Overview

Get your miRNA ready for qPCR profiling

When you’d like to profile miRNA with qPCR, SBI’s QuantiMir™ Kit can get your samples ready. By efficiently converting all of your miRNA into cDNA, you can get accurate, sensitive, and quantitative qPCR data on the miRs in your sample. Great for understanding differential miRNA expression, simultaneously profiling siRNA and mRNA, and more.

• Simple and robust procedure
• Rapidly and efficiently convert all small RNAs into cDNAs for qPCR
• Suitable for high-throughput screening of clinical samples
• Sensitive and accurate (see the data in the Supporting Data section below)
• Versatile—design your own miRNA assays

Each kit includes three reference assays—human U6, mouse U6 and human/mouse miR-16. We also offer the following reference assays as individual 50-assay kits:

How It Works

The QuantiMir Kit enables robust miRNA quantitation through a simple and efficient workflow

Highly efficient poly-A tailing and reverse transcription in a single reaction tube provides uniform cDNA synthesis of miRNAs. The optimized reaction conditions and buffer components maximize cDNA yield when starting with several micrograms down to picograms of input total RNA. The universal 3′ tag sequence incorporated during reverse transcription enables easily scalable and accurate miRNA expression analysis by qPCR—profile thousands of different miRNAs from a single reverse transcription reaction.

• Tag all small RNAs with a poly-A tail
• Anneal an oligo-dT adaptor
• Reverse transcribe to create first-strand cDNA

The result is a cDNA pool of anchor-tailed miRNAs that are ready for qPCR.

Create custom miRNA assays

To design your own miRNA assays, simply synthesize an oligo using the sequence of the mature miRNA you’d like to profile as the forward primer in your miRNA qPCR assay, and use with the universal reverse primer included in the QuantiMir Kit.

Supporting Data

Converting miRNA into cDNA for accurate qPCR profiling and quantitation

The QuantiMir Kit is sensitive

Figure 1. The QuantiMir Kit is highly sensitive and enables measurement across a wide dynamic range. (A) Starting with total, Trizol-extracted RNA or fractionated small RNA samples you can measure from several micrograms down to picograms of input RNA with excellent accuracy. (B) You can also detect differential miRNA expression across a dynamic range of at least 6 logs.

The QuantiMir Kit is accurate

Figure 2. The QuantiMir Kit is accurate. We used the QuantiMir Kit to synthesize first strand cDNAs from 18 different human tissues: adipose, bladder, brain, cervix, colon, esophagus, heart, kidney, liver, lung, ovary, placenta, prostate, skeletal muscle, small intestine, spleen, testes, and thymus. The cDNAs were balanced to yield equal C_t values for the U6 snRNA normalizing transcript (bottom left plot, green bars). Real-time PCR results demonstrate that the normalizing snRNA is uniformly expressed across the 18 tissues examined. As expected, assays for miR-1 are show specific expression in heart and musculoskeletal tissues (top left plot, red bars) whereas assays for miR-122 show specific expression in the liver (top right plot, blue bar).

Use the QuantiMir Kit to profile miRNAs from cancerous and normal tissues

Figure 3. Use the QuantiMir Kit to profile miRNAs from cancerous and normal tissues. Example of quantitative miRNA profiling of nine miRNAs in five different normal and tumor-derived samples.

Measure both siRNA and mRNA knockdown in a single QuantiMir cDNA sample

Figure 4. Measure both siRNA and mRNA knockdown in a single QuantiMir cDNA sample. A time-course using anti-p53 shRNA-directed knockdown of the endogenous p53 mRNA transcript demonstrates how the QuantiMir Kit can be used to measure both p53 siRNA (orange bars) and p53 mRNA (blue line) via qPCR.

Resources

Citations

• Bhome, R, et al. (2017) Profiling the MicroRNA Payload of Exosomes Derived from Ex Vivo Primary Colorectal Fibroblasts. Methods Mol. Biol..2017 Nov 9; 1509:115-122. PM ID:27826922
• Dluzen, DF, et al. (2017) MicroRNAs Modulate Oxidative Stress in Hypertension through PARP-1 Regulation. Oxid Med Cell Longev.2017 Jun 29; 2017:3984280. PM ID:28660007
• Su, YK, et al. (2017) Antrodia cinnamomea sensitizes radio-/chemo-therapy of cancer stem-like cells by modulating microRNA expression. J Ethnopharmacol.2017 Jun 8;. PM ID:28602756
• Min, KW, et al. (2017) AUF1 facilitates microRNA-mediated gene silencing. Nucleic Acids Res..2017 Jun 2; 45(10):6064-6073. PM ID:28334781
• Zheng, H, et al. (2017) Alcohol-dysregulated microRNAs in hepatitis B virus-related hepatocellular carcinoma. PLoS ONE.2017 May 31; 12(5):e0178547. PM ID:28562643
• Paul, D, et al. (2017) A-to-I editing in human miRNAs is enriched in seed sequence, influenced by sequence contexts and significantly hypoedited in glioblastoma multiforme. Sci Rep.2017 May 26; 7(1):2466. PM ID:28550310
• Reeves, ME, et al. (2017) Identification and characterization of RASSF1C piRNA target genes in lung cancer cells. Oncotarget.2017 May 23; 8(21):34268-34282. PM ID:28423657
• Brodie, S, et al. (2017) The novel long non-coding RNA TALNEC2, regulates tumor cell growth and the stemness and radiation response of glioma stem cells. Oncotarget.2017 May 9; 8(19):31785-31801. PM ID:28423669
• Gao, S, et al. (2017) Identification and characterization of miRNAs in two closely related C4 and C3 species of Cleome by high-throughput sequencing. Sci Rep.2017 Apr 19; 7:46552. PM ID:28422166
• Yang, Y, Bai, YS & Wang, Q. (2017) CDGSH Iron Sulfur Domain 2 Activates Proliferation and EMT of Pancreatic Cancer Cells via Wnt/β-Catenin Pathway and Has Prognostic Value in Human Pancreatic Cancer. Oncol. Res..2017 Apr 14; 25(4):605-615. PM ID:27983920
• Tsai, HC, et al. (2017) WISP-1 positively regulates angiogenesis by controlling VEGF-A expression in human osteosarcoma. Cell Death Dis.2017 Apr 13; 8(4):e2750. PM ID:28406476
• Ramachandran, M, et al. (2017) Safe and Effective Treatment of Experimental Neuroblastoma and Glioblastoma Using Systemically Delivered Triple MicroRNA-Detargeted Oncolytic Semliki Forest Virus. Clin. Cancer Res..2017 Mar 15; 23(6):1519-1530. PM ID:27637889
• Fafián-Labora, J, et al. (2017) Effect of age on pro-inflammatory miRNAs contained in mesenchymal stem cell-derived extracellular vesicles. Sci Rep.2017 Mar 6; 7:43923. PM ID:28262816
• Maeda, Y, et al. (2017) Synovium-Derived MicroRNAs Regulate Bone Pathways in Rheumatoid Arthritis. J. Bone Miner. Res..2017 Mar 1; 32(3):461-472. PM ID:27676131
• Chen, S, et al. (2017) H19 Overexpression Induces Resistance to 1,25(OH)2D3 by Targeting VDR Through miR-675-5p in Colon Cancer Cells. Neoplasia.2017 Mar 1; 19(3):226-236. PM ID:28189050
• Hou, S, et al. (2017) MicroRNA-939 governs vascular integrity and angiogenesis through targeting γ-catenin in endothelial cells. Biochem. Biophys. Res. Commun..2017 Feb 26; 484(1):27-33. PM ID:28115160
• Yeung, CL, et al. (2017) Human papillomavirus type 16 E6 suppresses microRNA-23b expression in human cervical cancer cells through DNA methylation of the host gene C9orf3. Oncotarget.2017 Feb 14; 8(7):12158-12173. PM ID:28077801
• Tsukita, S, et al. (2017) MicroRNAs 106b and 222 Improve Hyperglycemia in a Mouse Model of Insulin-Deficient Diabetes via Pancreatic β-Cell Proliferation. EBioMedicine.2017 Feb 1; 15:163-172. PM ID:27974246
• Monsanto-Hearne, V, et al. (2017) Drosophila miR-956 suppression modulates Ectoderm-expressed 4 and inhibits viral replication. Virology.2017 Feb 1; 502:20-27. PM ID:27960110
• Small, RNA. (2017) QuantiMir RT Kit Small RNA Quantitation System. Product.;. Link:Product