MOLECULAR MEDICINE I Flashcards
enzymes derived from bacteria
highly specific
recognise and cleave DNA at specific sequences
restriction endonucleases
endonuclease may have a symmetric cut with ___ ends
endonuclease may have a asymmetric cut with ___ overhang
endonuclease may have an asymmetric cut with ___ overhang
blunt ends
5’ overhang
3’ overhang
use of restriction endonucleases give rise to procedures such as
- _____
- _____
- _____
DNA fingerprinting
formation of recombinant DNA
DNA propagating
commonly used to separate and analyze the nucleic acids based on their size
gel electrophoresis
_______: recombinant DNA replicating within host cell
______________: In-vitro DNA amplication
cell-based
polymerase chain reaction
_________ is required to clone either cDNAs or copies of genes
vector
3 classes of vectors
P, L, Y
plasmids
lambda
yeast artificial chromosome (YAC) vector
circular DNAs found in bacteria
autonomous replication separate from the host genome
plasmid
STEPS IN CELL BASED DNA PROPAGATION (CLONING)
(4 STEPS)
R-T-S-P
- recombinant DNA (rDNA) production
- transformation (introduce the rDNA to the host microorganism)
- selection (select the microorganism that has been fully transformed)
- propagation and purification
DNA PROPAGATION
________: recombinant DNA replicating within host cell
________: in-vitro DNA amplification
cell based
polymerase chain reaction
method of amplifying a target sequence of DNA up to 600bp
requires DNA polymerase enzyme to remain stable at high temperature
polymerase chain reaction
3 steps of PCR
DD, DP, PE
dna denaturation
dna primer annealing
primer extension by Taq polymerase
_____________ gene expansion can be detected by PCR
Huntingtin gene