4A-enzymes that manipulate DNA Flashcards
restriction enzymes/ restirction endonucleases
restriction enzymes cut DNA into smaller fragments in order to isolate the gene of interest.
circular DNA produces 1 less fragemnet then cut double check than linear dna eg. circular dna 3 cuts= 3 frgaemets but linear dna 3 cuts is 4 fragements
There are 100 base pairs in a DNA strand, 20 are A and 20 are T, how mamny are C and G.
NOTE if theres 100 base pairs that meanstheres 200 nulcoeitide,s which means that A=20, T=20 andwhats left ove ris 80 and 80 so
C=80 and G=80
recognition site
specific target sequence of the DNA that restriction enzyme cuts
what kind of bonds between nucleic acids
phosphodiester bond
restriction endonucleases also known as
restriction enzymes or molecular scissors because restrction endonucleases are cutting down the molecuels
where are endonucleases from
mostly from bacteria
EcoRI
is a restriction endonuclease enzyme isolated from species E. coli
overhanging nucleotides
unbonded nucleotides on the ends of the DNA strand
circular DNA
plasmid
compare thecuts between linear and circular dna
linear DNA has one more than circular DNA–the linear DNA has the same one more strand than the amount of cuts. eg 3 cuts linear dna will have 4 fragemnets, circular dna will have 3 cuts
ligase
enzymes that join fragments of DNA, forming a phosphodiester bond after the restricton enzymes cut DNA
why is it that the ligase is called a molecular glue, whilst restriction endo nucleases are known as moleculear scissors
ligase joins 2 fragments of DNA, whilst restrction endonuclease cut the sequences of DNA
types of ligase
DNA ligase
RNA ligase
differnece between polyermase and ligase
polymerase forms DNA and RNA from nucleotides, whereas, ligase joins DNA or RNA fragments together
primer
a short strand of nucleic acids that guides the polymerase to the right place to start copying the DNA and is the starting pointto which polyermase attahche
in the DNA sequence, the primer is always
at the beginning of the sequence, there should be a distinction to show where the primer is
where are restriction enzymes foudn
resctrictionenzymes are foundin bacteria
where do restriction enzymes cut and how do restriction enzymes cut
restriction enzymes cut a specific recognition site(code)
restriction enzymes cut specific reocngintion sites by cleaving the phosphodiester bond of the sugar-phosphate back bone.
The recognition site of a restriction enzyme
specific
how many nucleotides is a recognition site composed of
4-6 nucleotides
types of restriction enzymes
sticky end restriction enzyme
blunt end restriction enzyem
sticky end restriction enzymeshow is it produced and whenpairing with anotether DNA fragemnets
1.astaggered cut in mdae in the DNA,leaving overhanging nucleotides
- when pairing with another DNA fragments, the exposed nucleotides must be complementary.
Blunt end restriction enzyme
- Restriction enzymes cut in the middle of the recongition site and thereofre, this straight cut leaves no overhanging nucloeitdes.
if there are equal bases on each end of the dna sequence, then
its blunt sequence if not tis a sticky end(zig zag patterns)
ligase
enzymes that join fragments of DNA, forming a phosphodiester bond after the restricton enzymes cut DNA
annealing
process where ligase, joins fragments of DNA, forming a phosphodiester bond after the restriction enzymes cut the DNA, and leaving sticky ends complemenatary to their overhanign nucldeotiedes
sticky ends must be
not even amount of bases of each side of the sqeuence and complemenatary to their overhanging nucloetides.
polyermase
adds nucleotides to DNA or RNA, leading to genes being copied,
2 types of polyermease
DNA and RNA polymereaese
compare RNA polyermase and DNA polyemrease
RNA polyermease copies the DNA strand to produce an mRNA strand
DNA polyermease adds nucleotides to form DNA in cell replication
what is required for polyreemase to copygenes
a primer
