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. 2011 Nov 29;40(3):e21. doi: 10.1093/nar/gkr908

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© The Author(s) 2011. Published by Oxford University Press.

This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/3.0), which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

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Figure 2. — Isolation of homozygous mutants by selection for copy number increase. (A) Transposition using limiting amounts of transposon donor plasmid results in most G418 resistant subclones having one transposon per cell. (B) Selection scheme, showing predicted number and type of transposon present at each stage. (C) Typical result of double-selection stage. Well 5 has two non-allelic transposon integrations to begin with and thus all cells can become double resistant. (D) Genotyping scheme—primer positions (1–3) are shown relative to the transposon integration site. (E) All double-resistant subclones (derived from an integration in Dhx35) fail to amplify the wild-type PCR product. (F and G) Examples of double-resistant populations in which some (F) and all (G) subclones have a wild-type allele.