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Perfecta sybr green fastmix reaction mixes

Manufactured by Quanta Biosciences
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PerfeCTa SYBR Green FastMix Reaction Mixes is a laboratory product designed for real-time PCR applications. It contains a proprietary buffer, stabilizers, SYBR Green I dye, and a high-performance thermostable DNA polymerase.

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Lab products found in correlation

Quantstudio 6 flex pcr system, by Thermo Fisher Scientific (1 mentions) Qscript xlt cdna supermix, by Quanta Biosciences (1 mentions) Steponeplus real time pcr system, by Thermo Fisher Scientific (1 mentions) Direct zol rna microprep kit, by Zymo Research (1 mentions) Dnase, by Qiagen (1 mentions) Tri reagent, by Zymo Research (1 mentions) Realplex4, by Eppendorf (1 mentions)

3 protocols using perfecta sybr green fastmix reaction mixes

1

Quantitative Chromatin Immunoprecipitation (qChIP)

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Quantitative ChIP (qChIP) was performed using PerfeCTa SYBR Green FastMix Reaction Mixes (Quanta Biosciences, Gaithersburg, MD) on a QuantStudio 6 Flex PCR System (Thermo Fisher Scientific). The Ct (cycle threshold) of immunoprecipitates were normalized to their respective input control. P-values between regimens were calculated using Student’s t-test and were considered significant after passing the test for vehicle-treated samples. Primers used for qChIP are listed in TableS5. Additional antibodies used for qChIP were p300 (A300–358A, Bethyl laboratories, Montgomery, TX) and HDAC2 (A300–705A, Bethyl laboratories), whose immunocomplexes were washed according to the LSD1-ChIP protocol.
Duy C., Teater M., Garrett-Bakelman F.E., Lee T.C., Meydan C., Glass J.L., Li M., Hellmuth J.C., Mohammad H.P., Smitheman K.N., Shih A.H., Abdel-Wahab O., Tallman M.S., Guzman M.L., Muench D., Leighton Grimes H., Roboz G.J., Kruger R.G., Creasy C.L., Paietta E.M., Levine R.L., Carroll M, & Melnick A.M. (2019). Rational targeting of cooperating layers of the epigenome yields enhanced therapeutic efficacy against AML. Cancer discovery, 9(7), 872-889.
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2

Quantitative Real-Time PCR Analysis of Gene Expression

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Total RNA was isolated using TRI reagent (Zymo Research) and further purified with the Direct-zol RNA MicroPrep kit (Zymo Research), treated with DNase (30 U/µg total RNA, QIAGEN), and reverse transcribed using qScript XLT cDNA SuperMix (Quanta Bioscience). Expression of mRNA was quantitated using PerfeCTa SYBR Green FastMix Reaction Mixes (Quanta Bioscience) and the StepOnePlus Real-Time PCR System (Applied Biosystems) with cDNA equivalent to 7.5 ng total RNA.
Primers used for real-time PCR include the following (5′–3′): HPRT-Fw, TGA​CAC​TGG​CAA​AAC​AAT​GCA; HPRT-Rv, GGT​CCT​TTT​CAC​CAG​CAA​GCT; SPNS2-Fw, AAC​GTG​CTC​AAC​TAC​CTG​GAC; SPNS2-Rv, ATG​AAC​ACT​GAC​TGC​AGC​AG; LPAR1-Fw, ACT​GTG​GTC​ATT​GTG​CTT​GG; LPAR1-Rv, ACA​GCA​CAC​GTC​TAG​AAG​TAA​C; FAM156A-Fw, TAT​GCT​GTT​GGG​AGG​GAA​GC; and FAM156A-Rv, GCA​GTA​TCG​ACA​TTC​ACA​TCG​G.
Hisano Y., Kono M., Cartier A., Engelbrecht E., Kano K., Kawakami K., Xiong Y., Piao W., Galvani S., Yanagida K., Kuo A., Ono Y., Ishida S., Aoki J., Proia R.L., Bromberg J.S., Inoue A, & Hla T. (2019). Lysolipid receptor cross-talk regulates lymphatic endothelial junctions in lymph nodes. The Journal of Experimental Medicine, 216(7), 1582-1598.
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3

Quantifying Osteogenic and Angiogenic Genes

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The expression of the osteogenic genes bone morphogenic protein-2 (BMP-2), transcription factor RUNX2, and bone sialoprotein (BSP), and of the angiogenic genes von Willebrand factor (vWF), CD31, and vascular endothelial growth factor A (VEGF) were quantified in the cDNA samples using a real-time PCR reaction. The target genes were normalized with 18S rRNA as the housekeeping. The primers (200 nM) were designed to span exon-exon junctions using the primer-BLAST tool (sequences in Table S1). The real-time PCR reaction was done using the PerfeCTa SYBR Green FastMix Reaction Mixes (Quanta Biosciences), according to manufacturer’s specifications. The reactions were monitored in a Mastercycler (Realplex4, Eppendorf) using the software realplex version 2.2 (Eppendorf). The relative quantification of the osteogenic and angiogenic markers expression was performed using the 2−ΔΔCT method (Perkin-Elmer). Results are expressed relatively to gene expression levels of day 3.
Correia C.R., Pirraco R.P., Cerqueira M.T., Marques A.P., Reis R.L, & Mano J.F. (2016). Semipermeable Capsules Wrapping a Multifunctional and Self-regulated Co-culture Microenvironment for Osteogenic Differentiation. Scientific Reports, 6, 21883.
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