The reverse transcription of RNA was
performed in two steps according to the instructions of Roche Reverse
Transcription Kit.29 (link) The RNA concentration
and purity were determined by a Nano Drop spectrophotometer (Nano
Drop Technologies, Wilmington, DE), and the determination was performed
by ABI fluorescence quantitative PCR (Rrism 7500, Applied Biosystems,
Foster City, CA). The cDNA was obtained and stored in a refrigerator
at −20 °C for standby. The RT-PCR amplification reaction
was performed according to the instructions of FastStart DNA Master
SYBR Green I kit. Twenty microliters of the reaction system (10 μL
of SYBR Green Master + 6 μL of DNase/RNase-Free Water + 2 μL
of cDNA + 2 μL primer) was used to detect the effect of samples
on the expression of genes related to the ox-LDL-induced foam of macrophages.
Gene-specific primers were as below: GAPDH (forward, 5′-TTTGTCAAGCTCATTTCCTGGTATG-3′,
reverse, 5′-TGGGATAGGGCCTCTCTTGC-3′), ABCA1 (forward,
5′-CCCAGAGCAAAAAGGGACTC-3′, reverse, 5′-GGTCATCATCACTTTGGTCCTTG-3′),
ABCG1 (forward, 5′-CAAGACCCTTTTGAAAGGGATCTC-3′, reverse,
5′-GCCAGAATATTCATGAGTGTGGAC-3′), SRA1 (forward, 5′-TTAAAGGTGATCGGGGACAAA-3′,
reverse, 5′-CAACCAGTCGAACTGTCTTAAG-3′), SRB1 (forward,
5′-GCAAATTTGGCCTGTTTGTT-3′, reverse, 5′-GATCTTGCTGAGTCCGTTCC-3′),
and CD36 (forward, 5′-CAAGCTCCTTGGCATGGTAGA-3′, reverse,
5′-TGGATTTGCAAGCACAATATGAA-3′).
performed in two steps according to the instructions of Roche Reverse
Transcription Kit.29 (link) The RNA concentration
and purity were determined by a Nano Drop spectrophotometer (Nano
Drop Technologies, Wilmington, DE), and the determination was performed
by ABI fluorescence quantitative PCR (Rrism 7500, Applied Biosystems,
Foster City, CA). The cDNA was obtained and stored in a refrigerator
at −20 °C for standby. The RT-PCR amplification reaction
was performed according to the instructions of FastStart DNA Master
SYBR Green I kit. Twenty microliters of the reaction system (10 μL
of SYBR Green Master + 6 μL of DNase/RNase-Free Water + 2 μL
of cDNA + 2 μL primer) was used to detect the effect of samples
on the expression of genes related to the ox-LDL-induced foam of macrophages.
Gene-specific primers were as below: GAPDH (forward, 5′-TTTGTCAAGCTCATTTCCTGGTATG-3′,
reverse, 5′-TGGGATAGGGCCTCTCTTGC-3′), ABCA1 (forward,
5′-CCCAGAGCAAAAAGGGACTC-3′, reverse, 5′-GGTCATCATCACTTTGGTCCTTG-3′),
ABCG1 (forward, 5′-CAAGACCCTTTTGAAAGGGATCTC-3′, reverse,
5′-GCCAGAATATTCATGAGTGTGGAC-3′), SRA1 (forward, 5′-TTAAAGGTGATCGGGGACAAA-3′,
reverse, 5′-CAACCAGTCGAACTGTCTTAAG-3′), SRB1 (forward,
5′-GCAAATTTGGCCTGTTTGTT-3′, reverse, 5′-GATCTTGCTGAGTCCGTTCC-3′),
and CD36 (forward, 5′-CAAGCTCCTTGGCATGGTAGA-3′, reverse,
5′-TGGATTTGCAAGCACAATATGAA-3′).