RNAi and CRISPR/Cas9 as Functional Genomics Tools in the Neotropical Stink Bug, <i>Euschistus heros</i>
The Neotropical brown stink bug, <i>Euschistus heros,</i> is one of the most important stink bug pests in leguminous plants in South America. RNAi and CRISPR/Cas9 are important and useful tools in functional genomics, as well as in the future development of new integrated pest management...
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| Format: | Article |
| Language: | English |
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MDPI AG
2020-11-01
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| Series: | Insects |
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| Online Access: | https://www.mdpi.com/2075-4450/11/12/838 |
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| author | Deise Cagliari Guy Smagghe Moises Zotti Clauvis Nji Tizi Taning |
| author_facet | Deise Cagliari Guy Smagghe Moises Zotti Clauvis Nji Tizi Taning |
| author_sort | Deise Cagliari |
| collection | DOAJ |
| description | The Neotropical brown stink bug, <i>Euschistus heros,</i> is one of the most important stink bug pests in leguminous plants in South America. RNAi and CRISPR/Cas9 are important and useful tools in functional genomics, as well as in the future development of new integrated pest management strategies. Here, we explore the use of these technologies as complementing functional genomic tools in <i>E. heros</i>. Three genes, <i>abnormal wing disc</i> (<i>awd</i>), <i>tyrosine hydroxylase</i> (<i>th</i>) and <i>yellow</i> (<i>yel</i>), known to be involved in wing development (<i>awd</i>) and the melanin pathway (<i>th</i> and <i>yel</i>) in other insects, were chosen to be evaluated using RNAi and CRISPR/Cas9 as tools. First, the genes were functionally characterized using RNAi knockdown technology. The expected phenotype of either deformed wing or lighter cuticle pigmentation/defects in cuticle sclerotization was observed for <i>awd</i> and <i>th</i>, respectively. However, for <i>yel</i>, no obvious phenotype was observed. Based on this, <i>yel</i> was selected as a target for the development of a CRISPR/Cas9 workflow to study gene knockout in <i>E. heros.</i> A total of 719 eggs were injected with the Cas9 nuclease (300 ng/µL) together with the sgRNA (300 ng/µL) targeting <i>yel</i>. A total of six insects successfully hatched from the injected eggs and one of the insects showed mutation in the target region, however, the phenotype was still not obvious. Overall, this study for the first time provides a useful CRISPR/Cas9 gene editing methodology to complement RNAi for functional genomic studies in one of the most important and economically relevant stink bug species. |
| first_indexed | 2024-03-10T14:28:19Z |
| format | Article |
| id | doaj.art-955532ddc3fe4f1f996c519c62353f05 |
| institution | Directory Open Access Journal |
| issn | 2075-4450 |
| language | English |
| last_indexed | 2024-03-10T14:28:19Z |
| publishDate | 2020-11-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Insects |
| spelling | doaj.art-955532ddc3fe4f1f996c519c62353f052023-11-20T22:48:56ZengMDPI AGInsects2075-44502020-11-01111283810.3390/insects11120838RNAi and CRISPR/Cas9 as Functional Genomics Tools in the Neotropical Stink Bug, <i>Euschistus heros</i>Deise Cagliari0Guy Smagghe1Moises Zotti2Clauvis Nji Tizi Taning3Department of Plants and Crops, Faculty of Bioscience Engineering, Ghent University, 9000 Ghent, BelgiumDepartment of Plants and Crops, Faculty of Bioscience Engineering, Ghent University, 9000 Ghent, BelgiumDepartment of Crop Protection, Molecular Entomology Laboratory, Federal University of Pelotas, Pelotas 96160-000, BrazilDepartment of Plants and Crops, Faculty of Bioscience Engineering, Ghent University, 9000 Ghent, BelgiumThe Neotropical brown stink bug, <i>Euschistus heros,</i> is one of the most important stink bug pests in leguminous plants in South America. RNAi and CRISPR/Cas9 are important and useful tools in functional genomics, as well as in the future development of new integrated pest management strategies. Here, we explore the use of these technologies as complementing functional genomic tools in <i>E. heros</i>. Three genes, <i>abnormal wing disc</i> (<i>awd</i>), <i>tyrosine hydroxylase</i> (<i>th</i>) and <i>yellow</i> (<i>yel</i>), known to be involved in wing development (<i>awd</i>) and the melanin pathway (<i>th</i> and <i>yel</i>) in other insects, were chosen to be evaluated using RNAi and CRISPR/Cas9 as tools. First, the genes were functionally characterized using RNAi knockdown technology. The expected phenotype of either deformed wing or lighter cuticle pigmentation/defects in cuticle sclerotization was observed for <i>awd</i> and <i>th</i>, respectively. However, for <i>yel</i>, no obvious phenotype was observed. Based on this, <i>yel</i> was selected as a target for the development of a CRISPR/Cas9 workflow to study gene knockout in <i>E. heros.</i> A total of 719 eggs were injected with the Cas9 nuclease (300 ng/µL) together with the sgRNA (300 ng/µL) targeting <i>yel</i>. A total of six insects successfully hatched from the injected eggs and one of the insects showed mutation in the target region, however, the phenotype was still not obvious. Overall, this study for the first time provides a useful CRISPR/Cas9 gene editing methodology to complement RNAi for functional genomic studies in one of the most important and economically relevant stink bug species.https://www.mdpi.com/2075-4450/11/12/838gene knockdowngene knockoutPentatomidaegene editing |
| spellingShingle | Deise Cagliari Guy Smagghe Moises Zotti Clauvis Nji Tizi Taning RNAi and CRISPR/Cas9 as Functional Genomics Tools in the Neotropical Stink Bug, <i>Euschistus heros</i> Insects gene knockdown gene knockout Pentatomidae gene editing |
| title | RNAi and CRISPR/Cas9 as Functional Genomics Tools in the Neotropical Stink Bug, <i>Euschistus heros</i> |
| title_full | RNAi and CRISPR/Cas9 as Functional Genomics Tools in the Neotropical Stink Bug, <i>Euschistus heros</i> |
| title_fullStr | RNAi and CRISPR/Cas9 as Functional Genomics Tools in the Neotropical Stink Bug, <i>Euschistus heros</i> |
| title_full_unstemmed | RNAi and CRISPR/Cas9 as Functional Genomics Tools in the Neotropical Stink Bug, <i>Euschistus heros</i> |
| title_short | RNAi and CRISPR/Cas9 as Functional Genomics Tools in the Neotropical Stink Bug, <i>Euschistus heros</i> |
| title_sort | rnai and crispr cas9 as functional genomics tools in the neotropical stink bug i euschistus heros i |
| topic | gene knockdown gene knockout Pentatomidae gene editing |
| url | https://www.mdpi.com/2075-4450/11/12/838 |
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