Protein complex isolated from culture

A Nigeran-Protein Complex Isolated from Culture Filtrates... 

More complex proteins have been produced in mammalian cell culture using recombinant DNA techniques. The gene for Factor VIII, a protein involved in blood clotting, is defective in individuals with hemophilia. Before genetically engineered Factor VIII became available, a number of hemophiliac patients died of AIDS or hepatitis that they contracted from transfusions of contaminated blood or from Factor VIII isolated from contaminated blood. 

Figure 6. Time course of production of (%) cell wall nigeran and (O) exocellular nigeran protein complex by A. awamori on incomplete medium. After growth on complete medium, washed cells were transferred to medium lacking nitrogen source (16). Aliquots of the culture were withdrawn periodically and cells were separated by filtration. The nigeran-protein complex was isolated from the filtrate by centrifugation at 30,000 Xg CL d washed repeatedly with water.

For isolating all DNA contained in a cell, the cell culture or tissue sample is transferred into a buffer which contains a detergent such as SDS or Triton A-100. The detergent disrupts the cellular walls and dissociates any DNA-protein complexes. RNA molecules contained in the cell extract are broken up by treatment with a ribonuclease. Proteins can be digested by a proteolytic enzyme, most commonly proteinase K. The DNA can then be extracted from the mixture by precipitation with ethanol. Only long nucleic acid chains precipitate, single nucleotides and products of the RNA digestion remain in solution. 

---