unit 8 Flashcards
(21 cards)
what is PCR
-form of artifical dna replication
-this is used to amplify ( to increase the amount of) dna, so it can be analysed
what makes pcr very useful
only a small sample of it is required
state when PCR is used
-used in forensic dna analysis
-used in analysis of dna for genetic diseases
-used before dna sequencing
-used before making a genetic fingerprint
-used to clone genes in vivo
what happens before pcr
proteaze enzymes are added to the dna sample to purify the dna
what does pcr require
theromycler
-dna nucleotides, all 4 types to synethesie the dna strands
-primers-short fragments of single stranded dna strands that are comp to the bases at start and end of dna fragment
-dna polymerase, used to join dna nucleotides togethor, taq is used as its thermostable and wont denature at high temps
-dna fragment- the sample we wish to copy
state the two uses of primers
-they stop the original two strands of dna from re joining after separation
-they create double stranded sections of dna at the start of dna fragments to allow dna polymerase to attach and start building new complementory dna strands
state the process of pcr
95- high temperature breaks hydrogen bonds between complementory bases, seperating two strands, both single stranded dna acts as strands
55-temperature drops to 55 allowing primers to bind to the strands reforming hydrogen bonds
72-optimum temp for taq dna polymerase, enzymes joins nucleotides togethor by forming phosphodiester bonds to form new complementory strands
Primers are used in PCR – describe what they are
short, single strands of dna that are complementory to the bases at the start and end of dna fragments
Explain the role of the primers used in PCR.
-stop the orignal two strands of dna from re joining
-creates double stranded sections of dna so dna polymerase can attatch so it can synthesis dna
Explain why 2 different primers would be required.
different base sequences at the start and end of dna fragment
primers have to have complementory base sequence
state problems of PCR
sometimes does not produce the expected number of dna fragments, due to
-recation may run out of primers
-reaction may run out of free nucleotides
-high temps damage dna bases
-dna strands may re join after seperation
Suggest one use of the polymerase chain reaction. (1)
forensic analysis of dna
Give two ways in which the polymerase chain reaction differs from the process of transcription. (2)
in t rna used in p dna used
in t one strand act as templates in p both strands act as templates
Tissue traces from a crime scene often need to be identified. DNA from the tissue is ‘amplified’ by the
polymerase chain reaction (PCR) to get samples large enough for further analysis.
Modern PCR technique uses DNA polymerase from the bacterium Thermus aquaticus. Why is this enzyme
chosen? (2)
as it thermostable
so will not denature at high temps
so pcr can continue
It is important that the fragments of DNA used in PCR are not contaminated with any other biological
material. Suggest a reason why.
-biological molecules also contain dna
-therefore this dna would get amplified
How is PCR different to DNA replication? 3 reasons
used taq dna, uses normal dna
needs primers, does not need primers
uses high temp to break h bonds, uses dna helicase to break h bonds
genome
proteome
complete set of genes in an organism
variety of proteins produced by a cell
what is genetic engineering
involves trnafer of dna fragments from one species/organism to another
what if GE in vivo
gene is copied within a living orgnaism as organism divies it replicates the dna creating copies of the gene
state the steps of genetic engineering
-isolate gene that codes for desired protein
-insert gene into vector to make recombinent dna
-insert recombinant dna into host cells
-identification of succesfully transformed host cells using marker genes
-cloning of transformed cells
