Background
Methods
Tissue collection, follicular fluid aspiration and granulosa cell culture
Cell lysis, RNA preparation and cDNA synthesis
Quantification of transcripts and of the respective external standards by real-time PCR
Symbol | Name | Sequence |
---|---|---|
B2M
|
Beta-2-microglobulin
|
F:ACGCTGAGTTCACTCCCAACAGCAA
|
R:TCGATGGTGCTGCTTACAGGTCTCG
| ||
GAPDH
|
Glyceraldehyde-3-phosphate dehydrogenase
|
F:AGCGAGATCCTGCCAACATCAAG
|
R:GCAGGAGGCATTGCTGACAATCT
| ||
HMBS
|
Hydroxymethylbilane synthase
|
F:CAGCATGAAGATGGCCCTGAAGATG
|
R:CTCAGGTAGCAGAGGGCTGGGATGT
| ||
HPRT1
|
Hypoxanthine phosphoribosyltransferase 1
|
F:TGAAAAGGACCCCTCGAAGTGTTGG
|
R:CGCCAGGTATTTCCAAACTCAACTCG
| ||
RPLP0
|
Ribosomal protein, large, P0
|
F:TGGTTACCCAACCGTCGCATCTGTA
|
R:CACAAAGGCAGATGGATCAGCCAAG
| ||
RPS18
|
Ribosomal protein S18
|
F:GAGGTGGAACGTGTGATCACCATT
|
R: TGTATTTCCCGTCCTTCACGTCCT
| ||
TBP
|
TATA box binding protein
|
F: GCCTTGTGCTTACCCACCAACAGTTC
|
R: TGTCTTCCTGAAACCCTTCAGAATAGGG
|
Step | Description | Temperature | Time |
---|---|---|---|
1
|
Pre-incubation
|
95°C
|
5 min
|
2
|
Denaturation
|
95°C
|
20 sec
|
3
|
Annealing
|
60°C
|
15 sec
|
4
|
Extension
|
72°C
|
15 sec
|
5
|
Melting
|
95°C
|
5 sec
|
70°C
|
60 sec
| ||
97°C
|
1 s (continuous acquisition)
|