Comparative evaluation of different molecular methods for DNA extraction from individual Teladorsagia circumcincta nematodes
Abstract Background The purpose of this study was to develop a reliable DNA extraction protocol to use on individual Teladorsagia circumcincta nematode specimens to produce high quality DNA for genome sequencing and phylogenetic analysis. Pooled samples have been critical in providing the groundwork...
| Main Authors: | , , , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
BMC
2021-05-01
|
| Series: | BMC Biotechnology |
| Subjects: | |
| Online Access: | https://doi.org/10.1186/s12896-021-00695-6 |
| _version_ | 1819081852948840448 |
|---|---|
| author | S. Sloan C. J. Jenvey D. Piedrafita S. Preston M. J. Stear |
| author_facet | S. Sloan C. J. Jenvey D. Piedrafita S. Preston M. J. Stear |
| author_sort | S. Sloan |
| collection | DOAJ |
| description | Abstract Background The purpose of this study was to develop a reliable DNA extraction protocol to use on individual Teladorsagia circumcincta nematode specimens to produce high quality DNA for genome sequencing and phylogenetic analysis. Pooled samples have been critical in providing the groundwork for T. circumcincta genome construction, but there is currently no standard method for extracting high-quality DNA from individual nematodes. 11 extraction kits were compared based on DNA quality, yield, and processing time. Results 11 extraction protocols were compared, and the concentration and purity of the extracted DNA was quantified. Median DNA concentration among all methods measured on NanoDrop 2000™ ranged between 0.45–11.5 ng/μL, and on Qubit™ ranged between undetectable – 0.962 ng/μL. Median A260/280 ranged between 0.505–3.925, and median A260/230 ranged − 0.005 – 1.545. Larval exsheathment to remove the nematode cuticle negatively impacted DNA concentration and purity. Conclusions A Schistosoma sp. DNA extraction method was determined as most suitable for individual T. circumcincta nematode specimens due to its resulting DNA concentration, purity, and relatively fast processing time. |
| first_indexed | 2024-12-21T20:07:22Z |
| format | Article |
| id | doaj.art-1580beb2b82b49b9bc1f23cae3c8ae2c |
| institution | Directory Open Access Journal |
| issn | 1472-6750 |
| language | English |
| last_indexed | 2024-12-21T20:07:22Z |
| publishDate | 2021-05-01 |
| publisher | BMC |
| record_format | Article |
| series | BMC Biotechnology |
| spelling | doaj.art-1580beb2b82b49b9bc1f23cae3c8ae2c2022-12-21T18:51:48ZengBMCBMC Biotechnology1472-67502021-05-0121111310.1186/s12896-021-00695-6Comparative evaluation of different molecular methods for DNA extraction from individual Teladorsagia circumcincta nematodesS. Sloan0C. J. Jenvey1D. Piedrafita2S. Preston3M. J. Stear4AgriBio Centre for AgriBioscience, Department of Animal, Plant and Soil Sciences, School of Life Sciences, La Trobe UniversityAgriBio Centre for AgriBioscience, Department of Animal, Plant and Soil Sciences, School of Life Sciences, La Trobe UniversitySchool of Science, Psychology and Sport, Federation UniversitySchool of Science, Psychology and Sport, Federation UniversityAgriBio Centre for AgriBioscience, Department of Animal, Plant and Soil Sciences, School of Life Sciences, La Trobe UniversityAbstract Background The purpose of this study was to develop a reliable DNA extraction protocol to use on individual Teladorsagia circumcincta nematode specimens to produce high quality DNA for genome sequencing and phylogenetic analysis. Pooled samples have been critical in providing the groundwork for T. circumcincta genome construction, but there is currently no standard method for extracting high-quality DNA from individual nematodes. 11 extraction kits were compared based on DNA quality, yield, and processing time. Results 11 extraction protocols were compared, and the concentration and purity of the extracted DNA was quantified. Median DNA concentration among all methods measured on NanoDrop 2000™ ranged between 0.45–11.5 ng/μL, and on Qubit™ ranged between undetectable – 0.962 ng/μL. Median A260/280 ranged between 0.505–3.925, and median A260/230 ranged − 0.005 – 1.545. Larval exsheathment to remove the nematode cuticle negatively impacted DNA concentration and purity. Conclusions A Schistosoma sp. DNA extraction method was determined as most suitable for individual T. circumcincta nematode specimens due to its resulting DNA concentration, purity, and relatively fast processing time.https://doi.org/10.1186/s12896-021-00695-6DNA isolationTeladorsagia circumcinctaDNA extractionPolymerase chain reactionNematodeGenome sequencing |
| spellingShingle | S. Sloan C. J. Jenvey D. Piedrafita S. Preston M. J. Stear Comparative evaluation of different molecular methods for DNA extraction from individual Teladorsagia circumcincta nematodes BMC Biotechnology DNA isolation Teladorsagia circumcincta DNA extraction Polymerase chain reaction Nematode Genome sequencing |
| title | Comparative evaluation of different molecular methods for DNA extraction from individual Teladorsagia circumcincta nematodes |
| title_full | Comparative evaluation of different molecular methods for DNA extraction from individual Teladorsagia circumcincta nematodes |
| title_fullStr | Comparative evaluation of different molecular methods for DNA extraction from individual Teladorsagia circumcincta nematodes |
| title_full_unstemmed | Comparative evaluation of different molecular methods for DNA extraction from individual Teladorsagia circumcincta nematodes |
| title_short | Comparative evaluation of different molecular methods for DNA extraction from individual Teladorsagia circumcincta nematodes |
| title_sort | comparative evaluation of different molecular methods for dna extraction from individual teladorsagia circumcincta nematodes |
| topic | DNA isolation Teladorsagia circumcincta DNA extraction Polymerase chain reaction Nematode Genome sequencing |
| url | https://doi.org/10.1186/s12896-021-00695-6 |
| work_keys_str_mv | AT ssloan comparativeevaluationofdifferentmolecularmethodsfordnaextractionfromindividualteladorsagiacircumcinctanematodes AT cjjenvey comparativeevaluationofdifferentmolecularmethodsfordnaextractionfromindividualteladorsagiacircumcinctanematodes AT dpiedrafita comparativeevaluationofdifferentmolecularmethodsfordnaextractionfromindividualteladorsagiacircumcinctanematodes AT spreston comparativeevaluationofdifferentmolecularmethodsfordnaextractionfromindividualteladorsagiacircumcinctanematodes AT mjstear comparativeevaluationofdifferentmolecularmethodsfordnaextractionfromindividualteladorsagiacircumcinctanematodes |