D1.1 DNA replication Flashcards
What is semi conservative replication
when one strand comes from the original chromosome and the other is newly synthesized at the end of DNA replication to form a new double helix
Each original strand forms a new helix with a new strand
What is conservative replication
when an entirely new double helix is formed alongside the original with no unzipping of DNA
1 helix is completely original
1 helix is completely new
How can a high degree of accuracy be maintained when copying base sequences
through complementary base pairing and semi conservative replication
what does helicase do
unwinds the DNA double helix at one region, breaking hydrogen bonds and temporarily keeping the strands separated
how do both strands of DNA act as templates in replication
complementary nucleotides line up opposite each base on the exposed strands
hydrogen bonds then form
What are the purposes of DNA polymerase
catalyses a condensation reaction between sugar and phosphate groups of nucleotides, forming the new strands
also proofreads the new DNA for mistakes
what are the uses for analysing DNA to identify individuals
forensics and genetic relatedness
What is the use of polymerase chain reaction
they allow small fragments of DNA to be copied repeatedly in a fully automated process
What enzyme is used in PCR
a heat tolerant DNA polymerase enzyme known as tag polymerase
How can amplified DNA be separated
through gel electrophoresis, allowing genotypes of different individuals to be compared
what are the limitations of PCR
knowledge of the amino acid sequence of a desired genes is needed to synthesize nucleotide primers
non target sequence may be amplified because short nucleotide sequences may not be specific enough
what are DNA primers
short sequences of single stranded DNA, made synthetically with base sequences complementary to one end of DNA
what is gel electrophoresis
a process thats used to separate molecules such as proteins and nucleic acids
important for DNA sequencing
What is the separation of nucleic acid fragments/ proteins in gel electrophoresis due to
migration of these molecules through a sieve like medium ( agarose gel), essentially size
the electrical charge that molecules carry ( phosphate has negative charge, so migrate to positive side), basically charge
how can fragments be identified after separation in gel electrophoresis?
through gene probes and DNA stains
what is a gene probe
single stranded DNA that is complementary to that of a particular gene, must be radioactive so it can be identified
what is a DNA stain
locate the position of all DNA fragments once applied
stains can be ethidium bromide, or methylene blue
what is genetic profiling
used to identify organisms, species or individuals using DNA
uses both PCR and gel electrophoresis
what are Variable number tandem repeats
VNTR’s are short base sequences that show variation between individuals in terms of number of repeats
they are major lengths of non coding DNA used in genetic profiling
features of VNTRs
dont code for proteins are scattered throughout the length of the DNA molecule in clusters
half comes from each parent
everyone has unique sequence of nucleotides in our DNA
how are genetic profiles produced
a sample of DN is cut with a restriction enzyme that acts close to the VNTR region
electrophoresis then used to separate fragments, resulting in a pattern of bands
what is the process of southern blotting
copy of distrbuted DNA fragments on nylon membrane ( through southern blotting)
heat treatment of nylon membrane binds DNA to it
DNA probes added to bind to particular DNA
Nylon membrane overlaid with xray film which is fogged by emissions from probes
film is developed to show position of fragments
what are the applications of genetic fingerprinting
identification of suspects
identification of corpses
determining paternity
Which way does DNA polymerase move along the strand
synthesizes in 5’ to 3’ direction and moves along the template strand in the opposite direction
why does DNA polymerase move in a certain direction
because otherwise it would be unable to proofread, allows it to function as a self correcting enzyme
what is replication initiated by
an RNA primer
what are replication forks
semi conservative replication is initiated at many points along the double helix, known as replication forks
what is the result of enzyme helicase
bubbles form due to hydrogen bonds being split apart
what does DNA gyrase do
assists in reducing tension and prevents separate strands from rejoining
what do single strand binding proteins do
they attach and prevent separate strands from rejoining, allowing unwound section to act as templates for the synthesis of complementary DNA strands
why do DNA strands only elongate in the 5’ to 3’ direction
because nucleotides can only be added to the free 3’ direction
what happens in the leading strand
exposed 5’ to 3’ is the leading strand
DNA polymerase III adds nucleotides to free 3’ end in same direction as replication fork
continous process as helicase continues to move and new template is exposed
what is the function of the primer
a short length of RNA that is synthesized by primase where the new DNA can start from ( initial nucleotide chain)
what happens in the lagging strand
replication is discontinuous
strands synthesized in opposite direction
Okazaki fragments are formed, each primed separately
allows both strands to replicate at the same time
what are Okazaki fragments
shorts lengths of DNA created due to discontinuous process
what is the function of DNA ligase
present in the lagging strand to join the fragments together
function of enzymes in DNA replication
RNA primer formed by primase and then DNA polymerase III attached nucleotides to the RNA primer, forming a fragment
DNA polymerase I replaced the RNA nucleotides at the start of each fragment with DNA nucleotides
Enzyme DNA ligase joins the Okazaki fragments together
How is DNA polymerase III involved in DNA proofreading?
removes any mismatched base from the 3’ terminal which is then replaced with the correct nucleotide
ensures any wrong nucleotides are removed before DNA replication proceeds
How is proofreading carried out
by a nuclease that cleaves the phosphodiester bond
