Cargando…
Efficient and Rapid C. elegans Transgenesis by Bombardment and Hygromycin B Selection
We report a simple, cost-effective, scalable and efficient method for creating transgenic Caenorhabditis elegans that requires minimal hands-on time. The method combines biolistic bombardment with selection for transgenics that bear a hygromycin B resistance gene on agar plates supplemented with hyg...
Autores principales: | , , |
---|---|
Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Public Library of Science
2013
|
Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3794024/ https://www.ncbi.nlm.nih.gov/pubmed/24130756 http://dx.doi.org/10.1371/journal.pone.0076019 |
_version_ | 1782287166765793280 |
---|---|
author | Radman, Inja Greiss, Sebastian Chin, Jason W. |
author_facet | Radman, Inja Greiss, Sebastian Chin, Jason W. |
author_sort | Radman, Inja |
collection | PubMed |
description | We report a simple, cost-effective, scalable and efficient method for creating transgenic Caenorhabditis elegans that requires minimal hands-on time. The method combines biolistic bombardment with selection for transgenics that bear a hygromycin B resistance gene on agar plates supplemented with hygromycin B, taking advantage of our observation that hygromycin B is sufficient to kill wild-type C. elegans at very low concentrations. Crucially, the method provides substantial improvements in the success of bombardments for isolating transmitting strains, the isolation of multiple independent strains, and the isolation of integrated strains: 100% of bombardments in a large data set yielded transgenics; 10 or more independent strains were isolated from 84% of bombardments, and up to 28 independent strains were isolated from a single bombardment; 82% of bombardments yielded stably transmitting integrated lines with most yielding multiple integrated lines. We anticipate that the selection will be widely adopted for C. elegans transgenesis via bombardment, and that hygromycin B resistance will be adopted as a marker in other approaches for manipulating, introducing or deleting DNA in C. elegans. |
format | Online Article Text |
id | pubmed-3794024 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2013 |
publisher | Public Library of Science |
record_format | MEDLINE/PubMed |
spelling | pubmed-37940242013-10-15 Efficient and Rapid C. elegans Transgenesis by Bombardment and Hygromycin B Selection Radman, Inja Greiss, Sebastian Chin, Jason W. PLoS One Research Article We report a simple, cost-effective, scalable and efficient method for creating transgenic Caenorhabditis elegans that requires minimal hands-on time. The method combines biolistic bombardment with selection for transgenics that bear a hygromycin B resistance gene on agar plates supplemented with hygromycin B, taking advantage of our observation that hygromycin B is sufficient to kill wild-type C. elegans at very low concentrations. Crucially, the method provides substantial improvements in the success of bombardments for isolating transmitting strains, the isolation of multiple independent strains, and the isolation of integrated strains: 100% of bombardments in a large data set yielded transgenics; 10 or more independent strains were isolated from 84% of bombardments, and up to 28 independent strains were isolated from a single bombardment; 82% of bombardments yielded stably transmitting integrated lines with most yielding multiple integrated lines. We anticipate that the selection will be widely adopted for C. elegans transgenesis via bombardment, and that hygromycin B resistance will be adopted as a marker in other approaches for manipulating, introducing or deleting DNA in C. elegans. Public Library of Science 2013-10-09 /pmc/articles/PMC3794024/ /pubmed/24130756 http://dx.doi.org/10.1371/journal.pone.0076019 Text en © 2013 Radman et al http://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are properly credited. |
spellingShingle | Research Article Radman, Inja Greiss, Sebastian Chin, Jason W. Efficient and Rapid C. elegans Transgenesis by Bombardment and Hygromycin B Selection |
title | Efficient and Rapid C. elegans Transgenesis by Bombardment and Hygromycin B Selection |
title_full | Efficient and Rapid C. elegans Transgenesis by Bombardment and Hygromycin B Selection |
title_fullStr | Efficient and Rapid C. elegans Transgenesis by Bombardment and Hygromycin B Selection |
title_full_unstemmed | Efficient and Rapid C. elegans Transgenesis by Bombardment and Hygromycin B Selection |
title_short | Efficient and Rapid C. elegans Transgenesis by Bombardment and Hygromycin B Selection |
title_sort | efficient and rapid c. elegans transgenesis by bombardment and hygromycin b selection |
topic | Research Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3794024/ https://www.ncbi.nlm.nih.gov/pubmed/24130756 http://dx.doi.org/10.1371/journal.pone.0076019 |
work_keys_str_mv | AT radmaninja efficientandrapidceleganstransgenesisbybombardmentandhygromycinbselection AT greisssebastian efficientandrapidceleganstransgenesisbybombardmentandhygromycinbselection AT chinjasonw efficientandrapidceleganstransgenesisbybombardmentandhygromycinbselection |