Optimization of a large-scale gene disruption protocol in <it>Dictyostelium </it>and analysis of conserved genes of unknown function
<p>Abstract</p> <p>Background</p> <p>Development of the post-genomic age in <it>Dictyostelium </it>will require the existence of rapid and reliable methods to disrupt genes that would allow the analysis of entire gene families and perhaps the possibility to...
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| Format: | Article |
| Language: | English |
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BMC
2006-08-01
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| Series: | BMC Microbiology |
| Online Access: | http://www.biomedcentral.com/1471-2180/6/75 |
| _version_ | 1811297381311840256 |
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| author | Escalante Ricardo Robles Alicia Torija Patricia |
| author_facet | Escalante Ricardo Robles Alicia Torija Patricia |
| author_sort | Escalante Ricardo |
| collection | DOAJ |
| description | <p>Abstract</p> <p>Background</p> <p>Development of the post-genomic age in <it>Dictyostelium </it>will require the existence of rapid and reliable methods to disrupt genes that would allow the analysis of entire gene families and perhaps the possibility to undertake the complete knock-out analysis of all the protein-coding genes present in <it>Dictyostelium </it>genome.</p> <p>Results</p> <p>Here we present an optimized protocol based on the previously described construction of gene disruption vectors by in vitro transposition. Our method allows a rapid selection of the construct by a simple PCR approach and subsequent sequencing. Disruption constructs were amplified by PCR and the products were directly transformed in <it>Dictyostelium </it>cells. The selection of homologous recombination events was also performed by PCR. We have constructed 41 disruption vectors to target genes of unknown function, highly conserved between <it>Dictyostelium </it>and human, but absent from the genomes of <it>S. cerevisiae </it>and <it>S. pombe</it>. 28 genes were successfully disrupted.</p> <p>Conclusion</p> <p>This is the first step towards the understanding of the function of these conserved genes and exemplifies the easiness to undertake large-scale disruption analysis in <it>Dictyostelium</it>.</p> |
| first_indexed | 2024-04-13T06:03:21Z |
| format | Article |
| id | doaj.art-98adfb9aadba410ebfcf586551d5af2e |
| institution | Directory Open Access Journal |
| issn | 1471-2180 |
| language | English |
| last_indexed | 2024-04-13T06:03:21Z |
| publishDate | 2006-08-01 |
| publisher | BMC |
| record_format | Article |
| series | BMC Microbiology |
| spelling | doaj.art-98adfb9aadba410ebfcf586551d5af2e2022-12-22T02:59:22ZengBMCBMC Microbiology1471-21802006-08-01617510.1186/1471-2180-6-75Optimization of a large-scale gene disruption protocol in <it>Dictyostelium </it>and analysis of conserved genes of unknown functionEscalante RicardoRobles AliciaTorija Patricia<p>Abstract</p> <p>Background</p> <p>Development of the post-genomic age in <it>Dictyostelium </it>will require the existence of rapid and reliable methods to disrupt genes that would allow the analysis of entire gene families and perhaps the possibility to undertake the complete knock-out analysis of all the protein-coding genes present in <it>Dictyostelium </it>genome.</p> <p>Results</p> <p>Here we present an optimized protocol based on the previously described construction of gene disruption vectors by in vitro transposition. Our method allows a rapid selection of the construct by a simple PCR approach and subsequent sequencing. Disruption constructs were amplified by PCR and the products were directly transformed in <it>Dictyostelium </it>cells. The selection of homologous recombination events was also performed by PCR. We have constructed 41 disruption vectors to target genes of unknown function, highly conserved between <it>Dictyostelium </it>and human, but absent from the genomes of <it>S. cerevisiae </it>and <it>S. pombe</it>. 28 genes were successfully disrupted.</p> <p>Conclusion</p> <p>This is the first step towards the understanding of the function of these conserved genes and exemplifies the easiness to undertake large-scale disruption analysis in <it>Dictyostelium</it>.</p>http://www.biomedcentral.com/1471-2180/6/75 |
| spellingShingle | Escalante Ricardo Robles Alicia Torija Patricia Optimization of a large-scale gene disruption protocol in <it>Dictyostelium </it>and analysis of conserved genes of unknown function BMC Microbiology |
| title | Optimization of a large-scale gene disruption protocol in <it>Dictyostelium </it>and analysis of conserved genes of unknown function |
| title_full | Optimization of a large-scale gene disruption protocol in <it>Dictyostelium </it>and analysis of conserved genes of unknown function |
| title_fullStr | Optimization of a large-scale gene disruption protocol in <it>Dictyostelium </it>and analysis of conserved genes of unknown function |
| title_full_unstemmed | Optimization of a large-scale gene disruption protocol in <it>Dictyostelium </it>and analysis of conserved genes of unknown function |
| title_short | Optimization of a large-scale gene disruption protocol in <it>Dictyostelium </it>and analysis of conserved genes of unknown function |
| title_sort | optimization of a large scale gene disruption protocol in it dictyostelium it and analysis of conserved genes of unknown function |
| url | http://www.biomedcentral.com/1471-2180/6/75 |
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