Sybr premix ex taq tli rnase h plus

Total RNA was extracted from cells or tissues with Trizol reagent (Invitrogen). For isolating RNA from blood, we used the kit Mouse RiboPure–Blood RNA Isolation (Applied Biosystems) following manufacturer procedure. Then, 1 μg of RNA was reverse-transcribed using a QuantiTect Reverse Transcription Kit (Qiagen) on a Q-cycler II. Quantitative PCR was done using SYBR Premix Ex Taq (Tli RNase H Plus) (Ozyme) on a StepOne machine (Life Technologies). The relative amount of all mRNAs was calculated using the comparative ΔΔCT method, and either 36B4 (for mice) or RPL27 (for humans) was used as the housekeeping gene. Primer sequences are available upon request.

Total RNA was extracted from the AAA patient serum-treated primary human macrophages with Trizol reagent (Invitrogen, Waltham, MA, USA). Then, 1 μg of RNA was reverse-transcribed using a QuantiTect Reverse Transcription Kit (Qiagen, Hilden, Germany) on a Q-cycler II. A quantitative PCR was performed using SYBR Premix Ex Taq (Tli RNase H Plus) (Ozyme, Paris, France) on a StepOne machine (Life Technologies, Carlsbad, CA, USA). The relative amount of all mRNAs was calculated using the comparative ΔΔCT method, and cyclophilin A was used as the housekeeping gene. Primer sequences are available upon request.

Total RNA was extracted from cells or tissues with Trizol reagent (Invitrogen) and 1 μg of RNA was reverse-transcribed using a QuantiTect Reverse Transcription Kit (Qiagen) on a QcyclerII. Quantitative PCR was done using SYBR Premix Ex Taq (Tli RNase H Plus) (Ozyme) on a StepOne machine (Life Technologies). The mRNA levels of all genes were normalized to 36B4 transcript levels. Sequences of primers used are as follows: Pparβ/δ-F: GCA-GCC-TCA-ACA-TGG-AAT-GTC; Pparβ/δ-R CAT-ACT-CGA-GCT-TCA-TGC-GG. These primers allow the detection of intact exon 4 (wild-type form) and does not detect the recombined transcript, deleted from this exon (mutated form). Cpt1a-F: CTC-AGT-GGG-AGC-GAC-TCT-TCA; Cpt1a-R: GGC-CTC-TGT-GGT-ACA-CGA-CAA; 36B4-F: TCC-AGG-CTT-TGG-GCA-TCA; and 36B4-R: CTT-TAT-CAG-CTG-CAC-ATC-ACT-CAG-A.