Establishment and Characterization of a Novel Tissue-specific DNA Construct and Culture System with Potential for Avian Bioreactor Generation

• Published in: Molecular biotechnology Vol. 61; no. 6; pp. 400 - 409
• Main Authors: Pérez Sáez, Juan M., Bussmann, Leonardo E., Bussmann, Ursula Agnes
• Format: Journal Article
• Language: English
• Published: New York Springer US 01.06.2019; Springer Nature B.V
• Subjects: 5' Flanking Region; Animals; Animals, Genetically Modified; Avian Proteins - genetics; Avian Proteins - metabolism; Biochemistry; Biological Techniques; Bioreactors; Biotechnology; Cell Biology; Chemistry; Chemistry and Materials Science; Chick Embryo; Chickens; Chickens - genetics; Chickens - metabolism; Cloning, Molecular; Culture; Deoxyribonucleic acid; DNA; Eggs; Estradiol - pharmacology; Female; Fibroblasts - cytology; Fibroblasts - drug effects; Fibroblasts - metabolism; Gene expression; Gene Expression Regulation; Genes, Reporter; Genetic modification; Genetic Vectors - chemistry; Genetic Vectors - metabolism; Hepatocytes - cytology; Hepatocytes - drug effects; Hepatocytes - metabolism; Human Genetics; In vitro methods and tests; Luciferases - genetics; Luciferases - metabolism; Original Paper; Ovalbumin; Poultry; Primary Cell Culture; Promoter Regions, Genetic; Protein Science; Proteins; Receptors, Estrogen; Recombinant Fusion Proteins - genetics; Recombinant Fusion Proteins - metabolism; Tissues; Transfection - methods; Vitellogenin; Vitellogenins - genetics; Vitellogenins - metabolism; Zygote - drug effects; Zygote - growth & development; Zygote - metabolism; LMH cells; Avian bioreactors; Hepatocytes; Vitellogenin promoter; Transgenic Chicken; Recombinant proteins; Tissue-specific Promoters
• ISSN: 1073-6085; 1559-0305
• DOI: 10.1007/s12033-019-00170-w

Transgenic chickens are of great interest for the production of recombinant proteins in their eggs. However, the use of constitutive strong promoters or the tissue-specific ovalbumin promoter for the generation of the transgenic chickens have different drawbacks that have to be overcome in order to make chicken bioreactor an efficient production system. This prompted us to investigate the use of an alternative tissue-specific promoter, the vitellogenin promoter, which could overcome the difficulties currently found in the generation of chicken bioreactors. In the present work we establish and characterize a DNA construct consisting of a fragment of the 5´-flanking region of the chicken vitellogenin II gene cloned in a reporter vector. This construct is capable of showing the ability of the promoter to drive expression of a reporting gene in a tissue-specific manner and in a way that closely resembles physiologic regulation of vitellogenin, making it an ideal candidate to be used in the future for generation of avian bioreactors. Besides, we validate an in vitro culture system to test the performance of the DNA construct under study that could be used as a practical tool before generating any transgenic chicken. These results are important since they provide the proof of concept for the use of the vitellogenin promoter for future genetic modification of chickens bioreactors with improved characteristics in terms of quality of the recombinant protein produced.