MF-Millipore Membrane Filter, mixed cellulose esters, Hydrophilic, 0.22 µm, 25 mm, white, plain

Company: EMD Millipore

Catalog#: GSWP02500

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A Quick DNA Extraction Method for High Throughput Screening in Gram-positive Bacteria

Nuo Chen and Xiaoming Yuan

Published online: 2023-04-20

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Cell-to-cell DNA Transfer among Thermus Species

Author:

Alba Blesa and José Berenguer,

Date:

2016-11-20

[Abstract] The ability to transfer DNA via direct cell-to-cell contact-dependent process similar to conjugation has been described in Thermus thermophilus (Tth). Here, we detail the mating experiment protocol involving the lateral transfer of thermostable antibiotic resistance markers (i.e., kanamycin: Km^R; hygromycin: Hyg^R) between Thermus cells, enabling the selection and quantification of the transfer frequencies. Briefly, liquid cultures of both mates are mixed and laid onto a nitrocellulose filter on a TB plate. After incubation at 60 °C, filters are resuspended upon selective plating. The contribution of DNA uptake by transformation is abolished by the addition of DNase I to the mix. This protocol can be used for the transfer of large DNA ... [摘要] 通过与共轭相似的直接细胞间接触依赖性过程转移DNA的能力已在Thermus thermophilus（Tth）中描述。在这里，我们详细描述了包括热稳定抗生素抗性标记（即卡那霉素：Km），潮霉素：Hyg ^R的横向转移的交配实验方案>）在细胞之间，使得能够选择和量化转移频率。简言之，将两种配合物的液体培养物混合并铺在TB板上的硝酸纤维素滤膜上。在60℃温育后，在选择性平板培养基上重悬浮过滤器。通过向混合物中加入DNA酶I消除通过转化的DNA摄取的贡献。该方案可用于将大DNA片段（> 10kb）转移到Thermus 物种。

[背景] 是大多数细菌水平基因转移的主要机制，是一种高度专业化的过程，通过其DNA在直接接触的两个细胞之间转移（Lederberg和Tatum，1946）。经典缀合涉及DNA分子（通常是质粒编码的）从供体到受体细胞的单向转移，其仍然是被动的。然而，已经描述了用于多种细菌的替代模型。在实验室中，缀合实验对于标志物交换诱变是有效的。在其它遗传转移方法中，缀合在细菌包膜的最小破坏和转移大DNA片段（包括大的染色体区域（> 10kb））的可行性方面是有利的。在T之间的共轭样过程。嗜热细菌在十年前被报道，其中染色体标记在重组HFR - ...

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MF-Millipore Membrane Filter, mixed cellulose esters, Hydrophilic, 0.22 µm, 25 mm, white, plain

MF-Millipore Membrane Filter, mixed cellulose esters, Hydrophilic, 0.22 µm, 25 mm, white, plain

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