論文 - 詳細
| RRC ID | 46553 |
|---|---|
| 著者 | Kage-Nakadai E, Kobuna H, Funatsu O, Otori M, Gengyo-Ando K, Yoshina S, Hori S, Mitani S. |
| タイトル | Single/low-copy integration of transgenes in Caenorhabditis elegans using an ultraviolet trimethylpsoralen method. |
| ジャーナル | BMC Biotechnol |
| Abstract |
BACKGROUND:Transgenic strains of Caenorhabditis elegans are typically generated by injecting DNA into the germline to form multi-copy extrachromosomal arrays. These transgenes are semi-stable and their expression is silenced in the germline. Mos1 transposon or microparticle bombardment methods have been developed to create single- or low-copy chromosomal integrated lines. Here we report an alternative method using ultraviolet trimethylpsoralen (UV/TMP) to generate single/low-copy gene integrations. RESULTS:We successfully integrated low-copy transgenes from extrachromosomal arrays using positive selection based on temperature sensitivity with a vps-45 rescue fragment and negative selection based on benzimidazole sensitivity with a ben-1 rescue fragment. We confirmed that the integrants express transgenes in the germline. Quantitative PCR revealed that strains generated by this method contain single- or low-copy transgenes. Moreover, positive selection marker genes flanked by LoxP sites were excised by Cre recombinase mRNA microinjection, demonstrating Cre-mediated chromosomal excision for the first time in C. elegans. CONCLUSION:Our UV/TMP integration method, based on familiar extrachromosomal transgenics, provides a useful approach for generating single/low-copy gene integrations. |
| 巻・号 | 12 |
| ページ | 1 |
| 公開日 | 2012-1-5 |
| DOI | 10.1186/1472-6750-12-1 |
| PII | 1472-6750-12-1 |
| PMID | 22217006 |
| PMC | PMC3262153 |
| MeSH | Animals Animals, Genetically Modified Caenorhabditis elegans / genetics* Gene Dosage Gene Transfer Techniques* Integrases / genetics Trioxsalen Ultraviolet Rays |
| IF | 2.312 |
| 引用数 | 18 |
| WOS 分野 | BIOTECHNOLOGY & APPLIED MICROBIOLOGY |
| リソース情報 | |
| 線虫 | tm234 tm246 |