scholarly journals PRODUCTION AND BIOACTIVITY POTENTIAL OF THREE RECOMBINANT GROWTH HORMONES OF FARMED FISH

2010 ◽  
Vol 5 (1) ◽  
pp. 11 ◽  
Author(s):  
Alimuddin Alimuddin ◽  
Indra Lesmana ◽  
Agus Oman Sudrajat ◽  
Odang Carman ◽  
Irvan Faizal
Keyword(s):  
Growth Hormone ◽  
Body Weight ◽  
Protein Expression ◽  
Common Carp ◽  
Nile Tilapia ◽  
Expression Vector ◽  
Expression Vectors ◽  
Farmed Fish ◽  
Recombinant Growth Hormone ◽  
E Coli

This study was aimed to produce recombinant growth hormone (rGH) from giant grouper (Epinephelus lanceolatus), giant gouramy (Osphronemus gouramy) and common carp (Cyprinus carpio) and compare their bioactivity potential by means of inducing the growth hormone of juvenile Nile tilapia (Oreochromis niloticus) as the model. DNA fragment encoding mature GH protein of giant grouper (El-mGH), giant gouramy (Og-mGH) and common carp (Cc-mGH) was amplified by PCR method. The purified PCR products were ligated to pCold-1 to generate pCold/El-mGH, pCold/OgmGH, and pCold/Cc-mGH protein expression vector, respectively. Each of the expression vectors was transformed into the Escherichia coli BL21. E. coli BL21 was cultured using 2xYT medium and protein production was induced by cold shock at 15±1oC for overnight. The inclusion bodies of E. coli transformants containing protein expression vector were isolated by sonication method, and rGH production was analyzed by SDS-PAGE. Juvenile of Nile tilapia of average body weight of 12.41±3.28 g was intramuscularly injected once a week for 4 weeks with 1 μg inclusion body containing rGH per gram fish body weight. The result showed that rGH in molecular weight of about 25 kDa was obtained. Fish injected with rGH of El-mGH, Cc-mGH and Og-mGH grew 20.94%, 18.09%, and 16.99% faster, respectively, compared with the control. This result indicated that the three rGH produced in E. coli possessed biological activity when tested on Nile tilapia and further research is needed to find its effect on the growth of other aquaculture fish species.

scholarly journals Production and bioactivity test of recombinant protein common carp growth hormone

2011 ◽  
Vol 10 (1) ◽  
pp. 44
Author(s):  
Deny Sapto Chondro Utomo ◽  
. Alimuddin ◽  
Agus Oman Sudrajat ◽  
Irvan Faizal
Keyword(s):  
Escherichia Coli ◽  
Growth Hormone ◽  
Body Weight ◽  
Recombinant Protein ◽  
Common Carp ◽  
Cyprinus Carpio ◽  
Bacterial Cells ◽  
Recombinant Growth Hormone ◽  
E Coli ◽  
Sds Page

<p>This study aimed to produce recombinant growth hormone <em>(r</em>GH) protein of common carp (<em>Cyprinus carpio</em>) and evaluate its bioactivity. DNA fragment encoding mature GH protein of common carp (<em>mCc</em>GH) was amplified by polymerase chain reaction (PCR) method and PCR products were then ligated into pCold I to generate pCold I-<em>mCc</em>GH protein expression vector. <em>Escherichia coli </em>BL21 (DE3) harboring pCold I-<em>mCc</em>GH was cultured in the 2xYT medium at 15 °C for 24 hours and protein production was induced by isopropyl-beta-thio galactopyranoside (IPTG). The inclusion bodies containing rGH protein from <em>E. coli </em>transformants were isolated by sonication method and analyzed by SDS-PAGE. The result showed that rGH with molecular weight of about 25 kDa was obtained. Common carp juveniles with average body weight of 5.2±0.4 g were intramuscularly injected once a week for 4 weeks with rGH protein solution from 1 μg bacterial cells per gram fish body weight. The result showed that juveniles fish injected with rGH grew 106.56% higher than control. This result indicated that rGH produced in <em>E. coli </em>BL21 possessed biological activity and it may be useful to improve growth of aquaculture species.</p> <p>Key words: growth hormone, recombinant protein, common carp</p> <p> </p> <p>ABSTRAK</p> <p>Penelitian ini bertujuan menghasilkan protein rekombinan hormon pertumbuhan (<em>growth hormone</em>, GH) dari ikan mas (<em>Cyprinus carpio</em>) dan menguji bioaktivitasnya. Fragmen DNA penyandi protein matang (<em>mature</em>) GH ikan mas (<em>mCc</em>GH) diamplifikasi dengan menggunakan metode PCR dan hasilnya kemudian diligasi ke dalam pCold-I untuk menghasilkan konstruksi vektor ekspresi pCold-I-<em>mCc</em>GH. Plasmid pCold-I-<em>mCc</em>GH ditransformasi ke bakteri <em>Escherichia coli</em> BL21 (DE3), dikultur dalam media 2xYT cair pada suhu 15°C selama 24 jam dan produksi protein diinduksi dengan menggunakan isopropyl-beta-thio galactopyranoside (IPTG). Badan inklusi yang mengandung protein rekombinan GH (rGH) dari bakteri <em>E. coli</em> transforman diisolasi menggunakan metode sonikasi dan dianalisis dengan menggunakan SDS-PAGE. Hasil penelitian menunjukkan bahwa rGH dengan bobot molekul sekitar 25 kDa berhasil diproduksi. Benih ikan mas dengan bobot rata-rata 5,15±0,4 g diinjeksi secara intramuskular satu kali per minggu selama 4 minggu dengan larutan rGH hasil ekstraksi dari 1 µg pelet bakteri/g bobot ikan. Benih yang disuntik dengan rGH tumbuh sekitar 100% lebih cepat bila dibandingkan dengan kontrol yang tidak diinjeksi rGH. Hasil ini mengindikasikan bahwa rGH yang diproduksi dalam bakteri <em>E. coli</em> memiliki bioaktivitas dan dapat bermanfaat untuk memacu pertumbuhan spesies ikan-ikan budidaya.</p> <p>Kata kunci: hormon pertumbuhan, protein rekombinan, ikan mas</p>

scholarly journals Developmental and Evaluation of Advanced Expression Vectors with Both Enhanced Integration and Stable Expression for Transgenic Farmed Fish

2001 ◽  
Author(s):  
Zhanjiang John Liu ◽  
Rex Dunham ◽  
Boaz Moav
Keyword(s):  
Growth Hormone ◽  
Transposable Elements ◽  
Transgene Expression ◽  
Expression Vector ◽  
Transgenic Fish ◽  
Sleeping Beauty ◽  
Expression Vectors ◽  
Farmed Fish ◽  
Sleeping Beauty Transposon ◽  
Border Elements

The objectives of the project were to develop expression vectors using the Sleeping Beauty transposon technology and the genetic border elements to provide both enhanced integration rate and stable transgene expression, and to evaluate the application of such vectors in farmed fish such as catfish and carp. The panel recommended adding the objective of evaluating the endogenous transposable elements, particularly in catfish, in order to evaluate the applicability of the expression vectors while reduc1ng efforts in real production of transgenic fish considering the focus of the project was to develop the vector and evaluation of its applicability, not producing transgenic fish. Efficient production of transgenic farmed fish is hindered by two major problems: mosaicism due to delayed integration after single-cell stage, and silencing of transgene expression. In this project, we proposed to combat these problems by coupling the Sleeping Beauty transposon technology that can enhance integration rate and the border elements that can insulate transgene from position effect. Our major objective was to develop a new generation of expression vector that contains both of these elements. We have developed expression vectors containing both the Sleeping Beauty transposon signals, inverted repeats and direct repeats (IR and DR, respectively), and the border elements, scs and scs'. Growth hormone minigene has been cloned into this vector for applications of such vectors in growth enhancement. Luc reporter gene has been also cloned into this vector cascades for relative easy evaluation of transgene expression. Transgenic fish have been produced using these expression vectors in both catfish (US) and carp (Israel). Much effort was also devoted to evaluation of the endogenous transposable elements in catfish as recommended by the BARD grant panel. Multiple families of Tcl-like transposons were identified from catfish. Surprisingly, many Tc I-related transcripts were identified. Among these transcripts, both the sense and antisense transcripts were present. Some of the transcripts may be useful for development of novel transposase-based technology for aquaculture applications in the future. This project has both scientific and aquaculture implications. First, to develop expression vectors containing both IR/DR and scs/scs' repeated elements have been reported being extremely technically difficult due to excision of the repeated sequences by the E. coli host during cloning processes. We have successfully constructed this advanced vector that contained very complex cascades for both gene integration and gene regulation. We have produced transgenic fish using such vectors. This advanced expression vector should be useful for production of transgenic fish. By simply replacing the growth hormone gene, any gene of interest can be readily inserted in this vector. Thus this vector should provide technological possibility for early integration and stable expression of any economically important genes in aquaculture. We have also evaluated the applications of the Sleeping Beauty-based vectors in terms of the impact of gene size and found that the size of trans gene drastically affects transposition. The system will be only useful for transferring genes smaller than 5.6 kb. We have also identified novel transposase-related transcripts that may be useful for the development of novel transposase-based technologies for general scientific research and for aquaculture applications.  

Effect of supplementing Moringa oleifera essential oils on milk quality and fatty acid profile in dairy sheep

2019 ◽  
Author(s):  
N. Hemamalini ◽  
S. Ezhilmathi ◽  
A. Angela Mercy
Keyword(s):  
Recombinant Protein ◽  
Protein Expression ◽  
Cell Biology ◽  
Genetic Manipulation ◽  
Recombinant Protein Expression ◽  
Coli Strain ◽  
Expression Vectors ◽  
Functional Domain ◽  
Post Translational Modifications ◽  
E Coli

Escherichia coli is the most extensively used organism in recombinant protein production. It has several advantages including a very short life cycle, ease of genetic manipulation and the well-known cell biology etc. which makes E. coli as the perfect host for recombinant protein expression. Despite many advantages, E. coli also have few disadvantages such as coupled transcription and translation and lack of eukaryotic post-translational modifications. These challenges can be overcome by adopting several strategies such as, using different E. coli expression vectors, changing the gene sequence without altering the functional domain, modified E. coli strain usage, changing the culture parameters and co-expression with a molecular chaperone. In this review, we present the level of strategies used to enhance the recombinant protein expression and its stability in E. coli.

scholarly journals Expression and Inheritance of Growth Hormone Gene Constructs and Selective Breeding of Transgenic Farmed Fish

1994 ◽  
Author(s):  
Rex A. Dunham ◽  
Boaz Moav ◽  
Thomas Chen ◽  
Benzion Cavari
Keyword(s):  
Growth Hormone ◽  
Gene Transfer ◽  
Common Carp ◽  
Ictalurus Punctatus ◽  
Channel Catfish ◽  
Copy Number ◽  
Selective Breeding ◽  
Transgenic Fish ◽  
Farmed Fish ◽  
Aquaculture Production

Objectives: To accomplish stable expression, inheritance of transgenes and growth improvement in transgenic channel catfish, Ictalurus punctatus, and common carp, Cyprinus carpio, containing growth hormone (GH) genes, develop transgenic fish with all fish constructs, determine the relationships between copy number, expression and growth, determine the combined affect of selective breeding and gene transfer and assess environmental risk of transgenic fish. To develop mechanisms of triploidization for transgenic carp. Results: Performance of transgenic channel catfish was made uniform by selection. Growth of channel catfish and common carp was improved 40-50% more by combining gene transfer of GH genes with selection and crossbreeding than with either selection of crossbreeding. Growth improvement of transgenic catfish was not strongly correlated with copy number and expression levels. Progress was made in producting triploid transgenic common carp. Insertion of salmonid GH gene did not alter reproductive performance in channel catfish. Transgenic channel catfish grew no faster than controls when they had to forage on natural food and transgenic individuals were slightly more vulnerable to predation indicating that fitness of transgenic individuals in natural conditions is less than or equal to non-transgenic channel catfish. Contribution to Agriculture: These experiments are the first to demonstrate that transgenic fish can increase aquaculture production in the aquaculture production in the aquaculture environment. This research also demonstrated that maximum benefit of gene transfer in farmed fish is attained when combined with traditional selective breeding.

scholarly journals Growth and survival of giant gourami juvenile immersed in freshwater containing growth hormone

2013 ◽  
Vol 11 (2) ◽  
pp. 162
Author(s):  
Pustika Ratnawati ◽  
. Alimuddin ◽  
Harton Arfah ◽  
Agus Oman Sudrajat
Keyword(s):  
Growth Hormone ◽  
Growth Rate ◽  
Body Weight ◽  
Immersion Time ◽  
Egg Yolk ◽  
Recombinant Growth Hormone ◽  
Fish Hatchery ◽  
Growth And Survival ◽  
Controlled Conditions

<p>Growth of giant gourami is relatively slow. Administration of recombinant growth hormone (rGH) by several<br />times immersion in water containing 0.9% NaCl for one hour can significantly increase the growth. In this<br />study, giant gourami juvenile were immersed in freshwater containing 120 mg/L rGH to determine the long<br />immersion time that provides higher growth and survival. Juvenile at seventh-day-old after egg yolk absorbed<br />was immersed for 0.5, 1.0, 2.0, and 3.0 hours in rGH solution. As controls, fish was immersed in freshwater<br />without rGH (control 1: C1), and water containing 0.9% NaCl, and 120 mg/L rGH (control 2: C2). Fish was<br />kept under controlled conditions in 20 L aquarium for three weeks, then in 50 L aquarium until at end of<br />experiment, and fed at satiation. The results showed that highest growth in body weight, survival, and biomass<br />(p&lt;0.05) were obtained in rGH immersion treatment for 0.5 hour. Biomass of fish immersed by rGH for 0.5<br />hour was about 32% higher compared to control C1, and 13% compared to C2. Survival of the 0.5 hour rGHimmersed<br />fish (100%) was similar to control C1 and C2. Thus, immersion of giant gourami juvenile in<br />freshwater containing rGH for 0.5 hour allowed highest growth rate, and this method is relatively easy to be<br />adopted by fish hatchery.</p><p>Keywords: immersion, hormone, growth, freshwater, giant gourami</p>

scholarly journals PRODUKSI PROTEIN REKOMBINAN HORMON PERTUMBUHAN IKAN KERAPU

2012 ◽  
Vol 7 (2) ◽  
pp. 231
Author(s):  
Irvan Faizal ◽  
Ratu Siti Aliah ◽  
Muhammad Husni Amarullah ◽  
Novi Megawati ◽  
Sutanti Sutanti ◽  
...  
Keyword(s):  
Growth Hormone ◽  
Inclusion Bodies ◽  
Recombinant Growth Hormone ◽  
E Coli

Salah satu spesies ikan yang menjadi target produksi perikanan budidaya nasional adalah ikan kerapu tikus (Cromileptes altivelis). Ikan kerapu tikus merupakan ikan laut budidaya komoditas ekspor, namun laju pertumbuhannya sangat lambat. Oleh karena itu, perlu dilakukan usaha budidaya yang mampu menaikkan laju pertumbuhan ikan kerapu tikus. Pendekatan nutrisi melalui penggunaan hormon pertumbuhan (Growth Hormone, GH) pada usaha budidaya diyakini mampu meningkatkan kecepatan tumbuh ikan budidaya. Pada penelitian ini, melalui teknologi DNA rekombinan telah dilakukan isolasi dan identifikasi gen GH yang selanjutnya dilakukan produksi protein rekombinan GH (recombinant Growth Hormone, rGH) dengan memanfaatkan penggunaan bakteri E. coli. Dalam pelaksanaan penelitian dilakukan konstruksi rGH yang menghasilkan bakteri E. coli BL21 (DE3) yang mampu memproduksi protein rGH. Produksi rGH dilakukan pada skala bioreaktor. Proses isolasi produk rGH-nya dalam bentuk pellet inclusion bodies yang selanjutnya dicampur dengan pelet pakan komersil hingga konsentrasi akhir protein dalam pakan mencapai 1 ng/6 mg pakan, di mana setelah dikering-anginkan, pelet pakan protein rekombinan GH dapat diaplikasikan untuk budidaya ikan kerapu tikus dan ikan budidaya lainnya.

Recombinant growth hormone in growing lambs: effects on growth, feed utilization, body and carcass characteristics and on wool growth

1989 ◽  
Vol 40 (1) ◽  
pp. 195 ◽  
Author(s):  
AS Zainur ◽  
R Tassell ◽  
RC Kellaway ◽  
WR Dodemaide
Keyword(s):  
Growth Hormone ◽  
Body Weight ◽  
Recombinant Dna ◽  
Plasma Concentrations ◽  
Recombinant Growth Hormone ◽  
Linear Change ◽  
Wool Growth ◽  
Quadratic Response ◽  
The Mean ◽  
Plasma Gh

Groups of six crossbred lambs, three wethers and three ewes, about 22 kg liveweight (LW), were injected subcutaneously with 0, 50, 150 and 250 pg (kg LW) d a y 1 of recombinant DNA-derived bovine growth hormone (rebGH) for 60 days. From blood sampled twice weekly 4.5 h after injection, the mean plasma concentrations of growth hormone (GH) (P<0.01) and insulin (P<0.05) were significantly increased. The mean concentrations of GH were 2.9, 7.1, 9.4 and 8.8 8g/l, while insulin concentrations were 24.2, 29.5, 46.8 and 68.2 mU/l for the respective treatments. Maximum concentrations of plasma GH were recorded about 3 h after injection. Rates of liveweight gain were 180, 217,238 and 261 g/day (P<0.001), fccd conversion ratios were 5.71,4.97,4.66 and 4.05 g feed/g gain ( P < 0.001), and warm carcase weights were 14.4, 16.4, 16.6 and 16.8 kg (P < 0.0 1) for the respective treatments. The linear change in warm carcase weights became a quadratic response (P<0.05) when adjusted to an empty body weight (EBW) of 28.8 kg. Wool growth was not affcctcd by treatments. Of the non-carcase components, major organs including the head (P<0.01), feet (P<0.05), heart (P<0.05), liver and gallbladder (P < 0.05), reticula-rumen (P < 0.05), omasum (P<0.05) and blood (P<0.01) showed linear increases in weight with increasing dosage of rebGH, but no significant differences when data were adjusted to a standard EBW. The chemical composition of the carcase showed that crude protein increased linearly (P<0.001) with increasing doses of rebGH. When adjusted to 28.8 kg empty body weight, there was 16.0�/o more carcase protein (P<0.01) with 250 than with 0 8g (kg LW)-1 day-1 of rebGH.

scholarly journals Performance of Nile tilapia juvenile fed diet containing different dosages ofrecombinant common carpgrowth hormone

2013 ◽  
Vol 11 (1) ◽  
pp. 17
Author(s):  
Dian Hardiantho ◽  
. Alimuddin ◽  
Arief Eko Prasetiyo ◽  
Dwi Hany Yanti ◽  
Komar Sumantadinata
Keyword(s):  
Growth Hormone ◽  
Body Weight ◽  
Nile Tilapia ◽  
Rt Pcr ◽  
Commercial Diet ◽  
Commercial Feed ◽  
Igf I ◽  
And Control ◽  
Average Body ◽  
Average Body Weight

<p>The studywas conductedtodetermine thedose of recombinant common carp growth hormone (r<em>Cc</em>GH) supplemented in the commercial diet that generates the best performanceonbodyweight, biomass, andfeed conversion rate of juvenile tilapia.The dose of r<em>Cc</em>GH administered was10, 20, and30mg/kg of commercial feed, andnorGH supplementation as control. Eachtreatment was designed in triplicates. Juveniles at average bodylengthof about2cm(average body weight of 0.7 g) were reared in a density of25fish/m<sup>2</sup>, for three weeks in hapa net at a size of 2×2×1 m<sup>3</sup>. Fish were fed diet containing r<em>Cc</em>GH twicea week with an interval ofthreedays. The juvenile were fed with the rCcGH supplemented diet three times daily at satiation. The results showedthat average body weight andbiomass of fish treated by r<em>Cc</em>GH with the doses of 20 and 30 mg/kg feed was higher (p&lt;0.05) than that of 10 mg/kg and control. In addition, feed conversionrate of 20 and 30 mg/kg feed treatments was lower (p&lt;0.05) than that of 10 mg/kg and control. RT-PCR analysisshowedthat expression level of IGF-1 gene in juvenile liver was increased in parallel with the increasing of rCcGH dosages supplemented in diet.This suggestedthatIGF-1 plays an important rolein the induction ofgrowth,andfeed conversionefficiencyof tilapiafed diet containing r<em>Cc</em>GH. The bestperformance of juvenile can be obtainedby feeding with diet containingr<em>Cc</em>GH20‒30mg/kgof feed.</p> <p>Keywords: rCcGH, IGF-I, recombinant growthhormone,performance, Nile tilapia.</p>

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