| First Author | Nakahara M | Year | 2013 |
| Journal | Mamm Genome | Volume | 24 |
| Issue | 5-6 | Pages | 228-39 |
| PubMed ID | 23604909 | Mgi Jnum | J:198500 |
| Mgi Id | MGI:5496945 | Doi | 10.1007/s00335-013-9452-4 |
| Citation | Nakahara M, et al. (2013) Gene-trap mutagenesis using Mol/MSM-1 embryonic stem cells from MSM/Ms mice. Mamm Genome 24(5-6):228-39 |
| abstractText | The MSM/Ms strain is derived from the Japanese wild mouse Mus musculus molossinus and displays characteristics not observed in common laboratory strains. Functional genomic analyses using genetically engineered MSM/Ms mice will reveal novel phenotypes and gene functions/interactions. We previously reported the establishment of a germline-competent embryonic stem (ES) cell line, Mol/MSM-1, from the MSM/Ms strain. To analyze its usefulness for insertional mutagenesis, we performed gene-trapping using these cells. In the present study, we compared the gene-trap events between Mol/MSM-1 and a conventional ES cell line, KTPU8, derived from the F1 progeny of a C57BL/6 x CBA cross. We introduced a promoter-trap vector carrying the promoterless beta-galactosidase/neomycin-resistance fusion gene into Mol/MSM-1 and KTPU8 cells, isolated clones, and identified the trapped genes by rapid amplification of cDNA 5'-ends (5'-RACE), inverse PCR, or plasmid rescue. Unexpectedly, the success rate of 5'-RACE in Mol/MSM trap clones was 47 %, lower than the 87 % observed in KTPU8 clones. Genomic analysis of the 5'-RACE-failed clones revealed that most had trapped ribosomal RNA gene regions. The percentage of ribosomal RNA region trap clones was 41 % in Mol/MSM-1 cells, but less than 10 % in KTPU8 cells. However, within the Mol/MSM-1 5'-RACE-successful clones, the trapping frequency of annotated genes, the chromosomal distribution of vector insertions, the frequency of integration into an intron around the start codon-containing exon, and the functional spectrum of trapped genes were comparable to those in KTPU8 cells. By selecting 5'-RACE-successful clones, it is possible to perform gene-trapping efficiently using Mol/MSM-1 ES cells and promoter-trap vectors. |
