Abi 7900 ht real time thermal cycler

All buffy-coat cryotubes were stored in freezers at −80°C. Genomic DNA was extracted from peripheral white blood cells using the QG-Mini80 workflow with DB-S kit (Fujifilm Corporation, Tokyo, Japan) as instructed.
Telomere length ratio (T/S ration) was measured using a quantitative PCR-based technique [19 (link)]. In this method, the ratio of the telomere repeat copy number (T) and single-copy gene number (S) was compared for each sample. Reactions for DNA samples were run in 7 μL reaction volumes with ABI-7900 HT real-time thermal cycler (Applied Biosystems).
The primers are as follows. 

Telomere-F 5′-CGGTTTGTTTGGGTTTGGGTTTGGGTTTGGGTTTGGGTT-3′.

 

Telomere-R 5′-GGCTTGCCTTACCCTTACCCTTACCCTTACCCTTACCCT-3′.

 

β-globin-F 5′-GCTTCTGACACAACTGTGTTCACTAGC-3′.

 

β-globin-R 5′-CACCAACTTCATCCACGTTCACC-3′.

 

β-globin-P FTGCATCTGACTCCTGAGGP.

Cycling conditions of telomere were as follows: 95°C incubation for 10 minutes followed by 35 cycles of 95°C for 15 seconds and 56°C for 1 min. As for the cycling conditions of β-globin, 95°C incubation for 10 minutes followed by 40 cycles of 95°C for 15 sec and 56°C for 1 min. The specificity of all of the amplification was determined by melting curve analysis.