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Gene Cloning Application (Gene Subtraction)

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Faizah Rahman

on 28 January 2015

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Transcript of Gene Cloning Application (Gene Subtraction)

The Overall Process
The Theory
Introduction
Target gene & Engineered Characteristic

Polygalacturonase - Delay of fruit spoilage

Polyphenol oxidase - Preventation of discoloration in fruits and vegetables

Starch synthase - Reduction of starch content

Chalcone synthase - Modification of flower color

About Gene Cloning
Gene Subtraction
Application of Antisense RNA Technology in Gene Subtraction
Gene Cloning Application
(Gene Subtraction)

Gene cloning is the process in which a gene of interest is located and copied (cloned) out of DNA extracted from an organism.

1. Isolation
2. PCR
3. Purification
4. Ligation
5. Transformation
6. Plasmid extraction
7. Plasmid digestion
8. Gel electrophoresis
Application of Gene Cloning
Production of recombinant protein
-Insulin
-Interferon
-Vaccines
-Human Growth Hormone
-Gene Therapy
-Human Genome Project
Application in Forensic Science
-DNA Fingerprinting
-Paternity Test
-Sex identification
Definition of Gene Subtraction
> Inactivation of a target gene, not removal
> Is done by
antisense technique
> RNA produced by this technology is called the
antisense RNA (asRNA)
Antisense RNA
Antisense RNA is a single stranded RNA that is complementary to the messenger RNA(mRNA) transcribed within a cell.

Antisense RNA may be introduced into a cell to inhibit translation of a complementary mRNA by base pairing to it and physically obstructing the translation machinery
Antisense RNA
Vs
RNA Interferons
The intention of both technology is the same which is gene silencing but the process is a bit different.
Antisense technology degrades RNA by enzyme RNaseH
RNAi utilizes the enzyme Dicer & RISC(RNA-inducing Slicing Complex) to degrade mRNA.
Antisense Therapy
Mode of treatment for genetic disorder & treatment

Antisense drugs are being used in research to treat cancer, HIV(Human Immunodeficiency Virus), CMV(Cytomegalovirus) etc.
Challenges
Difficult to get into the body
, i.e for Huntington's Disease it must cross the blood brain barrier
Inevitable toxic effect
. Eventough it is very specific, it can still regulate both mutant and normal Huntington Disease alleles.
Conclusion
The antisense RNA technology in gene subtraction shows the potential for diverse application to basic research and therapy.
References
H.D, Kumar. (2007). A textbook of biotechnology 2nd edition. Selangor, SL. Pearson Malaysia Sdn Bhd.
Nakashima, Nobutaka; Goh, Shan; Tamura, Tomohiro.(2012).Application of biotechnology. Selangor, SL. Pearson Malaysia Sdn Bhd
Gene subtraction.(2013). Retrieved January 14 2015, from
www.ncbi.nim.com(pubM ID 17173627).
Addition and subtraction of gene. (2014). Retrieved on January 15 2015, from
www.nature biotechnology.com.
Full transcript