GROWTH HORMONE

PRODUCTION OF HUMAN GROWTH HORMONE(hGH)BY RECOMBINANT DNA TECHNOLOGY:

• The gene for human growth

hormone (hGH) is isolated from
human pituitary gland.
• Insertion of whole hGH gene into
plasmid vector and cloning into E.coli
results into production of biologically
inactive hormone because bacteria can
translate the region of gene that are
not translated in human thereby
producing a prehormone containing an
extra 26 aminoacids which might be
difficult to remove.
• Hence the segment of gene that codes
for the first 24 aminoacids of hormone
is constructed chemically from blocks
of nucleotide.
Step I: Chemical synthesis of gene for first 24 aminoacids:
• From the known aminoacids sequence of hGH, gene for first 24 aminoacids
are constructed chemically. These genes are constructed in three small
fragments and then they are joined by T4 DNA ligase to get whole gene for
first 24 aminoacids.
Step II: Isolation of mRNA for hGH
• In this step mRNA for hGH is isolated from human pituitary gland tissue.
Step III: Reverse transcription
• Using reverse transcriptase enzyme complimentary DNA (cDNA) is
synthesized from mRNA.
• The cDNA obtained by reverse transcription process, is the gene for hGH.
• The full gene is cut with restriction endonuclease enzyme to remove first 24
gene.
Step IV: Joining of synthetic gene and cDNA
• In this step synthetic gene (gene for first 24 aminoacids) and cDNA are joined in order to
obtain full gene with its own initiation codon (AUG). T4 DNA ligase join these genes.
Step V: selection of suitable vector and recombination:
• Expression vector phGH407 derived from plasmid vector PBR322 is used as carrier
vector.
• HGH gene is ligated into a restriction site just downstream of Lac; promotor/operator
region of the expression vector.
Step VI: selection and recombination into suitable host cell
• E. coli is used as suitable host cell.
• The recombinant expression vector is then transformed into E.coli.
• The recombinant E. coli then starts producing hGH.
• The recombinant E. coli are isolated from the culture and mass production by
fermentation technology to obtain hGH.
SUMMARY STEP:
• hGH isolated from pituitary gland.
• Gene is cleaved using restriction endonuclease.
• Add gene to a plasmid vector (carrier molecule) by using ligase
• contruct with desired gene is obtained called rDNA.
• Plasmid under self-replication.
• Introduces into E.coli cell.
Simplified of the procedure:
• Synthesis of first 24 nucleotides chemically with known sequence. (8x3= 24)
• Isolate growth hormone from human pituitary gland.
• cDNA is formed using reverse transcription- gene for hGH production.
• First 24 sequences is removed using restriction endonuclease.
• Synthesis gene and cDNA are joined using T4 ligase.
• Suitable vector is chosen
• DNA joined to plasmid by endonuclease and ligase.
• Kept at Lac Operon- Operator region.
• Selection of the host cell which is E.coli .
• Recombinant E.coli isolated from culture and mass production by fermentation
technology to obtain hGH.
Use of recombinant human growth hormone (hGH)
1. Treatment of children suffering from 6. Atletic supplement
growth deficiency 7. Grwoth deficiency
2. Treat the patient with Tumer’s 8. Tumer’s syndrome and chronic renal
syndrome and chronic renal insufficiency treatment
insufficiency
9. Renal carcinoma treatment
3. To treat patient with renal carcinoma
10. Bovine somatotropin-increases milk
4. Bovine somatotropin hormone is production in lactating cows.
used to increases milk production in
lactating cows and also to increase
body mass of cattles
5. Growth Hormone deficiency
treatment
• Time conservation
Advantages
• Cost effective
• Cheap
• There are more copies due to recombinant technology

Limitation
• Transformation of diseases
• Bone and heart problem
• Fatigue
• Muscle weakness
• Risk of type 1 diabetes