NUCLEOTIDE PROBES

Question 1

nucleotide probes are..

Answer 1

DNA or RNA fragment

Question 2

they are well known and used for identify… by…

Answer 2

-the complementary fragment
-hybridization

Question 3

DNA probes

Answer 3

oligoprobes single stranded or double stranded

Question 4

RNA probes

Answer 4

riboprobes

Question 5

hot labels

Answer 5

radioative nucleotides (32P,35S,3h) on alpha or gamma

Question 6

cold labels

Answer 6

ligand couple to a base (biotin,flurochrome)

Question 7

labelling techniques for DAN probes

Answer 7

• AT 5’ end
• at 3’ ends
• uniform

Question 8

5’ end technique : radioative phosphate on a nucleotide thanks to..

Answer 8

T4 kinase

Question 9

at 3’ end, incorporation by.. of a..

Answer 9

-T.Transferase
- labelled nucleotide (cold)

Question 10

Labelling techniques for RNA probes

Answer 10

uniform

Question 11

uniform technique

Answer 11

polymerase is labelled

Question 12

prehybridization

Answer 12

-saturation of non specific binding site of the probes by adding non salmon sperm DNA
-at 15 to 20 degree

Question 13

hybridization

Answer 13

addition of denatured probe

Question 14

washes

Answer 14

removing non specific binding by increasing T and decreasing Na concentration

Question 15

detection of hybrids if non radioactive :

Answer 15

autoradiography or Storm apparatus

Question 16

direct probes detection for radioactive probes

Answer 16

autoradiography

Question 17

direct probes detection for non radioactive probes

Answer 17

fluorimetry

Question 18

indirect detection for probes labelled wit dixiogenin

Answer 18

• fluorecent antibody + enzyme

Question 19

indirect detection for probes labelled with biotin

Answer 19

• straptavin

Question 20

what blot technique do we use for DNA

Answer 20

southern blot or dot blot

Question 21

for RNA

Answer 21

northern blot

Question 22

proteins

Answer 22

western blot