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Restriction Enzymes

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Published in: Biology
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Restriction Endo-nucleases for cleavage of Bacterial and Genomic DNA

Mukul D / Delhi

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Qualification: Bachelor's of Engineering

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  1. RESTRICTION ENZYMES MADE BY MUKUL DHINGRA 81 3/BT/1 2
  2. Restriction Enzymes Restriction enzymes are DNA-cutting enzymes found in bacteria (and harvested from them for use). Because they cut within the molecule, they are often called restriction endonucleases
  3. WORKING OF RESTRICTION ENZYMES DIGESTION OF DNA NON9EClFlC BY RESTRICTION ENDONUCLEASE DNA FRAGMENTS
  4. NOMENCLATURE Derivation of the EcoRI name Abbreviation co Meaning Escherichia Co/i RYI 3 Description Genus species Strain First identified order of identification
  5. WHY DO WE NEED TO CUT DNA? In order to be able to sequence DNA, it is first necessary to cut it into smaller fragments. The tools for this are the restriction endonucleases. The rarer the site it recognizes, the smaller the number of pieces produced by a given restriction endonuclease.
  6. Recognition of Cutting Site A restriction enzyme recognizes and cuts DNA only at a particular sequence of nucleotides. For example, the bacterium Hemophilus aegypticus produces an enzyme named Haelll that cuts DNA wherever it encounters the sequence 5'GGCC3' or 3'CCGG5' The cut is made between the adjacent G and C and the fragments can be separated from one another. So sequence of each is determined.
  7. Cutting of DNA by ENZYMES Barn" 3* 5* 5* EcoRl 3* and Bam"' I-lindlll RESTRICTION 3* GVc c G T c T R. c. T R G R c G R c. R. prod uce blunt ends produce "sticky* ends and
  8. END CHARACTERISTICS Haelll and Alul cut straight across the double helix producing "blunt" ends. The ends of the cut have an overhanging piece of single-stranded DNA. These are called "sticky ends" because they are able to form base pairs with any DNA molecule that contains the complementary sticky end.
  9. APPLICATION OF RESTRICTION ENZYMES Mixed together, these molecules can join with each other by the base pairing between their sticky ends. The union can be made permanent by another enzyme, a DNA ligase, that forms covalent bonds along the backbone of each strand. The result is a molecule of recombinant DNA (rDNA).
  10. APPLICATIONS Isolated restriction enzymes are used to manipulate DNA for different scientific applications. They are used to assist insertion of genes into plasmid vectors during gene cloning and protein expression experiments. Restriction enzymes can also be used to by specifically distinguish gene alleles recognizing single base changes in DNA known as single nucleotide polymorphisms (SNPs).
  11. THANK YOU

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