Enabling low-cost and robust essentiality studies with high-throughput transposon mutagenesis (HTTM).
Transposon-insertion sequencing (TIS) methods couple high density transposon mutagenesis with next-generation sequencing and are commonly used to identify essential or important genes in bacteria. However, this approach can be work-intensive and sometimes expensive depending on the selected protocol...
| Main Authors: | , , , , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
Public Library of Science (PLoS)
2023-01-01
|
| Series: | PLoS ONE |
| Online Access: | https://doi.org/10.1371/journal.pone.0283990 |
| _version_ | 1797843202062942208 |
|---|---|
| author | Antoine Champie Amélie De Grandmaison Simon Jeanneau Frédéric Grenier Pierre-Étienne Jacques Sébastien Rodrigue |
| author_facet | Antoine Champie Amélie De Grandmaison Simon Jeanneau Frédéric Grenier Pierre-Étienne Jacques Sébastien Rodrigue |
| author_sort | Antoine Champie |
| collection | DOAJ |
| description | Transposon-insertion sequencing (TIS) methods couple high density transposon mutagenesis with next-generation sequencing and are commonly used to identify essential or important genes in bacteria. However, this approach can be work-intensive and sometimes expensive depending on the selected protocol. The difficulty to process a high number of samples in parallel using standard TIS protocols often restricts the number of replicates that can be performed and limits the deployment of this technique to large-scale projects studying gene essentiality in various strains or growth conditions. Here, we report the development of a robust and inexpensive High-Throughput Transposon Mutagenesis (HTTM) protocol and validate the method using Escherichia coli strain BW25113, the parental strain of the KEIO collection. HTTM reliably provides high insertion densities with an average of one transposon every ≤20bp along with impressive reproducibility (Spearman correlation coefficients >0.94). A detailed protocol is available at protocol.io and a graphical version is also included with this article. |
| first_indexed | 2024-04-09T17:02:23Z |
| format | Article |
| id | doaj.art-c5f6a846594644809836bcfd0201b143 |
| institution | Directory Open Access Journal |
| issn | 1932-6203 |
| language | English |
| last_indexed | 2024-04-09T17:02:23Z |
| publishDate | 2023-01-01 |
| publisher | Public Library of Science (PLoS) |
| record_format | Article |
| series | PLoS ONE |
| spelling | doaj.art-c5f6a846594644809836bcfd0201b1432023-04-21T05:32:16ZengPublic Library of Science (PLoS)PLoS ONE1932-62032023-01-01184e028399010.1371/journal.pone.0283990Enabling low-cost and robust essentiality studies with high-throughput transposon mutagenesis (HTTM).Antoine ChampieAmélie De GrandmaisonSimon JeanneauFrédéric GrenierPierre-Étienne JacquesSébastien RodrigueTransposon-insertion sequencing (TIS) methods couple high density transposon mutagenesis with next-generation sequencing and are commonly used to identify essential or important genes in bacteria. However, this approach can be work-intensive and sometimes expensive depending on the selected protocol. The difficulty to process a high number of samples in parallel using standard TIS protocols often restricts the number of replicates that can be performed and limits the deployment of this technique to large-scale projects studying gene essentiality in various strains or growth conditions. Here, we report the development of a robust and inexpensive High-Throughput Transposon Mutagenesis (HTTM) protocol and validate the method using Escherichia coli strain BW25113, the parental strain of the KEIO collection. HTTM reliably provides high insertion densities with an average of one transposon every ≤20bp along with impressive reproducibility (Spearman correlation coefficients >0.94). A detailed protocol is available at protocol.io and a graphical version is also included with this article.https://doi.org/10.1371/journal.pone.0283990 |
| spellingShingle | Antoine Champie Amélie De Grandmaison Simon Jeanneau Frédéric Grenier Pierre-Étienne Jacques Sébastien Rodrigue Enabling low-cost and robust essentiality studies with high-throughput transposon mutagenesis (HTTM). PLoS ONE |
| title | Enabling low-cost and robust essentiality studies with high-throughput transposon mutagenesis (HTTM). |
| title_full | Enabling low-cost and robust essentiality studies with high-throughput transposon mutagenesis (HTTM). |
| title_fullStr | Enabling low-cost and robust essentiality studies with high-throughput transposon mutagenesis (HTTM). |
| title_full_unstemmed | Enabling low-cost and robust essentiality studies with high-throughput transposon mutagenesis (HTTM). |
| title_short | Enabling low-cost and robust essentiality studies with high-throughput transposon mutagenesis (HTTM). |
| title_sort | enabling low cost and robust essentiality studies with high throughput transposon mutagenesis httm |
| url | https://doi.org/10.1371/journal.pone.0283990 |
| work_keys_str_mv | AT antoinechampie enablinglowcostandrobustessentialitystudieswithhighthroughputtransposonmutagenesishttm AT ameliedegrandmaison enablinglowcostandrobustessentialitystudieswithhighthroughputtransposonmutagenesishttm AT simonjeanneau enablinglowcostandrobustessentialitystudieswithhighthroughputtransposonmutagenesishttm AT fredericgrenier enablinglowcostandrobustessentialitystudieswithhighthroughputtransposonmutagenesishttm AT pierreetiennejacques enablinglowcostandrobustessentialitystudieswithhighthroughputtransposonmutagenesishttm AT sebastienrodrigue enablinglowcostandrobustessentialitystudieswithhighthroughputtransposonmutagenesishttm |