southern blotting

synthesizes DNA

northern blotting

synthesizes RNA

western blotting

synthesizes protiens

1st step PCR

Primers target a specific region

2nd step PCR

Denature DNA (~95°C)

3rd step PCR

Anneal primers (~55°-65°C)

1st step cDNA synthesis

RNA purification

2nd step cDNA synthesis

RNA incubated with reverse transcriptase

3rd step of cDNA synthesis

oligo-dT primer anneals polyA tail of mRNA so reverse transcriptase synthesizes single strands DNA

4th step of cDNA synthesis

mRNA is removed and DNA polymerse copies the cDNA strand

5th step of cDNA synthesis

nick hairpin

4th step PCR

Elongate new DNA strands, using Taq or other DNA polymerase(~72°C)

5th step PCR

Repeat denaturation, annealing, and elongation for 20-40 cycles.

miRNA and siRNA similarities

21-22 nucleotides

undergo post-transcriptional processing.
involve the formation of double-stranded RNA

participate in RNA interference (RNAi), targeted, RNA-
mediated repression of gene expression

siRNAs

miRNA

Many are produced by RNA pol II

regulate other genes

may have a mismatch with target transcripts

are implicated in developmental processes

may block translation and/or trigger RNA degradation