Practical laboratory class to assess genesilencing using CRISPR interference (CRISPRi) technology in the archaeonHaloferax volcanii.
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| Title: | Practical laboratory class to assess genesilencing using CRISPR interference (CRISPRi) technology in the archaeonHaloferax volcanii. |
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| Authors: | Paggi, R. A.1 (AUTHOR) paggi@mdp.edu.ar, Ferrari, M. C.1 (AUTHOR), Cerletti, M.1 (AUTHOR), Giménez, M. I.1 (AUTHOR), Schwarz, T. S.2 (AUTHOR), Marchfelder, A.2 (AUTHOR), De Castro, R. E.1 (AUTHOR) decastro@mdp.edu.ar |
| Source: | Biochemistry & Molecular Biology Education. Mar/Apr2025, Vol. 53 Issue 2, p155-164. 10p. |
| Subject Terms: | Gene expression, Molecular biology, RNA interference, Small interfering RNA, CRISPRs |
| Abstract: | Perturbation of gene expression using RNA interference (RNAi) or CRISPR interference (CRISPRi) is a useful strategy to explore the function of essential genes. In the archaeon Haloferax volcanii, the CRISPR‐Cas system has been adapted as a CRISPRi tool to silence the expression of specific genes. We developed a laboratory class (LC) to conceptualize gene silencing through inactivation of the H. volcanii LonB protease gene, a negative regulator of carotenoid pigments biosynthesis, using CRISPRi. This LC has been successfully applied in the Biology and Biochemistry of Microorganisms course for undergraduate students of Biology in 2022 and 2023. The following objectives were proposed: (a) generate H. volcanii mutant strains with reduced expression of the lonB gene using CRISPRi; (b) examine the effect of lonB gene silencing on cell pigmentation and growth rate; (c) assess lonB gene repression by Western blotting (WB). This LC allows students to obtain and screen CRISPRi silenced‐mutants by means of simple procedures using a non‐pathogenic organism as well as handle basic microbiology, biochemistry and molecular biology protocols. Additionally, the LC fosters social actions through collaborative work (experimental work), the interpretation and discussion of data and the ability to communicate outcomes orally and in a written format (scientific report). [ABSTRACT FROM AUTHOR] |
| Copyright of Biochemistry & Molecular Biology Education is the property of Wiley-Blackwell and its content may not be copied or emailed to multiple sites without the copyright holder's express written permission. Additionally, content may not be used with any artificial intelligence tools or machine learning technologies. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.) | |
| Database: | Education Research Complete |
| FullText | Text: Availability: 0 |
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| Header | DbId: ehh DbLabel: Education Research Complete An: 183690672 AccessLevel: 6 PubType: Academic Journal PubTypeId: academicJournal PreciseRelevancyScore: 0 |
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| Items | – Name: Title Label: Title Group: Ti Data: Practical laboratory class to assess genesilencing using CRISPR interference (CRISPRi) technology in the archaeonHaloferax volcanii. – Name: Author Label: Authors Group: Au Data: <searchLink fieldCode="AR" term="%22Paggi%2C+R%2E+A%2E%22">Paggi, R. A.</searchLink><relatesTo>1</relatesTo> (AUTHOR)<i> paggi@mdp.edu.ar</i><br /><searchLink fieldCode="AR" term="%22Ferrari%2C+M%2E+C%2E%22">Ferrari, M. C.</searchLink><relatesTo>1</relatesTo> (AUTHOR)<br /><searchLink fieldCode="AR" term="%22Cerletti%2C+M%2E%22">Cerletti, M.</searchLink><relatesTo>1</relatesTo> (AUTHOR)<br /><searchLink fieldCode="AR" term="%22Giménez%2C+M%2E+I%2E%22">Giménez, M. I.</searchLink><relatesTo>1</relatesTo> (AUTHOR)<br /><searchLink fieldCode="AR" term="%22Schwarz%2C+T%2E+S%2E%22">Schwarz, T. S.</searchLink><relatesTo>2</relatesTo> (AUTHOR)<br /><searchLink fieldCode="AR" term="%22Marchfelder%2C+A%2E%22">Marchfelder, A.</searchLink><relatesTo>2</relatesTo> (AUTHOR)<br /><searchLink fieldCode="AR" term="%22De+Castro%2C+R%2E+E%2E%22">De Castro, R. E.</searchLink><relatesTo>1</relatesTo> (AUTHOR)<i> decastro@mdp.edu.ar</i> – Name: TitleSource Label: Source Group: Src Data: <searchLink fieldCode="JN" term="%22Biochemistry+%26+Molecular+Biology+Education%22">Biochemistry & Molecular Biology Education</searchLink>. Mar/Apr2025, Vol. 53 Issue 2, p155-164. 10p. – Name: Subject Label: Subject Terms Group: Su Data: <searchLink fieldCode="DE" term="%22Gene+expression%22">Gene expression</searchLink><br /><searchLink fieldCode="DE" term="%22Molecular+biology%22">Molecular biology</searchLink><br /><searchLink fieldCode="DE" term="%22RNA+interference%22">RNA interference</searchLink><br /><searchLink fieldCode="DE" term="%22Small+interfering+RNA%22">Small interfering RNA</searchLink><br /><searchLink fieldCode="DE" term="%22CRISPRs%22">CRISPRs</searchLink> – Name: Abstract Label: Abstract Group: Ab Data: Perturbation of gene expression using RNA interference (RNAi) or CRISPR interference (CRISPRi) is a useful strategy to explore the function of essential genes. In the archaeon Haloferax volcanii, the CRISPR‐Cas system has been adapted as a CRISPRi tool to silence the expression of specific genes. We developed a laboratory class (LC) to conceptualize gene silencing through inactivation of the H. volcanii LonB protease gene, a negative regulator of carotenoid pigments biosynthesis, using CRISPRi. This LC has been successfully applied in the Biology and Biochemistry of Microorganisms course for undergraduate students of Biology in 2022 and 2023. The following objectives were proposed: (a) generate H. volcanii mutant strains with reduced expression of the lonB gene using CRISPRi; (b) examine the effect of lonB gene silencing on cell pigmentation and growth rate; (c) assess lonB gene repression by Western blotting (WB). This LC allows students to obtain and screen CRISPRi silenced‐mutants by means of simple procedures using a non‐pathogenic organism as well as handle basic microbiology, biochemistry and molecular biology protocols. Additionally, the LC fosters social actions through collaborative work (experimental work), the interpretation and discussion of data and the ability to communicate outcomes orally and in a written format (scientific report). [ABSTRACT FROM AUTHOR] – Name: AbstractSuppliedCopyright Label: Group: Ab Data: <i>Copyright of Biochemistry & Molecular Biology Education is the property of Wiley-Blackwell and its content may not be copied or emailed to multiple sites without the copyright holder's express written permission. Additionally, content may not be used with any artificial intelligence tools or machine learning technologies. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract.</i> (Copyright applies to all Abstracts.) |
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| RecordInfo | BibRecord: BibEntity: Identifiers: – Type: doi Value: 10.1002/bmb.21872 Languages: – Code: eng Text: English PhysicalDescription: Pagination: PageCount: 10 StartPage: 155 Subjects: – SubjectFull: Gene expression Type: general – SubjectFull: Molecular biology Type: general – SubjectFull: RNA interference Type: general – SubjectFull: Small interfering RNA Type: general – SubjectFull: CRISPRs Type: general Titles: – TitleFull: Practical laboratory class to assess genesilencing using CRISPR interference (CRISPRi) technology in the archaeonHaloferax volcanii. Type: main BibRelationships: HasContributorRelationships: – PersonEntity: Name: NameFull: Paggi, R. A. – PersonEntity: Name: NameFull: Ferrari, M. C. – PersonEntity: Name: NameFull: Cerletti, M. – PersonEntity: Name: NameFull: Giménez, M. I. – PersonEntity: Name: NameFull: Schwarz, T. S. – PersonEntity: Name: NameFull: Marchfelder, A. – PersonEntity: Name: NameFull: De Castro, R. E. IsPartOfRelationships: – BibEntity: Dates: – D: 01 M: 03 Text: Mar/Apr2025 Type: published Y: 2025 Identifiers: – Type: issn-print Value: 14708175 Numbering: – Type: volume Value: 53 – Type: issue Value: 2 Titles: – TitleFull: Biochemistry & Molecular Biology Education Type: main |
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