High-throughput detection of T-DNA insertion sites for multiple transgenes in complex genomes
Abstract Background Genetic engineering of crop plants has been successful in transferring traits into elite lines beyond what can be achieved with breeding techniques. Introduction of transgenes originating from other species has conferred resistance to biotic and abiotic stresses, increased effici...
| Main Authors: | , , , |
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| Format: | Article |
| Language: | English |
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BMC
2022-10-01
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| Series: | BMC Genomics |
| Subjects: | |
| Online Access: | https://doi.org/10.1186/s12864-022-08918-6 |
| _version_ | 1828161248785596416 |
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| author | Brianne Edwards Eli D. Hornstein Nathan J. Wilson Heike Sederoff |
| author_facet | Brianne Edwards Eli D. Hornstein Nathan J. Wilson Heike Sederoff |
| author_sort | Brianne Edwards |
| collection | DOAJ |
| description | Abstract Background Genetic engineering of crop plants has been successful in transferring traits into elite lines beyond what can be achieved with breeding techniques. Introduction of transgenes originating from other species has conferred resistance to biotic and abiotic stresses, increased efficiency, and modified developmental programs. The next challenge is now to combine multiple transgenes into elite varieties via gene stacking to combine traits. Generating stable homozygous lines with multiple transgenes requires selection of segregating generations which is time consuming and labor intensive, especially if the crop is polyploid. Insertion site effects and transgene copy number are important metrics for commercialization and trait efficiency. Results We have developed a simple method to identify the sites of transgene insertions using T-DNA-specific primers and high-throughput sequencing that enables identification of multiple insertion sites in the T1 generation of any crop transformed via Agrobacterium. We present an example using the allohexaploid oil-seed plant Camelina sativa to determine insertion site location of two transgenes. Conclusion This new methodology enables the early selection of desirable transgene location and copy number to generate homozygous lines within two generations. |
| first_indexed | 2024-04-12T00:35:28Z |
| format | Article |
| id | doaj.art-a66837680c6c48a89e5ff577ac0c8e13 |
| institution | Directory Open Access Journal |
| issn | 1471-2164 |
| language | English |
| last_indexed | 2024-04-12T00:35:28Z |
| publishDate | 2022-10-01 |
| publisher | BMC |
| record_format | Article |
| series | BMC Genomics |
| spelling | doaj.art-a66837680c6c48a89e5ff577ac0c8e132022-12-22T03:55:11ZengBMCBMC Genomics1471-21642022-10-0123112010.1186/s12864-022-08918-6High-throughput detection of T-DNA insertion sites for multiple transgenes in complex genomesBrianne Edwards0Eli D. Hornstein1Nathan J. Wilson2Heike Sederoff3Department of Plant and Microbial Biology, North Carolina State UniversityDepartment of Plant and Microbial Biology, North Carolina State UniversityDepartment of Plant and Microbial Biology, North Carolina State UniversityDepartment of Plant and Microbial Biology, North Carolina State UniversityAbstract Background Genetic engineering of crop plants has been successful in transferring traits into elite lines beyond what can be achieved with breeding techniques. Introduction of transgenes originating from other species has conferred resistance to biotic and abiotic stresses, increased efficiency, and modified developmental programs. The next challenge is now to combine multiple transgenes into elite varieties via gene stacking to combine traits. Generating stable homozygous lines with multiple transgenes requires selection of segregating generations which is time consuming and labor intensive, especially if the crop is polyploid. Insertion site effects and transgene copy number are important metrics for commercialization and trait efficiency. Results We have developed a simple method to identify the sites of transgene insertions using T-DNA-specific primers and high-throughput sequencing that enables identification of multiple insertion sites in the T1 generation of any crop transformed via Agrobacterium. We present an example using the allohexaploid oil-seed plant Camelina sativa to determine insertion site location of two transgenes. Conclusion This new methodology enables the early selection of desirable transgene location and copy number to generate homozygous lines within two generations.https://doi.org/10.1186/s12864-022-08918-6TransgeneInsertion sitePolyploidT-DNASequencing |
| spellingShingle | Brianne Edwards Eli D. Hornstein Nathan J. Wilson Heike Sederoff High-throughput detection of T-DNA insertion sites for multiple transgenes in complex genomes BMC Genomics Transgene Insertion site Polyploid T-DNA Sequencing |
| title | High-throughput detection of T-DNA insertion sites for multiple transgenes in complex genomes |
| title_full | High-throughput detection of T-DNA insertion sites for multiple transgenes in complex genomes |
| title_fullStr | High-throughput detection of T-DNA insertion sites for multiple transgenes in complex genomes |
| title_full_unstemmed | High-throughput detection of T-DNA insertion sites for multiple transgenes in complex genomes |
| title_short | High-throughput detection of T-DNA insertion sites for multiple transgenes in complex genomes |
| title_sort | high throughput detection of t dna insertion sites for multiple transgenes in complex genomes |
| topic | Transgene Insertion site Polyploid T-DNA Sequencing |
| url | https://doi.org/10.1186/s12864-022-08918-6 |
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