RECOMBINANT DNA TECHNOLOGY
it is a method used in genetic engineering to manipulate DNA to produce a desired trait.
RECOMBINANT DNA TECHNOLOGY
who and when is the DNA technology invented
1973 by HERBERT BOYER AND STANLEY COHEN
what is the first plasmid used
pSC101
important feature of the pSC101
IT HAS A SINGLE REGION WITH GAATTC SEQUENCE AND THE PRESENCE OF TETRACYCLINE RESISTANCE GENE
who invented gene splicing
PAUL BERG
participants in recombinant DNA
ENZYME RESTRICTION ENDONUCLEASE, DONOR DNA AND VECTOR DNA
example of restriction endonuclease
EcoR1
recognizes and cleaves the GAATTC
EcoR1
importamce of the restriction endonuclease
serves as a scissor that cuts the donor and vector dna at the very specific site that contains palindromes
these are sequences na kahit pagbali baiktarin mo, same parin
PALINDROMES
it has the ability to attract complementary bases
STICKY ENDS
it contains the gene of interest where palindromic sequence is nandon
DONOR DNA
carrier of the gene of interest
VECTOR DNA
common type of vector DNA
PLASMID
major roles of the vector DNA
PRESENCE OF ORIGIN OF REPLICATION AND HAS MARKERS THAT SERVES AS A GUIDE
it is a guide to locate the recombinant DNA later
GENETIC MARKERS
2 types of vector DNA
pBR322 and pUC18
pBR22 has ...
Psti and BamHI
it has ampicillin resistance gene
Psti
it has tetracycline resistance
BamHI
it is made up of 2.7 kilobases with ampicillin resistance gene and LacZ marker
pUC18
it is for the synthesis of galactosidase and contains poly linker
LACZ
it breaks down the x gal and release blue color once done
GALACTOSIDASE
it contains several restriction sites / palindromic sequence
POLY LINKER
steps in recombinant dna technology
1. ISOLATE THE DONOR AND VECTOR DNA
2. CUT THE DNAs WITH RESTRICTION ENYME
3. COMBINE THEM WITH DNA LIGASE
4. HOST CELL TRANSFORMATION THROUGH CHEMICAL TRANSFORMATION OR ELECTROPORATION
5. RECOMBINANT DNA CLONING
6. SCREENING OF THE TRANSFORMED CELL (EX: BLUE WHITE SCREENING)
it is the process where you force the bacterium by exposing through heat temp
CHEMICAL TRANSFORMATION
it uses electricity to increase the permeability of the cell membrane allowing the molecules to pass through
ELECTROPORATION