Effect of Bacterial Distribution and Activity on Conjugal Gene Transfer on the Phylloplane of the Bush Bean (Phaseolus vulgaris)
Effect of Bacterial Distribution and Activity on Conjugal Gene Transfer on the Phylloplane of the Bush Bean (Phaseolus vulgaris)
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细菌分布和活性对布什豆（菜豆）叶面接合基因转移的影响
DOI：
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发表时间：
1998
影响因子：
4.4
通讯作者：
N. Kroer
中科院分区：
文献类型：
--
作者：
B. Normander;B. Christensen;S. Molin;N. Kroer
本文研究了菜豆叶平面上的接合质粒转移，并探讨了接合质粒转移与菜豆叶平面上细菌空间分布和代谢活性的关系。供体（恶臭假单胞菌KT 2442）含有TOL质粒的衍生物，该衍生物赋予卡那霉素抗性，并将gfp基因插入lac启动子的下游。染色体插入lacIq阻止了gfp基因的表达。受体（恶臭假单胞菌KT 2440）具有染色体四环素抗性标记。因此，可以通过平板接种计数接合子，并在原位观察为绿色荧光细胞。用供体和受体以约105个细胞/cm 2的密度接种无菌菜豆幼苗。为了操纵接种细菌的密度和代谢活性（通过掺入[3 H]亮氨酸测量），使植物在各种相对湿度（RH）下生长。在100%RH下，接合子的密度达到3 × 103 CFU/cm 2，相当于受体群体的约三分之一。在25%RH下，接合子的数量低于检测限。接种到叶片上后，接种物的每细胞代谢活性立即增加了8倍（100%RH），然后在96小时后降低到初始水平。细菌的代谢活性对接合没有速率限制，并且未观察到两个参数之间的相关性。显然，叶片分泌物确保了细菌的活性高于发生转移的阈值。Transconjugants主要观察到表皮细胞之间的交界处和气孔下腔。接合子的分布与非不育叶片上土著细菌的分布相似。与聚碳酸酯过滤器相比，细胞密度等于叶片上的总密度，叶片上的转移率高达30倍。因此，细菌聚集到叶面上的微生境中对转移有很大的刺激作用。
ABSTRACT Conjugal plasmid transfer was examined on the phylloplane of bean (Phaseolus vulgaris) and related to the spatial distribution pattern and metabolic activity of the bacteria. The donor (Pseudomonas putida KT2442) harbored a derivative of the TOL plasmid, which conferred kanamycin resistance and had thegfp gene inserted downstream of a lac promoter. A chromosomal insertion of lacIq prevented expression of the gfp gene. The recipient (P. putida KT2440) had a chromosomal tetracycline resistance marker. Thus, transconjugants could be enumerated by plating and visualized in situ as green fluorescent cells. Sterile bean seedlings were inoculated with donors and recipients at densities of approximately 105 cells per cm2. To manipulate the density and metabolic activity (measured by incorporation of [3H]leucine) of the inoculated bacteria, plants were grown at various relative humidities (RH). At 100% RH, the transconjugants reached a density of 3 × 103CFU/cm2, corresponding to about one-third of the recipient population. At 25% RH, numbers of transconjugants were below the detection limit. Immediately after inoculation onto the leaves, the per-cell metabolic activity of the inocula increased by up to eight times (100% RH), followed by a decrease to the initial level after 96 h. The metabolic activity of the bacteria was not rate limiting for conjugation, and no correlation between the two parameters was observed. Apparently, leaf exudates insured that the activity of the bacteria was above a threshold value for transfer to occur. Transconjugants were primarily observed in junctures between epidermal cells and in substomatal cavities. The distribution of the transconjugants was similar to the distribution of indigenous bacteria on nonsterile leaves. Compared to polycarbonate filters, with cell densities equal to the overall density on the leaves, transfer ratios on leaves were up to 30 times higher. Thus, aggregation of the bacteria into microhabitats on the phylloplane had a great stimulatory effect on transfer.
影响因子：
3.5
作者：
Cormack, BP;Valdivia, RH;Falkow, S
通讯作者：
Falkow, S