碩士 / 國立臺灣大學 / 漁業科學研究所 / 87 / It is difficult to gain pure growth hormone (GH) via extraction of animal pituitary glands. However, with genetic engineering technology, large amount of recombinant GH (rGH) can be obtained through either prokaryotic or eukaryotic expression system. In prokaryotic system, 33.66 ± 0.16 mg of rGH was obtained out of 100 ml E. coli culture containing yellowfin porgy GH (ypGH) cDNA. Mass spectrometer and densito-pattern analyzer analysis show that the molecular weight of the E. coli-expressed rGH is 2,1397 kilo Dalton (kDa) and constitutes 80% of the purified protein. Biological activity of the rGH was assayed via immersing fries of grouper (Epinephelus malabaricus) in seawater containing rGH. After 17 days of experiment, our result showed that 30 min immersion with 500 mg rGH/liter promotes fries growth most prominently. Compared with control group, the weight gain rate and average body weight were 170% and 69% higher.
In eukaryotic system, plasmid (pSJ04) containing ypGH cDNA was introduced into yeast (P. pastoris). The transformed yeast was selected by nutrient limited medium (RDB) and antibiotic-containing (YPD-G418, 4 mg/ml) culture plates. One unique protein band of 22 kDa was identified exclusively in experimental group in SDS-PAGE analysis, and the 22-kDa protein reacted positively against rabbit anti-ypGH polyclonal antibody in western blotting analysis. After being induced for 72 hr, 800 milliliters of the transformed yeast were subjected to ELISA. The rGH concentraction was 0.425 ± 0.014 μg/ml. The transformed yeast harvested from large scale preparation was frozen-dried, added with various proportion to the fodder of tilapia fries (Oreochromis niloticus x O. aures). After 29 days of experiment, our result showed that the fodder with 0.16% transformed yeast containing rGH promoted the growth of tilapia fries most prominently (P<0.01). Compared with control group, the weight gain rate and average body weight were 257% and 101% higher.
The experiments showed above indicate that using both E. coli and P. pastoris expressing systems, large amount of biologically active rGH could be obtained, and the qiantity of rGH in the transformed yeast could be measured. Furthermore, it is feasible to apply rGH to aquaculture by either immersing or feeding.
| Identifer | oai:union.ndltd.org:TW/087NTU00451010 |
| Date | January 1999 |
| Creators | Bing-Chang Lee, 李秉璋 |
| Contributors | H. J. Tsai, 蔡懷楨 |
| Source Sets | National Digital Library of Theses and Dissertations in Taiwan |
| Language | zh-TW |
| Detected Language | English |
| Type | 學位論文 ; thesis |
| Format | 55 |
Page generated in 0.0017 seconds