LOCALIZATION OF AGROBACTERIUM-RHIZOGENES T-DNA IN PLANT CHROMOSOMES BY INSITU HYBRIDIZATION
LOCALIZATION OF AGROBACTERIUM-RHIZOGENES T-DNA IN PLANT CHROMOSOMES BY INSITU HYBRIDIZATION
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DOI：
10.1002/j.1460-2075.1986.tb04468.x
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发表时间：
1986-09-01
期刊：
影响因子：
11.4
通讯作者：
MATZKE, MA
中科院分区：
文献类型：
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作者：
AMBROS, PF;MATZKE, AJM;MATZKE, MA
我们使用原位杂交来确定发根农杆菌 Ri T-DNA 在 Crepis capillaris (2n = 6) 转化根的染色体中的插入位点。通过用发根农杆菌感染 Crepis 茎段获得了四个转化的根系。 Southern 杂交分析表明，每个根系都是一个或多个独立 T-DNA 插入事件的结果。在两个根系中，存在一份 T-DNA 拷贝；另外两个根系各含有两个 T-DNA 拷贝。为了将这些 T-DNA 插入片段定位在 Crepis 染色体上，从每个根系制备中期扩散，并与生物素标记的 T-DNA 探针原位杂交。结果表明，T-DNA 存在于每个根系的不同染色体位置，并且每个染色体都至少成为 T-DNA 插入的目标一次。在含有两个 T-DNA 插入片段的根系中，观察到两种整合模式：在一种情况下，T-DNA 存在于单独的染色体上；在另一种情况下，T-DNA 存在于不同的染色体上。在另一种情况下，两个 T-DNA 在一条染色体上紧密相连（但不是串联排列）。将这些结果与之前获得的第五个 Crepis 转化根系的结果进行比较表明，Ri T-DNA 不会优先插入特定的染色体位置。
We have used in situ hybridization to determine the sites of insertion of Agrobacterium rhizogenes Ri T-DNA in the chromosomes of Crepis capillaris (2n = 6) transformed roots. Four transformed root lines were obtained by infecting Crepis stem segments with A. rhizogenes. Southern hybridization analysis indicated that each root line was the result of one or more independent T-DNA insertion events. In two root lines, one copy of T-DNA was present; the other two root lines each contained two copies of T-DNA. To localize these T-DNA inserts on Crepis chromosomes, metaphase spreads were prepared from each root line, and hybridized in situ to a biotin-labeled T-DNA probe. The results indicated that T-DNA was present in a different chromosomal location in each root line, and that each chromosome had been a target for T-DNA insertion at least once. In the root lines containing two T-DNA inserts, two patterns of integration were observed: in one case the T-DNAs were present on separate chromosomes; in the other case the two T-DNAs were close together (but not tandemly arranged) on a single chromosome. A comparison of these results and those obtained previously for a fifth Crepis-transformed root line demonstrated that Ri T-DNA does not insert preferentially into a particular chromosomal location.