A convenient method for the identification and expression of eukaryotic genes

Area of Science:

• Biotechnology
• Molecular Biology
• Biochemistry

Background:

• Traditional methods for assessing eukaryotic gene product activity involve in vitro systems.
• Mammalian expression vectors are commonly used for gene expression studies.
• An efficient in vivo system offers advantages for protein production and biological activity assessment.

Purpose of the Study:

• To develop an efficient in vivo system for expressing eukaryotic genes in lactating-goat mammary glands.
• To assess the biological activity of expressed proteins using a mammary-gland-specific vector.
• To evaluate the potential of this system as a bioreactor for biological protein production.

Main Methods:

• Synthesized lumbrokinase cDNA variant (LK-m) and tissue-type plasminogen activator (tPA) cDNA were cloned downstream of the goat beta-casein regulatory sequence.
• Expression plasmids were injected into lactating-goat mammary gland tissues.
• Fibrin-agarose plate assay was used to detect fibrin-lysis activity, indicating protein expression.

Main Results:

• High-level expression of LK-m and tPA was detected, demonstrating significant fibrin-lysis activity.
• Specific expression initiated immediately post-injection, peaking within 6-9 hours.
• Peak expression persisted for 20-24 hours and lasted for 4 days with a gradual decline.

Conclusions:

• An efficient in vivo expression system was successfully developed in lactating-goat mammary glands.
• This system enables high-level production and functional assessment of therapeutic proteins.
• The developed system shows potential as a bioreactor for producing biological proteins, offering an alternative to transgenic animals.