Gene probes, preparation of chimeric DNA molecules, cloning

This is a presentation of Biochemistry and Molecular Biology entitled "Gene probes, preparation of chimeric DNA molecules, cloning" which was fundamental for my Advanced Biochemical Technique course.

1. Gene probes, preparation of chimeric DNA molecules, cloning Md. Hanzala Rahaman , MS Student, BAU

2. Outline Gene probes Working of gene probes Application of gene probes Chimeric DNA Cloning Types of Cloning Gene Cloning Reproductive cloning Artificial embryo twinning Somatic cell nuclear transfer Therapeutic Cloning Advantages and disadvantages of Cloning

3. Gene probes S ingle-stranded DNA or RNA fragment U sed in genetic engineering S earch for a particular gene or other DNA sequence

4. Gene probes Base sequence complementary to the target sequence A ttached by base pairing Used in the Southern blotting  technique

5. Working of gene probes

6. Application of gene probes Identify a particular DNA sequence in a complex mixture Identify the presence of a pathogen in a sample Identify microorganisms compared to the traditional methods

7. Chimeric DNA Means of the cloning genes or other small DNA sequences Insertion of a foreign DNA segment into a DNA molecule of the vector Three techniques available for the preparation of chimeric DNA

8. C loning In simple terms cloning designates duplicating In biological sense, clone means genetically identical cell or organism In biomedical research, cloning is broadly defined to mean the duplication of any kind of biological material for scientific study, such as a piece of DNA or an individual cell.

9. Types of Cloning Cloning is basically of three different types Gene cloning Reproductive cloning Therapeutic cloning

10. Gene Cloning copy of particular gene or an exact segment of DNA DNA that need to be cloned is extracted from the specific organism purified and cut into smaller pieces using restriction enzyme enzymes create small fragments of specific DNA with cohesive ends This cohesive ends help in attachment of this DNA with other strand of DNA which is cut using same restriction  enzyme the obtained fragments of DNA are integrated into plasmid

11. Gene Cloning (Cont.) Plasmid cleaved using same restriction enzyme as that in DNA; it generates the same cohesive ends cohesive ends of DNA segment and plasmid gets linked by DNA ligase transformed plasmids are incorporated into bacteria that act as host organism by the process of transformation different plasmids contain different section of targeted DNA, inserting it into host helps in creating a library, termed as DNA library Finally when placed in agar medium, the identification and investigation of colonies help in detection and extraction of desired gene.

12. Reproductive cloning copy of complete organism is termed as reproductive cloning The best example of reproductive cloning is Dolly the sheep It was the first successfully done reproductive cloning

13. Reproductive cloning Two ways for reproductive cloning Artificial embryo twinning Somatic Cell Nuclear Transfer (SCNT )

14. Artificial embryo twinning T he natural method of cloning The only difference being the method used in petri dish instead of mother womb The early embryo is divided into cells and allowed to divide and develop in petri dish Then it is injected into surrogate mother where they complete the developmental process Since all the embryos came from same fertilized egg therefore it turns out to be identical

15. Somatic cell nuclear transfer This was the method used to create dolly the sheep Firstly, the somatic cell is isolated from an adult organism suppose it as A and DNA is extracted from it Secondly, from the egg cell of another adult organism B, nucleus and DNA is completely removed DNA obtained from A is fused with egg cell from sheep B This generates a clonal embryo that is kept under the laboratory environment for some time this cloned embryo is placed in the uterus of female organism also called surrogate organism B irth of clone of organism A has contributed somatic cell

16. Somatic cell nuclear transfer

17. Therapeutic Cloning Therapeutic cloning and reproductive cloning are almost similar The only difference is the end product In case of reproductive cloning, the clone is allowed to develop and live whereas in case of therapeutic cloning the clone is destroyed to obtain the stem cell from it. Therefore , the clone is allowed to grow for some days and then killed to conduct research on embryonic stem cell The advantage of this cloning is it helps in replacement of injured and diseases tissues of human body.

18. Advantages of C loning Replacement of organs. Helps infertile couple to have children. Introducing desired character in the plants. It is a cure for several diseases.

19. Disadvantages of Cloning Possibility of introducing new diseases. High chances of organ rejection. Decrease in gene diversity. Unbalanced society.

20. "Cloning is great. If God made the original, then making copies should be fine. --- Douglas Coupland Thank You