Cargando…

A biolistic method for high-throughput production of transgenic wheat plants with single gene insertions

BACKGROUND: The relatively low efficiency of biolistic transformation and subsequent integration of multiple copies of the introduced gene/s significantly complicate the genetic modification of wheat (Triticum aestivum) and other plant species. One of the key factors contributing to the reproducibil...

Descripción completa

Detalles Bibliográficos
Autores principales: Ismagul, Ainur, Yang, Nannan, Maltseva, Elina, Iskakova, Gulnur, Mazonka, Inna, Skiba, Yuri, Bi, Huihui, Eliby, Serik, Jatayev, Satyvaldy, Shavrukov, Yuri, Borisjuk, Nikolai, Langridge, Peter
Formato: Online Artículo Texto
Lenguaje:English
Publicado: BioMed Central 2018
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6020210/
https://www.ncbi.nlm.nih.gov/pubmed/29940859
http://dx.doi.org/10.1186/s12870-018-1326-1
_version_ 1783335244402786304
author Ismagul, Ainur
Yang, Nannan
Maltseva, Elina
Iskakova, Gulnur
Mazonka, Inna
Skiba, Yuri
Bi, Huihui
Eliby, Serik
Jatayev, Satyvaldy
Shavrukov, Yuri
Borisjuk, Nikolai
Langridge, Peter
author_facet Ismagul, Ainur
Yang, Nannan
Maltseva, Elina
Iskakova, Gulnur
Mazonka, Inna
Skiba, Yuri
Bi, Huihui
Eliby, Serik
Jatayev, Satyvaldy
Shavrukov, Yuri
Borisjuk, Nikolai
Langridge, Peter
author_sort Ismagul, Ainur
collection PubMed
description BACKGROUND: The relatively low efficiency of biolistic transformation and subsequent integration of multiple copies of the introduced gene/s significantly complicate the genetic modification of wheat (Triticum aestivum) and other plant species. One of the key factors contributing to the reproducibility of this method is the uniformity of the DNA/gold suspension, which is dependent on the coating procedure employed. It was also shown recently that the relative frequency of single copy transgene inserts could be increased through the use of nanogram quantities of the DNA during coating. RESULTS: A simplified DNA/gold coating method was developed to produce fertile transgenic plants, via microprojectile bombardment of callus cultures induced from immature embryos. In this method, polyethyleneglycol (PEG) and magnesium salt solutions were utilized in place of the spermidine and calcium chloride of the standard coating method, to precipitate the DNA onto gold microparticles. The prepared microparticles were used to generate transgenics from callus cultures of commercial bread wheat cv. Gladius resulting in an average transformation frequency of 9.9%. To increase the occurrence of low transgene copy number events, nanogram amounts of the minimal expression cassettes containing the gene of interest and the hpt gene were used for co-transformation. A total of 1538 transgenic wheat events were generated from 15,496 embryos across 19 independent experiments. The variation of single copy insert frequencies ranged from 16.1 to 73.5% in the transgenic wheat plants, which compares favourably to published results. CONCLUSIONS: The DNA/gold coating procedure presented here allows efficient, large scale transformation of wheat. The use of nanogram amounts of vector DNA improves the frequency of single copy transgene inserts in transgenic wheat plants.
format Online
Article
Text
id pubmed-6020210
institution National Center for Biotechnology Information
language English
publishDate 2018
publisher BioMed Central
record_format MEDLINE/PubMed
spelling pubmed-60202102018-07-06 A biolistic method for high-throughput production of transgenic wheat plants with single gene insertions Ismagul, Ainur Yang, Nannan Maltseva, Elina Iskakova, Gulnur Mazonka, Inna Skiba, Yuri Bi, Huihui Eliby, Serik Jatayev, Satyvaldy Shavrukov, Yuri Borisjuk, Nikolai Langridge, Peter BMC Plant Biol Methodology Article BACKGROUND: The relatively low efficiency of biolistic transformation and subsequent integration of multiple copies of the introduced gene/s significantly complicate the genetic modification of wheat (Triticum aestivum) and other plant species. One of the key factors contributing to the reproducibility of this method is the uniformity of the DNA/gold suspension, which is dependent on the coating procedure employed. It was also shown recently that the relative frequency of single copy transgene inserts could be increased through the use of nanogram quantities of the DNA during coating. RESULTS: A simplified DNA/gold coating method was developed to produce fertile transgenic plants, via microprojectile bombardment of callus cultures induced from immature embryos. In this method, polyethyleneglycol (PEG) and magnesium salt solutions were utilized in place of the spermidine and calcium chloride of the standard coating method, to precipitate the DNA onto gold microparticles. The prepared microparticles were used to generate transgenics from callus cultures of commercial bread wheat cv. Gladius resulting in an average transformation frequency of 9.9%. To increase the occurrence of low transgene copy number events, nanogram amounts of the minimal expression cassettes containing the gene of interest and the hpt gene were used for co-transformation. A total of 1538 transgenic wheat events were generated from 15,496 embryos across 19 independent experiments. The variation of single copy insert frequencies ranged from 16.1 to 73.5% in the transgenic wheat plants, which compares favourably to published results. CONCLUSIONS: The DNA/gold coating procedure presented here allows efficient, large scale transformation of wheat. The use of nanogram amounts of vector DNA improves the frequency of single copy transgene inserts in transgenic wheat plants. BioMed Central 2018-06-26 /pmc/articles/PMC6020210/ /pubmed/29940859 http://dx.doi.org/10.1186/s12870-018-1326-1 Text en © The Author(s). 2018 Open AccessThis article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.
spellingShingle Methodology Article
Ismagul, Ainur
Yang, Nannan
Maltseva, Elina
Iskakova, Gulnur
Mazonka, Inna
Skiba, Yuri
Bi, Huihui
Eliby, Serik
Jatayev, Satyvaldy
Shavrukov, Yuri
Borisjuk, Nikolai
Langridge, Peter
A biolistic method for high-throughput production of transgenic wheat plants with single gene insertions
title A biolistic method for high-throughput production of transgenic wheat plants with single gene insertions
title_full A biolistic method for high-throughput production of transgenic wheat plants with single gene insertions
title_fullStr A biolistic method for high-throughput production of transgenic wheat plants with single gene insertions
title_full_unstemmed A biolistic method for high-throughput production of transgenic wheat plants with single gene insertions
title_short A biolistic method for high-throughput production of transgenic wheat plants with single gene insertions
title_sort biolistic method for high-throughput production of transgenic wheat plants with single gene insertions
topic Methodology Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6020210/
https://www.ncbi.nlm.nih.gov/pubmed/29940859
http://dx.doi.org/10.1186/s12870-018-1326-1
work_keys_str_mv AT ismagulainur abiolisticmethodforhighthroughputproductionoftransgenicwheatplantswithsinglegeneinsertions
AT yangnannan abiolisticmethodforhighthroughputproductionoftransgenicwheatplantswithsinglegeneinsertions
AT maltsevaelina abiolisticmethodforhighthroughputproductionoftransgenicwheatplantswithsinglegeneinsertions
AT iskakovagulnur abiolisticmethodforhighthroughputproductionoftransgenicwheatplantswithsinglegeneinsertions
AT mazonkainna abiolisticmethodforhighthroughputproductionoftransgenicwheatplantswithsinglegeneinsertions
AT skibayuri abiolisticmethodforhighthroughputproductionoftransgenicwheatplantswithsinglegeneinsertions
AT bihuihui abiolisticmethodforhighthroughputproductionoftransgenicwheatplantswithsinglegeneinsertions
AT elibyserik abiolisticmethodforhighthroughputproductionoftransgenicwheatplantswithsinglegeneinsertions
AT jatayevsatyvaldy abiolisticmethodforhighthroughputproductionoftransgenicwheatplantswithsinglegeneinsertions
AT shavrukovyuri abiolisticmethodforhighthroughputproductionoftransgenicwheatplantswithsinglegeneinsertions
AT borisjuknikolai abiolisticmethodforhighthroughputproductionoftransgenicwheatplantswithsinglegeneinsertions
AT langridgepeter abiolisticmethodforhighthroughputproductionoftransgenicwheatplantswithsinglegeneinsertions
AT ismagulainur biolisticmethodforhighthroughputproductionoftransgenicwheatplantswithsinglegeneinsertions
AT yangnannan biolisticmethodforhighthroughputproductionoftransgenicwheatplantswithsinglegeneinsertions
AT maltsevaelina biolisticmethodforhighthroughputproductionoftransgenicwheatplantswithsinglegeneinsertions
AT iskakovagulnur biolisticmethodforhighthroughputproductionoftransgenicwheatplantswithsinglegeneinsertions
AT mazonkainna biolisticmethodforhighthroughputproductionoftransgenicwheatplantswithsinglegeneinsertions
AT skibayuri biolisticmethodforhighthroughputproductionoftransgenicwheatplantswithsinglegeneinsertions
AT bihuihui biolisticmethodforhighthroughputproductionoftransgenicwheatplantswithsinglegeneinsertions
AT elibyserik biolisticmethodforhighthroughputproductionoftransgenicwheatplantswithsinglegeneinsertions
AT jatayevsatyvaldy biolisticmethodforhighthroughputproductionoftransgenicwheatplantswithsinglegeneinsertions
AT shavrukovyuri biolisticmethodforhighthroughputproductionoftransgenicwheatplantswithsinglegeneinsertions
AT borisjuknikolai biolisticmethodforhighthroughputproductionoftransgenicwheatplantswithsinglegeneinsertions
AT langridgepeter biolisticmethodforhighthroughputproductionoftransgenicwheatplantswithsinglegeneinsertions