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. 2015 Sep 25;4:e10565. doi: 10.7554/eLife.10565

Figure 7. Structure-function analysis of PGBD5-induced DNA transposition using alanine scanning mutagenesis.

(A) Quantitative PCR analysis of genomic integration activity of alanine point mutants of GFP-PGBD5, as compared to wild-type and GFP control-expressing cells. D168A, D194A, and D386A mutants (red) exhibit significant reduction in apparent activity (Asterisks denote statistical significance: p = 0.00011, p = 0.000021, p = 0.000013 vs GFP-PGBD5, respectively). Dotted line marks threshold at which less than 1 transposon copy was detected per haploid human genome. Error bars represent standard errors of the mean of 3 biological replicates. (B) Western immunoblot showing equal expression of GFP-PGBD5 mutants, as compared to wild-type GFP-PGBD5 (green). β-actin (red) serves as loading control.

DOI: http://dx.doi.org/10.7554/eLife.10565.019

Figure 7.

Figure 7—figure supplement 1. Sanger sequencing trances of pRecLV103-GFP-PGBD5 D>A and E>A mutants (D168A, D175A, E188A, D192A, D194A, E203A, E205A, E236A).

Figure 7—figure supplement 1.

DOI: http://dx.doi.org/10.7554/eLife.10565.020

Figure 7—figure supplement 2. Sanger sequencing trances of pRecLV103-GFP-PGBD5 D>A and E>A mutants (D241A, D244A, E284A, E285A, E287A, D303A, E365A, E373A).

Figure 7—figure supplement 2.

DOI: http://dx.doi.org/10.7554/eLife.10565.021

Figure 7—figure supplement 3. Sanger sequencing trances of pRecLV103-GFP-PGBD5 D>A and E>A mutants (D386A, D387A, D425A, E439A, E444A, E449A, D450A).

Figure 7—figure supplement 3.

DOI: http://dx.doi.org/10.7554/eLife.10565.022