03 - Lecture 18 Flashcards
What help you see the changes in DNA?
Probes, fluorence
What happens if there is a mutation in the promoter region?
The RNA polymerase will not see the promoter region and will not make RNA to begin with
Gel electrophoresis?
Top is big and bottom is small
What is used to chop up the DNA into fragments?
Restriction endonuclease
Does Northern blot use a restriction endonuclease?
No because RNA is single stranded
What helps isolate mRNA from the cell?
Poly-T tail attached to a bead
What is the charge of the proteins when doing Western blot?
The proteins are negatively charged because of the detergent that coats the proteins
What technique would you use if you do not know what the mutation is?
di-deoxy method
Base pairs are incorporated in to a growing chain until what?
Synthesis stops because of ddNTP (no 3’ OH)
Steps of PCR?
Denature with heat into single strand DNA, the primer anneals to complementary DNA with cool temp, then you elongate the sample with warm temp
Steps of RFLP?
Use a restriction endonuclease to recognize and cut a certain base sequence in the DNA (different genes = different length of fragments)
- after you cut out the fragments you can use Southern blotting to compare sizes
How can southern blotting/RFLP be used for sickle cell?
In a normal beta globin gene there are two fragments (1.15 and 0.2) in the sickle cell fragment there is only one fragment (1.35)
