An established protocol for generating transgenic wheat for wheat functional genomics via particle bombardment.
An established protocol for generating transgenic wheat for wheat functional genomics via particle bombardment.
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通过粒子轰击生成用于小麦功能基因组学的转基因小麦的既定方案
DOI：
10.3389/fpls.2022.979540
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发表时间：
2022
影响因子：
5.6
通讯作者：
中科院分区：
文献类型：
--
作者：
小麦是世界上最重要的粮食作物之一，被认为是作物生物技术的首要目标之一。随着小麦及其近缘种高质量参考基因组的出现和近年来小麦族基因组资源的激增，对小麦基因功能研究和遗传改良的需求迅速增加，这就要求转基因小麦的生产应成为常规技术。虽然建立了20多年，但粒子轰击介导的小麦转化尚未成为常规，只有少数实验室精通这项技术。这可能至少部分是由于转化效率低和技术困难。在这里，我们通过适应和优化来描述该方法的当前版本。我们报告了通过粒子枪生产转基因小麦的详细方案，包括几个关键步骤，从选择合适的外植体（即，未成熟的小盾片），DNA包被的金颗粒的制备，以及几种已建立的组织培养策略。更重要的是，我们实验室在小麦转化方面有超过20年的经验，我们分享了许多技术细节和建议，并强调与农杆菌介导的方法相比，粒子轰击介导的方法在基因型依赖性和载体构建方面具有更少的限制。粒子轰击介导的方法已经成功地用于30多个小麦基因型，从四倍体硬粒小麦到六倍体普通小麦，从现代优良品种到地方品种。总之，粒子轰击介导的小麦转化已显示出其潜在的和广泛的应用，和全套的协议，经验，并在许多小麦基因型的成功报告，这里描述将进一步其影响，使其成为常规和强大的技术在作物研究实验室在世界各地。
Wheat is one of the most important food crops in the world and is considered one of the top targets in crop biotechnology. With the high-quality reference genomes of wheat and its relative species and the recent burst of genomic resources in Triticeae, demands to perform gene functional studies in wheat and genetic improvement have been rapidly increasing, requiring that production of transgenic wheat should become a routine technique. While established for more than 20 years, the particle bombardment-mediated wheat transformation has not become routine yet, with only a handful of labs being proficient in this technique. This could be due to, at least partly, the low transformation efficiency and the technical difficulties. Here, we describe the current version of this method through adaptation and optimization. We report the detailed protocol of producing transgenic wheat by the particle gun, including several critical steps, from the selection of appropriate explants (i.e., immature scutella), the preparation of DNA-coated gold particles, and several established strategies of tissue culture. More importantly, with over 20 years of experience in wheat transformation in our lab, we share the many technical details and recommendations and emphasize that the particle bombardment-mediated approach has fewer limitations in genotype dependency and vector construction when compared with the Agrobacterium-mediated methods. The particle bombardment-mediated method has been successful for over 30 wheat genotypes, from the tetraploid durum wheat to the hexaploid common wheat, from modern elite varieties to landraces. In conclusion, the particle bombardment-mediated wheat transformation has demonstrated its potential and wide applications, and the full set of protocol, experience, and successful reports in many wheat genotypes described here will further its impacts, making it a routine and robust technique in crop research labs worldwide.
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影响因子：
3.8
作者：
Gadaleta, A;Giancaspro, A;Blanco, A
通讯作者：
Blanco, A
DOI：
10.1007/978-1-4939-7337-8_3
发表时间：
2017-01-01
期刊：
WHEAT BIOTECHNOLOGY: METHODS AND PROTOCOLS
影响因子：
--
作者：
Han, Jiapeng;Yu, Xiaofen;He, Guangyuan
通讯作者：
He, Guangyuan
影响因子：
56.9
作者：
Avni, Raz;Nave, Moran;Distelfeld, Assaf
通讯作者：
Distelfeld, Assaf
DOI：
10.1007/s11627-019-10042-2
发表时间：
2020-01-02
影响因子：
2.6
作者：
Hoerster, George;Wang, Ning;Gordon-Kamm, Bill
通讯作者：
Gordon-Kamm, Bill
DOI：
10.1111/tpj.13446
发表时间：
2017-03
期刊：
The Plant journal : for cell and molecular biology
影响因子：
--
作者：
Gil-Humanes J;Wang Y;Liang Z;Shan Q;Ozuna CV;Sánchez-León S;Baltes NJ;Starker C;Barro F;Gao C;Voytas DF
通讯作者：
Voytas DF