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|Title:||Microbial expression of a gene for human growth hormone|
|Abstract:||Described are methods and means for the construction and microbial expression of quasi-synthetic genes arising from the combination of organic synthesis and enzymatic reverse transcription from messenger RNA sequences incomplete from the standpoint of the desired protein product. Preferred products of expression lack bio-inactivating leader sequences common in eukaryotic expression products but problematic with regard to microbial cleavage to yield bioactive material. Illustrative is a preferred embodiment in which a gene coding for human growth hormone (useful in, e.g., treatment of hypopituitary dwarfism) is constructed and expressed.|
|Inventor(s):||Goeddel; David V. (Burlingame, CA), Heyneker; Herbert L. (Burlingame, CA)|
|Assignee:||Genentech Inc. (South San Francisco, CA)|
|Filing Date:||Mar 09, 1982|
|Claims:||1. A method of producing human growth hormone which comprises: |
(a) disposing a culture of bacterial transformants comprising plasmids which, in a transformant bacterium, will express a gene for human growth hormone unaccompanied by the leader sequence of human growth hormone or other extraneous protein bound thereto within a fermenter vessel comprising aeration and agitation means in an aqueous, nutriment-containing fermentation broth;
(b) growing up the culture under aeration and agitation while supplying additional nutriments as required to maintain vigorous growth;
(c) separating the resulting cellular mass from the fermentation broth;
(d) lysing the cells to free the contents thereof;
(e) separating cellular debris from supernatant; and
(f) isolating and purifying human growth hormone contained in the supernatant.
2. A method for producing human growth hormone which method comprises culturing bacterial transformants containing recombinant plasmids which will, in a transformant bacterium, express a gene for human growth hormone unaccompanied by the leader sequence of human growth hormone or other extraneous protein bound thereto, and isolating and purifying said expressed human growth hormone.
3. The method of claim 1 wherein the bacteria are transformant E. coli having a selection characteristic and wherein selection pressure is applied to discourage competition by wild-type E. coli.
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