To determine the genetic relatedness of the isolates, pulsed-field gel electrophoresis (PFGE) was used, as described by Benson et al. [38 (link)]. The complete bacterial genome was embedded in agarose plugs and lysed in several steps (with lysozyme, mutanolysin and proteinase K) to purify DNA. For the digestion, Sma I restriction enzyme was used (3 h, 25 °C). The digested samples were run in a 1% agarose gel along with N0340S Lambda Ladder PFG Marker (New England Biolabs, Hitchin, Hertfordshire, UK), in a Bio-Rad CHEF-DR® II PFGE machine, for 21 h at 14 °C, with the following pulse times: block 1, 5 s/15 s for 10 h, and block 2, 15 s/60 s for 11 h. After the gel image was captured, dendrograms were created by the Fingerprinting II software (Bio-Rad, Marnes-la-Coquette, France) and the PFGE patterns were analysed according to the criteria of van Belkum et al. [39 (link)].
N0340s lambda ladder pfg marker
Manufactured by New England Biolabs
Sourced in United Kingdom
The N0340S Lambda Ladder PFG Marker is a DNA size standard used for estimating the molecular weight of DNA fragments in pulsed-field gel electrophoresis (PFGE) applications. It consists of DNA fragments ranging from 48.5 to 1,018.5 kilobase pairs (kb) in size, providing a reference for determining the size of high molecular weight DNA samples.
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Lab products found in correlation
2 protocols using n0340s lambda ladder pfg marker
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Pulsed-Field Gel Electrophoresis for Bacterial Genotyping
2
Genetic Relatedness Analysis by PFGE
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To determine the genetic relatedness of the isolates, pulsed-field gel electrophoresis (PFGE) was used as described earlier [14] . The complete bacterial genome was embedded in agarose plugs and lysed in several steps to purify DNA. For the digestion, Sma I restriction enzyme was used (3 h, 25 °C). The digested samples were run in a 1% agarose gel along with N0340S Lambda Ladder PFG Marker (New England Biolabs, Hitchin, Hertfordshire, UK), in a Bio-Rad CHEF-DR ® II PFGE machine, for 21 h at 14 °C, with the following pulse times: block one -5 s/15 s for 10 h, and block two -15 s/60 s for 11 h. After the gel image was captured, the PFGE patterns were analyzed and the dendrograms were created by the Fingerprinting II software (Bio-Rad).
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