NDLI: Minimum length of direct repeat sequences required for efficient homologous recombination induced by zinc finger nuclease in yeast

Content Provider	Springer Nature Link
Author	Ren, ChongHua Yan, Qiang Zhang, ZhiYing
Copyright Year	2014
Abstract	Zinc finger nuclease (ZFN) technology is a powerful molecular tool for targeted genome modifications and genetic engineering. However, screening for specific ZFs and validation of ZFN activity are labor intensive and time consuming. We previously designed a yeast-based ZFN screening and validation system by inserting a ZFN binding site flanked by a 164 bp direct repeat sequence into the middle of a Gal4 transcription factor, disrupting the open reading frame of the yeast Gal4 gene. Expression of the ZFN causes a double stranded break at its binding site, which promotes the cellular DNA repair system to restore expression of a functional Gal transcriptional factor via homologous recombination. Expression of Gal4 transcription factor leads to activation of three reporter genes in an AH109 yeast two-hybrid strain. However, the 164 bp direct repeat appears to generate spontaneous homologous recombination frequently, resulting in many false positive ZFNs. To overcome this, a series of DNA fragments of various lengths from 10 to 150 bp with 10 bp increase each and 164 bp direct repeats flanking the ZFN binding site were designed and constructed. The results demonstrated that the minimum length required for ZFN-induced homologous recombination was 30 bp, which almost eliminated spontaneous recombination. Using the 30 bp direct repeat sequence, ZFN could efficiently induce homologous recombination, while false positive ZFNs resulting from spontaneous homologous recombination were minimized. Thus, this study provided a simple, fast and sensitive ZFN screening and activity validation system in yeast.
Starting Page	6939
Ending Page	6948
Page Count	10
File Format	PDF
ISSN	03014851
Journal	Molecular Biology Reports
Volume Number	41
Issue Number	10
e-ISSN	15734978
Language	English
Publisher	Springer Netherlands
Publisher Date	2014-07-15
Publisher Place	Dordrecht
Access Restriction	One Nation One Subscription (ONOS)
Subject Keyword	Zinc finger nuclease (ZFN) Homologous recombination Direct repeats Yeast Animal Biochemistry Animal Anatomy / Morphology / Histology
Content Type	Text
Resource Type	Article
Subject	Genetics Medicine Molecular Biology

Sl.	Authority	Responsibilities	Communication Details
1	Ministry of Education (GoI), Department of Higher Education	Sanctioning Authority	https://www.education.gov.in/ict-initiatives
2	Indian Institute of Technology Kharagpur	Host Institute of the Project: The host institute of the project is responsible for providing infrastructure support and hosting the project	https://www.iitkgp.ac.in
3	National Digital Library of India Office, Indian Institute of Technology Kharagpur	The administrative and infrastructural headquarters of the project	Dr. B. Sutradhar bsutra@ndl.gov.in
4	Project PI / Joint PI	Principal Investigator and Joint Principal Investigators of the project	Dr. B. Sutradhar bsutra@ndl.gov.in Prof. Saswat Chakrabarti will be added soon
5	Website/Portal (Helpdesk)	Queries regarding NDLI and its services	support@ndl.gov.in
6	Contents and Copyright Issues	Queries related to content curation and copyright issues	content@ndl.gov.in
7	National Digital Library of India Club (NDLI Club)	Queries related to NDLI Club formation, support, user awareness program, seminar/symposium, collaboration, social media, promotion, and outreach	clubsupport@ndl.gov.in
8	Digital Preservation Centre (DPC)	Assistance with digitizing and archiving copyright-free printed books	dpc@ndl.gov.in
9	IDR Setup or Support	Queries related to establishment and support of Institutional Digital Repository (IDR) and IDR workshops	idr@ndl.gov.in

Zinc finger nuclease: a new approach for excising HIV-1 proviral DNA from infected human T cells

Targeted disruption of the sheep MSTN gene by engineered zinc-finger nucleases

Combination of a Zinc Finger and Homeodomain Required for Protein-Interaction

Targeted editing of goat genome with modular-assembly zinc finger nucleases based on activity prediction by computational molecular modeling

Efficient Knockin Mouse Generation by ssDNA Oligonucleotides and Zinc-Finger Nuclease Assisted Homologous Recombination in Zygotes

Mapping of Immune-Mediated Disease Genes

Characterization of grape C-repeat-binding factor 2 and B-box-type zinc finger protein in transgenic Arabidopsis plants under stress conditions

A suicidal zinc finger nuclease expression coupled with a surrogate reporter for efficient genome engineering

Identification and Characterization of the Human SCAN Domain Zinc-Finger Gene ZNF449

Minimum length of direct repeat sequences required for efficient homologous recombination induced by zinc finger nuclease in yeast

Similar Documents

Zinc finger nuclease: a new approach for excising HIV-1 proviral DNA from infected human T cells

Targeted disruption of the sheep MSTN gene by engineered zinc-finger nucleases

Combination of a Zinc Finger and Homeodomain Required for Protein-Interaction

Targeted editing of goat genome with modular-assembly zinc finger nucleases based on activity prediction by computational molecular modeling

Efficient Knockin Mouse Generation by ssDNA Oligonucleotides and Zinc-Finger Nuclease Assisted Homologous Recombination in Zygotes

Mapping of Immune-Mediated Disease Genes

Characterization of grape C-repeat-binding factor 2 and B-box-type zinc finger protein in transgenic Arabidopsis plants under stress conditions

A suicidal zinc finger nuclease expression coupled with a surrogate reporter for efficient genome engineering

Identification and Characterization of the Human SCAN Domain Zinc-Finger Gene ZNF449

Minimum length of direct repeat sequences required for efficient homologous recombination induced by zinc finger nuclease in yeast