Manipulating DNA Flashcards
What does isolating a specific DNA segment allow researchers to do?
Compare DNA samples and transfer DNA from one organism to another.
What enzyme cuts DNA segments?
Restriction endonucleases.
Where and exactly what do restriction endonucleases cut?
dsDNA at specific recognition sites.
What indicates a recognition site?
A palindrome: read the same 3’to5’ as 5’to3’.
How are the pieces of DNA cut?
Unevenly, leaving sticky ends (short pieces of ssDNA)
What will the sticky ends do?
H-bond to similar ssDNA pieces.
How many recognition sites does DNA generally have?
A recognition site for any one restriction enzyme. Every 5000 bp or so.
Give an example of the spacing of recognition sites.
2000, 9000, 1400, 16000, 23000.
How are restriction enzymes named?
After the bacteria they are extracted from. (Genus, species, strain, and isolate).
What does a methylase do?
Adds methyl groups to nitrogenous bases
What happens when a base is methylated?
It will be easily recognized by a restriction enzyme. The DNA will not be bisected.
What do methylases allow researchers to do?
Protect sections of DNA before using restriction enzymes.
What does DNA ligase do?
Attaches ssDNA complementary base pairs.
what happens after DNA ligase attaches ssDNA complementary base pairs?
The sticky ends form phosphdiester bonds to hold the bases together.
Explain the 4 stages in manipulating DNA
- Uncut DNA
- Unprotected DNA is cut with restriction enzymes.
Optional: - Protected DNA strand is cut with restriction enzymes.
- Protected DNA remains and first 2 segments were ligated.
