Probe labelling and hybridization

Probe labeling is defined as sequence use to search the mixture of nucleic acid for molecule containing complementary sequence.In molecular biology, hybridization is a phenomenon in which single-stranded deoxyribonucleic acid (DNA) or ribonucleic acid (RNA) molecules anneal to complementary DNA or RNA

1. AMBER HASSAN PhD Scholar Aznostics the diagnostic center PROBE LABELLING AND HYBRIDIZATION

2. PROBE: A probe is a piece of DNA of the sequence of the interest PROBE LABELLING: a labeled and defined sequence use to search the mixture of nucleic acid for molecule containing complementary sequence Probe concentration: The higher the probe concentration, the faster the hybridization occurs. However, the higher the probe concentration, the higher the blot background (i.e. probe will stick everywhere ). Low probe concentrations and long incubations (overnight) usually produce the best results.

3. LABELING OF DNA AND RNA PROBES RADIOACTIVE LABELING: DISPLAY OR MAGNIFY THE SIGNALSBY RADIOACTIVITY. NON RADIOACTIVE LABELING: DISPLAY OR MAGNIFY THE SIGNALS BY ANTIGEN LABELING. END LABELING: PUT THE LABELES AT THE END. UNIFORM LABELING :PUT THE LABELS INTERNALLY.

4. Classic procedure : Radioactive labelling of complementary RNA

5. The Probe: Artificial RNA Labelled with DIG Produced in vitro

6. HYBRIDIZATION: In molecular biology, hybridization is a phenomenon in which single-stranded deoxyribonucleic acid ( DNA ) or ribonucleic acid ( RNA ) molecules anneal to complementary DNA or RNA. double-stranded DNA sequence is generally stable under physiological conditions, changing these conditions in the laboratory (generally by raising the surrounding temperature) will cause the molecules to separate into single strands. These strands are complementary to each other but may also be complementary to other sequences present in their surroundings. Lowering the surrounding temperature allows the single-stranded molecules to anneal or “hybridize” to each other. DNA replication and transcription of DNA into RNA both rely upon nucleotide hybridization, as do molecular biology techniques including Southern blots and Northern blots ,the polymerase chain reaction (PCR), and most approaches to DNA sequencing .

7. 1.DNA-DNA HYBRIDIZATION:

8. 2.FLUORSCENT IN SITU HYBRIDIZATION :

9. RNA/DNA hybridization is influenced by… Probe concentration Sequence (GC bonds more stable than AT ) Stringency