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. 2008 Oct;18(10):1670-9.
doi: 10.1101/gr.078352.108. Epub 2008 Sep 17.

Large-scale gene trapping in C57BL/6N mouse embryonic stem cells

Gwenn M Hansen  1 , Diane C Markesich, Michael B Burnett, Qichao Zhu, Karen M Dionne, Lizabeth J Richter, Richard H Finnell, Arthur T Sands, Brian P Zambrowicz, Alejandro Abuin
Affiliations

Large-scale gene trapping in C57BL/6N mouse embryonic stem cells

Gwenn M Hansen et al. Genome Res. 2008 Oct.

Abstract

We report the construction and analysis of a mouse gene trap mutant resource created in the C57BL/6N genetic background containing more than 350,000 sequence-tagged embryonic stem (ES) cell clones. We also demonstrate the ability of these ES cell clones to contribute to the germline and produce knockout mice. Each mutant clone is identified by a genomic sequence tag representing the exact insertion location, allowing accurate prediction of mutagenicity and enabling direct genotyping of mutant alleles. Mutations have been identified in more than 10,000 genes and show a bias toward the first intron. The trapped ES cell lines, which can be requested from the Texas A&M Institute for Genomic Medicine, are readily available to the scientific community.

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Figures

Figure 1.
Figure 1.
OmniBankII gene acquisition and chromosomal distribution. (A) Gene acquisition rate over the course of OmniBankII production, shown as a function of the number of clones processed for sequence acquisition. For comparison, a plot of OmniBank gene acquisition (Zambrowicz et al. 2003) is shown. (B) Frequency of gene trap acquisition across the mouse genome. Frequency is calculated as a percentage of RefSeq genes trapped in each library. (C) Correlation between chromosomal gene content and gene acquisition for OmniBankII. RefSeq genes mapped to the random contig set were excluded from analysis. (D) Density of gene trap mutations per gene for the OmniBankII library.
Figure 2.
Figure 2.
OmniBankII gene trap insertions show first intron bias. (A) All insertions occurring within 10 kb upstream or downstream of a TSS were identified and used to plot the percentage of insertions occurring within 500-bp intervals on either side of the TSS. (B) Graph showing the percentage of total genic insertions occurring within exon 1 (e1) or introns 1 through 20 (i1–i20). The percentage was calculated as a function of total genic insertions, or as a function of genes containing insertions in each category.
Figure 3.
Figure 3.
Symmetrical sequence preferences at OmniBankII gene trap insertion sites. (A–C) Base compositions surrounding each gene trap insertion site are shown according to chromosomal cleavage offset. Insertion occurs between position −1 and 1 on the top strand (with respect to the orientation of the retroviral backbone rather than the gene trap construct components). Confirmed duplicated junction sequences are shown within the solid box. (Arrows) Base positions for DNA strand transfer. Positions showing statistically different base frequencies are highlighted. (Gray) Differences of 10%; differences of >10% (blue, increased frequency; or yellow, decreased frequency). Additional discussion and an expanded version of this table can be found in the Supplemental material and Supplemental Figure 7.
Figure 4.
Figure 4.
Germline transmission rates as a function of percent chimerism. The C57BL/6N derived chimeras approach those of 129S5/SvEvBrd chimeras only at the highest percent contribution.
See this image and copyright information in PMC

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