的无菌肿瘤组织的克隆显示，在所有病例中，肿瘤细胞(10-26%)与大多数正常细胞相邻。用野生型钛质粒菌株诱导的肿瘤克隆均为无定形愈伤组织。由枪手突变体诱导的肿瘤克隆有两种不同的类型。其中一种类型的无性系在不含激素的培养基上生长良好。这种类型总是在这种培养基上再生出大量的肿瘤芽。另一种克隆只有在添加生长素和细胞分裂素后才能生长，但在单独添加生长素时也会出现生长缓慢的现象。这种类型的植株从来不会自发再生。然而，在高水平激动素的刺激下，这些克隆也获得了一些新芽。其中一个与其他肿瘤枝条一样，含有肿瘤特异性酶OCS(octopinesynthase，OCS)，但与其他肿瘤枝条不同的是，它形成了一个根系。利用Northern杂交技术对2号进行了研究。除tr.2外，在所分析的肿瘤枝条中均检测到T-DNA转录本，包括生长素基因转录本tr.1和细胞分裂素基因转录本tr.4。这表明，这些转录本的存在不会干扰芽细胞的分化。其中一个肉瘤克隆(TSO38)在章鱼碱合成酶活性(OCS±)方面表现出不稳定的特征。对于TSO38及其部分亚无性系，只有4%的再生芽是OCS+。Northern印迹杂交结果表明，章鱼碱合成酶基因在TSO38衍生植株群体中的表达水平极低，证明了章鱼碱合成酶基因存在于植株的−中，并且该基因在转录水平上受转录水平的调控而表现出不稳定的表达。TSO38及其亚克隆TSO38-23−均为OCS+。

SummaryTumors were induced by anAgrobacterium tumefaciens strain with a wild-type octopine Ti plasmid and by shooter mutants with a transposon insertion in the auxin-locus of the T-region. Cloning of isolated axenic tumor tissues revealed that in all cases they consisted of tumor cells (10–26%) next to a majority of normal cells. The tumor clones that had been induced by the strain with the wild-type Ti plasmid all grew as amorphous calli. Tumor, clones induced by a shooter mutant were of two different types. One type of clone grew well on phytohormone-free medium. this type invariably regenerated tumorous shoots abundantly on this medium. The other type of clone only grew after the addition of auxin and cytokinin to the culture medium, but slow growth also took place in the presence of only auxin. This type never regenerated shoots spontaneously. After stimulation by a high level of kinetin, however, a few shoots were also obtained from these clones. One of these shoots, like other tumorous shoots, contained the tumor-specific enzyme octopinesynthase (Ocs), but in contrast to other tumorous shoots formed a root-system.The expression of T-DNA genes in shoots proliferating from the cloned tumor tissues induced by a mutant with an insertion in the region for transcript tr. 2 was studied by northern blot hybridization. Except for tr.2 the T-DNA transcripts were detected in the tumorous shoots analysed, including the transcript, tr.1 from the auxin-locus and tr.4 from the cytokinin-locus. This shows that the presence of these transcripts, which are assumed to be responsible for the tumorigenic character of tumor cells, does not interfere with the differentiation of shoot cells.One of the shooty tumor clones (TSO38) showed an unstable character with regard to octopine synthase activity (Ocs±). For, TSO38 and some of its subclones, it was found that only 4% of the regenerated shoots were Ocs+. Northern blot hybridization revealed that the mRNA for octopine synthase was present in extremely low quantity in the population of TSO38 derived shoots.The finding that it was possible to force shoots from clone TSO38 and from subclone TSO38-23− that were Ocs− to become Ocs+, proved that the gene for octopine synthase was present in the Ocs− shoots and that this gene showed unstable expression due to regulation at the level of transcription.

Sizes and map positions of several plasmid-DNA-encoded transcripts in octopine-type crown gall tumors.