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Bio Exam II

DNA Tech

PCR: Polymerase Chain Reaction - generates a lot of copies of DNA in short amount of time - Important when studying genes
PCR Components - Template DNA - primers - dNTPs ( dideoxynucleosides) A, T, C, G - Taq. polymerase
Process of PCR- done in thermal cycle to modulate temp. of mixture 1) denature: heat to 90 degrees C , breaks h-bonds in double helix 2) annealing: 40-65 degrees C, primers bind w/ complementary sequence in template stand 3) Extension: Taq. polymerase replicates DNA from 3'-5' end of primers. Strand become template
Gel Electrophoresis
Agarose Gel - polysaccharide gel - porous - passes electrical current through causing DNA to move from - to + end - Uses: confirms successful PCR, DNA sequencing, DNA profiling, Disease Diagnosis
Deoxyribonucleotide chain termination sequencing - 1st automated sequencing method - Sanger: won Nobel prize in 1980 -can sequence DNA fragments to about 1000 bp. - same process as PCR but w/ one major diff
Ingredients for Deoxyribonucleotide chain termination squencing - target dna - primer - DNA pol. -4 deoxyribonucleotides -4 dideoxyribonucleotides
Dideoxyribonucleotides - missing the 3' hydroxyl group making in responsible for chain termination because the phosphate cant bind
Process of Deoxyribonucleotide chain termination squencing - target DNA denatured - complementary strand synthesized - deoxyribonucleotide randomly incorporated - elongation stops (termination) at dideoxyribonucleotide
Polyacrylamide Gel - made from polymer - DNA fragments put in get and they migrate based on size (shorter strands move faster) - Detector reads bases as they migrate
Uses of sequencing - evolutionary relationships - determine loci & function - disease diagnosis
Applications - species diversity: know organisms in community, otherwise very hard to measure small & rare organisms - DNA bar coding: sequence of small region of genome to identify species
Gene Therapy - FDA approved gene therapy to cure type of genetic blindness; uses a retrovirus to insert correct genes into cell of retina
CRISPR - silence or insert genes
Created by: tennadeleta.
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