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Integrating Omics and CRISPR Technology for Identification and Verification of Genomic Safe Harbor Loci in the Chicken Genome.
- Source :
-
Biological Procedures Online . 6/24/2023, Vol. 25 Issue 1, p1-20. 20p. - Publication Year :
- 2023
-
Abstract
- Background: One of the most prominent questions in the field of transgenesis is 'Where in the genome to integrate a transgene?'. Escape from epigenetic silencing and promoter shutdown of the transgene needs reliable genomic safe harbor (GSH) loci. Advances in genome engineering technologies combined with multi-omics bioinformatics data have enabled rational evaluation of GSH loci in the host genome. Currently, no validated GSH loci have been evaluated in the chicken genome. Results: Here, we analyzed and experimentally examined two GSH loci in the genome of chicken cells. To this end, putative GSH loci including chicken HIPP-like (cHIPP; between DRG1 and EIF4ENIF1 genes) and chicken ROSA-like (cROSA; upstream of the THUMPD3 gene) were predicted using multi-omics bioinformatics data. Then, the durable expression of the transgene was validated by experimental characterization of continuously-cultured isogenous cell clones harboring DsRed2-ΔCMV-EGFP cassette in the predicted loci. The weakened form of the CMV promoter (ΔCMV) allowed the precise evaluation of GSH loci in a locus-dependent manner compared to the full-length CMV promoter. Conclusions: cHIPP and cROSA loci introduced in this study can be reliably exploited for consistent bio-manufacturing of recombinant proteins in the genetically-engineered chickens. Also, results showed that the genomic context dictates the expression of transgene controlled by ΔCMV in GSH loci. [ABSTRACT FROM AUTHOR]
Details
- Language :
- English
- ISSN :
- 14809222
- Volume :
- 25
- Issue :
- 1
- Database :
- Academic Search Index
- Journal :
- Biological Procedures Online
- Publication Type :
- Academic Journal
- Accession number :
- 164489208
- Full Text :
- https://doi.org/10.1186/s12575-023-00210-5