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Full-Text Articles in Biochemical and Biomolecular Engineering
Obtaining Of The “Green Flourecent Protein - Gfp” In Yeast Of Pichia Pastoris As Repoter Of Heterological Protein Expression, Ashirоv Оybek, Sasmakоv Sobirdjan, Abdurakhmanоv Jaloliddin, Hasanоv Shuhrat
Obtaining Of The “Green Flourecent Protein - Gfp” In Yeast Of Pichia Pastoris As Repoter Of Heterological Protein Expression, Ashirоv Оybek, Sasmakоv Sobirdjan, Abdurakhmanоv Jaloliddin, Hasanоv Shuhrat
CHEMISTRY AND CHEMICAL ENGINEERING
Based on pPIC3.5 and pPIC9 yeast vectors the recombinant plasmid DNA pPIC3.5-GFP (8464 bp) and pPIC9-GFP (8718 bp) containing cDNA (719 bp) of Green fluorescent protein (GFP) were cloned. The obtained plasmids were transformed into yeast cells of the GS115 Pichia pastoris strain. Transformants are selected on histidine-deficient medium. Recombinant clones of pPIC3.5-GFP and pPIC9-GFP were identified by detecting cell fluorescence. GFP expression in yeast is confirmed by green fluorescence. Using electrophoresis in PAAG the molecular weight of the synthesized protein (≈27 kDa) is determined. The resulting reporter constructs and recombinant strains can be used to study the expression level …
Engineering A Fluorescent Protease Sensor For In Vivo Protein Detection, Thomas C. Kinard
Engineering A Fluorescent Protease Sensor For In Vivo Protein Detection, Thomas C. Kinard
Honors Scholar Theses
This report details the results of an ongoing project to engineer a mutant form of Red Fluorescent Protein (RFP) variant mCherry that acts as a real-time in vivo protease sensor. The sought-after mutant only becomes fluorescent when exposed to Tobacco Etch Virus (TEV) Protease, this system’s model protease. This will be accomplished via the insertion of the TEV Protease Recognition Site (TEV-PRS) in such a position that, before cleavage, will prevent the protein from folding to fluorescent conformation, but upon cleavage, will allow for fluorescent conformation to occur. The cylindrical structure of the protein, composed of beta-pleated sheets, contains “loops” …