Dual- RNA Guided Editing of E.coli's DnaB Helicase Using the CRISPR/Cas9 System

Hdl Handle:
http://hdl.handle.net/10675.2/582733
Title:
Dual- RNA Guided Editing of E.coli's DnaB Helicase Using the CRISPR/Cas9 System
Authors:
Jones, Preston Dimitri
Abstract:
Precise manipulation of the genomic DNA is necessary for the characterization and identification of new genes and proteins. New technologies have enabled more facile and precise genomic engineering. Genome editing using the clustered regularly interspaced short palindromic repeats (CRISPR)- CRISPR associated protein method affords precise manipulation of the genome. This system relies on a precise RNA guide (crRNA) that guides the cas9 nuclease to a specific location on the genomic, where it creates a double strand break (DSB). We can take advantage of this adaptive immunological advance and reprogram crRNAs to target whatever gene we like and introduce precise mutations to the genome by offering a designer template to introduce the changes and help the cell remain viable. This method is on the forefront of biochemistry and is being implemented in a numerous eukaryotic systems. However, it has yet to be fully utilized in bacterial genome editing. We are interested in exploring the bacterial helicase EcDnaB. The EcDnaB helicase has been hypothesized to unwind DNA in two fashions; both strands simultaneously or by unwinding one strand while excluding the other. Our lab postulates that there is a steric exclusion of one of the strands as it winds around the exterior of the helicase via electrostatic interactions. We used the CRISPR-Cas9 system to precisely mutate six loci on the EcDnaB gene, corresponding to amino acid residues on the external surface of the helicase, in order to better understand the mechanism of unwinding and to support our proposed method of unwinding. Begin Time: 37:59 End Time: 59:59
Affiliation:
College of Science and Mathematics
Issue Date:
11-Sep-2015
URI:
http://hdl.handle.net/10675.2/582733
Additional Links:
https://lecture.gru.edu/ess/echo/presentation/eeb7244a-d357-43ba-b880-53326b5018cb?ec=true
Type:
Presentation
Language:
en_US
Description:
Presentation given at the CURS Brown Bag Seminar Series on September 11, 2015
Series/Report no.:
Fall; 2015
Sponsors:
Center for Undergraduate Research and Scholarship; College of Science and Mathematics; Department of Chemistry and Physics
Appears in Collections:
CURS Brown Bag Presentations

Full metadata record

DC FieldValue Language
dc.contributor.authorJones, Preston Dimitrien
dc.date.accessioned2015-11-25T15:34:29Zen
dc.date.available2015-11-25T15:34:29Zen
dc.date.issued2015-09-11en
dc.identifier.urihttp://hdl.handle.net/10675.2/582733en
dc.descriptionPresentation given at the CURS Brown Bag Seminar Series on September 11, 2015en
dc.description.abstractPrecise manipulation of the genomic DNA is necessary for the characterization and identification of new genes and proteins. New technologies have enabled more facile and precise genomic engineering. Genome editing using the clustered regularly interspaced short palindromic repeats (CRISPR)- CRISPR associated protein method affords precise manipulation of the genome. This system relies on a precise RNA guide (crRNA) that guides the cas9 nuclease to a specific location on the genomic, where it creates a double strand break (DSB). We can take advantage of this adaptive immunological advance and reprogram crRNAs to target whatever gene we like and introduce precise mutations to the genome by offering a designer template to introduce the changes and help the cell remain viable. This method is on the forefront of biochemistry and is being implemented in a numerous eukaryotic systems. However, it has yet to be fully utilized in bacterial genome editing. We are interested in exploring the bacterial helicase EcDnaB. The EcDnaB helicase has been hypothesized to unwind DNA in two fashions; both strands simultaneously or by unwinding one strand while excluding the other. Our lab postulates that there is a steric exclusion of one of the strands as it winds around the exterior of the helicase via electrostatic interactions. We used the CRISPR-Cas9 system to precisely mutate six loci on the EcDnaB gene, corresponding to amino acid residues on the external surface of the helicase, in order to better understand the mechanism of unwinding and to support our proposed method of unwinding. Begin Time: 37:59 End Time: 59:59en
dc.description.sponsorshipCenter for Undergraduate Research and Scholarship; College of Science and Mathematics; Department of Chemistry and Physicsen
dc.language.isoen_USen
dc.relation.ispartofseriesFallen
dc.relation.ispartofseries2015en
dc.relation.urlhttps://lecture.gru.edu/ess/echo/presentation/eeb7244a-d357-43ba-b880-53326b5018cb?ec=trueen
dc.subjectDNAen
dc.subjectCRISPR-Associated Proteinsen
dc.subjectGenomicsen
dc.subjectClustered Regularly Interspaceden
dc.titleDual- RNA Guided Editing of E.coli's DnaB Helicase Using the CRISPR/Cas9 Systemen_US
dc.typePresentationen
dc.contributor.departmentCollege of Science and Mathematicsen
dc.contributor.mentorSpencer, Angieen
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