Characterization of bacterial operons consisting of two tubulins and a kinesin-like gene by the novel Two-Step Gene Walking method.
Characterization of bacterial operons consisting of two tubulins and a kinesin-like gene by the novel Two-Step Gene Walking method.
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DOI：
10.1093/nar/gkm836
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发表时间：
2007
影响因子：
14.9
通讯作者：
Petroni G
中科院分区：
文献类型：
--
作者：
Pilhofer M;Bauer AP;Schrallhammer M;Richter L;Ludwig W;Schleifer KH;Petroni G
微管蛋白仍被认为是真核生物的典型蛋白质。然而，最近在不寻常的细菌Prostheclobulin（btubAB）中发现了它们。在这项研究中，btub基因的基因组组织和他们的基因组环境的特点，使用新开发的两步基因步行方法。在所有研究的原杆菌中，btubAB被组织在典型的细菌操纵子中。引人注目的是，所有btub操纵子都包含与驱动蛋白轻链序列相似的第三个基因。所表征的btub操纵子的基因组环境总是不同的。这支持了这组基因代表一个独立的功能单位的假设，这是通过水平基因转移获得的。新开发的两步基因步移法是基于随机引物聚合酶链反应（PCR）。它提出了一个简单的工作流程，其中只包括两个主要步骤-一个步行PCR与一个单一的特异性向外指向引物（步骤1）和直接测序的产物使用巢式特异性引物（步骤2）。两步基因步移被证明是高效的，并被成功地用于表征超过20 kb的序列，不仅在纯培养物中，甚至在复杂的非纯培养物样品中。
Tubulins are still considered as typical proteins of Eukaryotes. However, more recently they have been found in the unusual bacteria Prosthecobacter (btubAB). In this study, the genomic organization of the btub-genes and their genomic environment were characterized by using the newly developed Two-Step Gene Walking method. In all investigated Prosthecobacters, btubAB are organized in a typical bacterial operon. Strikingly, all btub-operons comprise a third gene with similarities to kinesin light chain sequences. The genomic environments of the characterized btub-operons are always different. This supports the hypothesis that this group of genes represents an independent functional unit, which was acquired by Prosthecobacter via horizontal gene transfer. The newly developed Two-Step Gene Walking method is based on randomly primed polymerase chain reaction (PCR). It presents a simple workflow, which comprises only two major steps—a Walking-PCR with a single specific outward pointing primer (step 1) and the direct sequencing of its product using a nested specific primer (step 2). Two-Step Gene Walking proved to be highly efficient and was successfully used to characterize over 20 kb of sequence not only in pure culture but even in complex non-pure culture samples.
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影响因子：
14.9
作者：
PARKS, CL;CHANG, LS;SHENK, T
通讯作者：
SHENK, T
影响因子：
9.4
作者：
Drancourt, M;Carlioz, A;Raoult, D
通讯作者：
Raoult, D
影响因子：
2.7
作者：
Mishra, RN;Singla-Pareek, SL;Reddy, MK
通讯作者：
Reddy, MK
影响因子：
2.2
作者：
Nthangeni, MB;Ramagoma, F;Litthauer, D
通讯作者：
Litthauer, D
DOI：
10.1073/pnas.012516899
发表时间：
2002-12-24
影响因子：
11.1
作者：
Jenkins, C;Samudrala, R;Staley, JT
通讯作者：
Staley, JT