Louie Cris Lopos, Natalia V Bykova, Janeen Robinson, Susan Brown, Kerry Ward, Andriy Bilichak
{"title":"利用农杆菌介导转化产生的小麦(Triticum aestivum L.)转基因植株中转基因整合和基因编辑事件的多样性。","authors":"Louie Cris Lopos, Natalia V Bykova, Janeen Robinson, Susan Brown, Kerry Ward, Andriy Bilichak","doi":"10.3389/fgeed.2023.1265103","DOIUrl":null,"url":null,"abstract":"<p><p>Improvement in agronomic traits in crops through gene editing (GE) relies on efficient transformation protocols for delivering the CRISPR/Cas9-coded transgenes. Recently, a few embryogenesis-related genes have been described, the co-delivery of which significantly increases the transformation efficiency with reduced genotype-dependency. Here, we characterized the transgenic and GE events in wheat (cv. Fielder) when transformed with <i>GROWTH-REGULATING FACTOR 4</i> (<i>GRF4</i>) and its cofactor <i>GRF-INTERACTING FACTOR 1</i> (<i>GIF1</i>) chimeric gene. Transformation efficiency in our experiments ranged from 22% to 68%, and the editing events were faithfully propagated into the following generation. Both low- and high-copy-number integration events were recovered in the T<sub>0</sub> population with various levels of integrity of the left and right T-DNA borders. We also generated a population of wheat plants with 10 different gRNAs targeting 30 loci in the genome. A comparison of the epigenetic profiles at the target sites and editing efficiency revealed a significant positive correlation between chromatin accessibility and mutagenesis rate. Overall, the preliminary screening of transgene quality and GE events in the T<sub>0</sub> population of plants regenerated through the co-delivery of <i>GRF-GIF</i> can allow for the propagation of the best candidates for further phenotypic analysis.</p>","PeriodicalId":73086,"journal":{"name":"Frontiers in genome editing","volume":null,"pages":null},"PeriodicalIF":4.9000,"publicationDate":"2023-12-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10773716/pdf/","citationCount":"0","resultStr":"{\"title\":\"Diversity of transgene integration and gene-editing events in wheat (<i>Triticum aestivum</i> L.) transgenic plants generated using <i>Agrobacterium</i>-mediated transformation.\",\"authors\":\"Louie Cris Lopos, Natalia V Bykova, Janeen Robinson, Susan Brown, Kerry Ward, Andriy Bilichak\",\"doi\":\"10.3389/fgeed.2023.1265103\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<p><p>Improvement in agronomic traits in crops through gene editing (GE) relies on efficient transformation protocols for delivering the CRISPR/Cas9-coded transgenes. Recently, a few embryogenesis-related genes have been described, the co-delivery of which significantly increases the transformation efficiency with reduced genotype-dependency. Here, we characterized the transgenic and GE events in wheat (cv. Fielder) when transformed with <i>GROWTH-REGULATING FACTOR 4</i> (<i>GRF4</i>) and its cofactor <i>GRF-INTERACTING FACTOR 1</i> (<i>GIF1</i>) chimeric gene. Transformation efficiency in our experiments ranged from 22% to 68%, and the editing events were faithfully propagated into the following generation. Both low- and high-copy-number integration events were recovered in the T<sub>0</sub> population with various levels of integrity of the left and right T-DNA borders. We also generated a population of wheat plants with 10 different gRNAs targeting 30 loci in the genome. A comparison of the epigenetic profiles at the target sites and editing efficiency revealed a significant positive correlation between chromatin accessibility and mutagenesis rate. Overall, the preliminary screening of transgene quality and GE events in the T<sub>0</sub> population of plants regenerated through the co-delivery of <i>GRF-GIF</i> can allow for the propagation of the best candidates for further phenotypic analysis.</p>\",\"PeriodicalId\":73086,\"journal\":{\"name\":\"Frontiers in genome editing\",\"volume\":null,\"pages\":null},\"PeriodicalIF\":4.9000,\"publicationDate\":\"2023-12-19\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10773716/pdf/\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Frontiers in genome editing\",\"FirstCategoryId\":\"1085\",\"ListUrlMain\":\"https://doi.org/10.3389/fgeed.2023.1265103\",\"RegionNum\":0,\"RegionCategory\":null,\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"2023/1/1 0:00:00\",\"PubModel\":\"eCollection\",\"JCR\":\"Q1\",\"JCRName\":\"BIOTECHNOLOGY & APPLIED MICROBIOLOGY\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Frontiers in genome editing","FirstCategoryId":"1085","ListUrlMain":"https://doi.org/10.3389/fgeed.2023.1265103","RegionNum":0,"RegionCategory":null,"ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"2023/1/1 0:00:00","PubModel":"eCollection","JCR":"Q1","JCRName":"BIOTECHNOLOGY & APPLIED MICROBIOLOGY","Score":null,"Total":0}
Diversity of transgene integration and gene-editing events in wheat (Triticum aestivum L.) transgenic plants generated using Agrobacterium-mediated transformation.
Improvement in agronomic traits in crops through gene editing (GE) relies on efficient transformation protocols for delivering the CRISPR/Cas9-coded transgenes. Recently, a few embryogenesis-related genes have been described, the co-delivery of which significantly increases the transformation efficiency with reduced genotype-dependency. Here, we characterized the transgenic and GE events in wheat (cv. Fielder) when transformed with GROWTH-REGULATING FACTOR 4 (GRF4) and its cofactor GRF-INTERACTING FACTOR 1 (GIF1) chimeric gene. Transformation efficiency in our experiments ranged from 22% to 68%, and the editing events were faithfully propagated into the following generation. Both low- and high-copy-number integration events were recovered in the T0 population with various levels of integrity of the left and right T-DNA borders. We also generated a population of wheat plants with 10 different gRNAs targeting 30 loci in the genome. A comparison of the epigenetic profiles at the target sites and editing efficiency revealed a significant positive correlation between chromatin accessibility and mutagenesis rate. Overall, the preliminary screening of transgene quality and GE events in the T0 population of plants regenerated through the co-delivery of GRF-GIF can allow for the propagation of the best candidates for further phenotypic analysis.