MicroRNA Research Resources

Frequently Asked Questions (FAQs)

MicroRNA profiling

Q:	How should I prepare or purify my RNA prior to QuantiMir cDNA synthesis?
A:	We have tested both small RNA fractionation using Ambion's miRVana kit and compared it to using Trizol-extracted total RNA preparations with comparable results. For most applications, simple Trizol extraction will suffice. We do not recommend using spin column purification of small RNAs for this procedure.
Q:	What is the sensitivity of the QuantiMir™ system?
A:	For detecting most microRNAs, we recommend that you use 100 picograms of starting total RNA for cDNA synthesis per microRNA you wish to quantitate. Example: Start with 10 ng total RNA=Measure 100 microRNAs
Q:	How can I determine whether I am measuring the mature versus the precursor form of the microRNA?
A:	SBI and others have found that the mature form of the microRNA exists in about 90% of the steady-state RNA preparations versus 8% precursor form. QuantiMir cDNA can be used to measure both forms and discrimination with relative abundance quantitation is simple to perform. View our application note. (Dissociation analysis (PDF) »)
Q:	Is it true that I can quantitate BOTH messenger RNA and small RNAs from QuantiMir cDNA?
A:	Yes. The system converts BOTH small RNAs as well as messenger RNAs that have polyA tails into cDNA compatible with qPCR. View our application note. How to measure shRNA and mRNA knockdown from QuantiMir cDNA (PDF) »

MicroRNA precursor clone collection (Lenti-miRs)

Q:	Are Human mature microRNA sequences homologous to my species model system (Mouse, Rat, etc.)?
A:	Mature microRNA sequences have a very high degree of sequence conservation across all vertebrates. To verify that SBI's Lenti-miR clones will produce the mature microRNA in your model system, view our application note. How to use the Sanger miRBase to align microRNA sequences(PDF) »
Q:	How are mature microRNA sequences expressed from SBI's Lenti-miR clones?
A:	SBI's microRNA precursor clones express precursors that have about 200 bp of native, flanking sequence to the hairpin RNA. This ensures the proper expression and processing of the transcript and the eventual robust expression of the mature microRNAs.
Q:	How can I make a stable cell line overexpressing a microRNA?
A:	SBI's microRNA precursor clones have a built-in GFP fluorescent marker that is co-expressed along with the microRNA. Transduce (infect) your target cells with packaged lentivirus for the microRNA construct of choice. After 48 to 72 hours, sort for GFP-positive cells using flow cytometry. These cells will serve as your founding colony clones.
Q:	How is the pooled Lenti-miR virus library quality-controlled?
A:	The pooled Lenti-miR virus library is transduced into HT1080 cells at an MOI of about 5. After 48 hours, the cells are lysed and genomic DNA is recovered. This genomic DNA is then interrogated for 95 different microRNA sequences along with vector-specific primers to determine that the proper representaion of greater than 90% is achieved.

Global MicroRNA amplification and cloning

Q:	Can I use total RNA or should I fractionate for small RNA prior to using the cloning kit?
A:	SBI's has observed better success in cloning of microRNA sequences when starting with small RNA (200 nt and below) size-fractionated starting RNA.

 

microRNA Expression Vectors
Name	Catalog #	Vector sequence
pMIF-cGFP-Zeo	MIFCZ300PA-1	MIFCZ300PA-1.txt
pCDH-CMV-MCS-EF1-copGFP	CD511B-1	CD511B-1.txt
pGreenPuro (miRZip empty vector)	SI505A-1	SI505A-1.pdf
miRZip scramble hairpin control vector	MZIP000-PA-1	MZ000Scramble_seq.pdf
pPS-EF1-copGFP-LCS	LF522A-1	LF522A-1.txt
pPS-EF1-RFP-LCS	LF522B-1	LF522B-1.txt

Posters and Presentations

Study and Analysis of MicroRNAs, ASCB 2008 Presentation (PDF) »

Identification and Stratification of Clear Cell Renal Carcinoma
using microRNA:Preliminary linkage of microRNA and mRNA in Human Kidney Cancer (PDF) »

Technical information for QuantiMir reference controls

SBI's reference control datasheets for Human RNU6 (U6 snRNA), Mouse RNU6 (U6-snRNA), RNU43 (snoRNA-43) and RNUA1 (U1 snRNA). (PDF) »

Product Citations

QuantiMir™ Small RNA Quantitaton System and qPCR Arrays

Shan SW, Lee DY, Deng Z, Shatseva T, Jeyapalan Z, Du WW, Zhang Y, Xuan JW, Yee SP, Siragam V, Yang BB. MicroRNA MiR-17 retards tissue growth and represses fibronectin expression. Nature Cell Biol. 2009 Aug;11(8):1031-8.

Li X, Shen Y, Ichikawa H, Antes T, Goldberg GS. Regulation of miRNA expression by Src and contact normalization: effects on nonanchored cell growth and migration. Oncogene. (2009 Sep 21): 1-12.

Andrea Krühn, Aijin wang, Johannes H. fruehauf and Hermann Lage. Delivery of short hairpin RNAs by transkingdom RNA interference modulates the classical ABCB1-mediated multidrug-resistant phenotype of cancer cells. Cell Cycle 8:20, 1-6; October 15, 2009.

Ng EK, Tsang WP, Ng SS, Jin HC, Yu J, Li JJ, Röcken C, Ebert MP, Kwok TT, Sung JJ MicroRNA-143 targets DNA methyltransferases 3A in colorectal cancer. Br J Cancer. 2009 Jul 28. [Epub ahead of print]

Shi L, Ko ML, Ko GY. Rhythmic expression of MicroRNA-26a (mir-26a) regulates the L-type voltage-gated calcium channel alpha-1C subunit (VGCC alpha-1C) in chicken cone photoreceptors. J Biol Chem. 2009 Jul 16. [Epub ahead of print]

Yih-Lin Chung, Ming-Yuan Lee and Newman N.M.Pui. Epigenetic therapy using the histone deacetylase inhibitor for increasing therapeuticgain in oral cancer: prevention of radiation-induced oral mucositis and inhibition ofchemical-induced oral carcinogenesis. Carcinogenesis vol.30 no.8 pp.1387–1397, 2009.

To KK, Robey RW, Knutsen T, Zhan Z, Ried T, Bates SE. Escape from hsa-miR-519c enables drug-resistant cells to maintain high expression of ABCG2. Mol Cancer Ther. 2009 Oct;8(10):2959-68.

Wu F, Zhu S, Ding Y, Beck WT, Mo YY. MicroRNA-mediated Regulation of Ubc9 Expression in Cancer Cells. Clin Cancer Res. 2009 Mar 1;15(5):1550-1557.

Chambers C, Shuai B. Profiling microRNA expression in Arabidopsis pollen using microRNA array and real-time PCR. BMC Plant Biol. 2009 Jul 10;9:87.

Tsang WP, Kwok TT. The miR-18a* microRNA functions as a potential tumor suppressor by targeting on K-Ras. Carcinogenesis. 2009 Jun;30(6):953-9.

Wang X, Lam EK, Zhang J, Jin H, Sung JJ. MicroRNA-122a functions as a novel tumor suppressor downstream of adenomatous polyposis coli in gastrointestinal cancers. Biochem Biophys Res Commun. 2009 Sep 18;387(2):376-80.

S. Ruiz, B.D. Schyth, P. Encinas, C. Tafalla, A. Estepa, N. Lorenzen, J.M. Coll. New tools to study RNA interference to fish viruses: Fish cell lines permanently expressing siRNAs targeting the viral polymerase of viral hemorrhagic septicemia virus. Antiviral Research 82 (2009) 148–156.

Qiong Jiang, Mingguang Feng and Yin-Yuan Mo. Systematic validation of predicted microRNAs for cyclin D1. BMC Cancer 2009, 9:194doi:10.1186/1471-2407-9-194 (2009).

R. Grant-Downton, S. Hafidh, D. Twell, and H. G. Dickinson Small RNA Pathways Are Present and Functional in the Angiosperm Male Gametophyte. Mol Plant, March 12, 2009; (2009) ssp003v1.

Lo HL, Chang T, Yam P, Marcovecchio PM, Li S, Zaia JA, Yee JK. Inhibition of HIV-1 replication with designed miRNAs expressed from RNA polymerase II promoters.Gene Ther. 2007 Sep 6

Zhang Y, Li M, Wang H, Fisher WE, Lin PH, Yao Q, Chen C. Profiling of 95 MicroRNAs in Pancreatic Cancer Cell Lines and Surgical Specimens by Real-Time PCR Analysis. World J Surg. 2008 Nov 22.(PDF) »

Stein U, Walther W, Stege A, Kaszubiak A, Fichtner I, Lage H. Complete In Vivo Reversal of the Multidrug Resistance Phenotype by Jet-injection of Anti-MDR1 Short Hairpin RNA-encoding Plasmid DNA. Mol Ther. 2007 Sep 18;1-9.

Siebert, P. & Antes, T. (2007). Enrichment Amplification of Human RNase III-cleaved RNA Reveals New MicroRNAs and Other Small Non-Coding RNAs. PHILICA.COM Article number 100.

Lee NS, Kim JS, Cho WJ, Lee MR, Steiner R, Gompers A, Ling D, Zhang J, Strom P, Behlke M, Moon SH, Salvaterra PM, Jove R, Kim KS. miR-302b maintains "stemness" of human embryonal carcinoma cells by post-transcriptional regulation of Cyclin D2 expression.Biochem Biophys Res Commun. 2008 Dec 12;377(2):434-40.

Paul E. Neiman, Katrina Elsaesser, Gilbert Loring and Robert Kimmel. Myc Oncogene-Induced Genomic Instability: DNA Palindromes in Bursal Lymphomagenesis. PLoS Genet. 2008 July; 4(7): e1000132.

Click for AACR 2008 QuantiMir Technology Highlights

MicroRNA Discovery and Cloning Kits

Tesfaye D, Worku D, Rings F, Phatsara C, Tholen E, Schellander K, Hoelker M. (2009). Identification and expression profiling of microRNAs during bovine oocyte maturation using heterologous approach. Mol Reprod Dev. 2009 Jul;76(7):665-77. (PDF) »

Weiping Peng, Olga Vitvitskaia, Dale Carpenter, Steven L. Wechsler, Clinton Jones. 2008. Identification of two small RNAs within the first 1.5-kb of the herpes simplex virus type 1-encoded latency-associated transcript. Journal of Neurovirology, Volume 14, Issue 1, pages:41–52.

Siebert, P. & Antes, T. (2007). Enrichment Amplification of Human RNase III-cleaved RNA Reveals New MicroRNAs and Other Small Non-Coding RNAs. PHILICA.COM Article number 100. (PDF) »

MicroRNA Precursor Expression Clones

Cordes KR, Sheehy NT, White MP, Berry EC, Morton SU, Muth AN, Lee TH, Miano JM, Ivey KN, Srivastava D. miR-145 and miR-143 regulate smooth muscle cell fate and plasticity. Nature. 2009 Aug 6;460(7256):705-10.

Li X, Shen Y, Ichikawa H, Antes T, Goldberg GS. Regulation of miRNA expression by Src and contact normalization: effects on nonanchored cell growth and migration. Oncogene. (2009 Sep 21): 1-12.

Dongsheng Yan, Xiang Da (Eric) Dong, Xiaoyan Chen, Lihua Wang, Chunjing Lu, Jiao Wang, Jia Qu and LiLi Tu. MicroRNA-1/206 Targets c-Met and Inhibits Rhabdomyosarcoma Development. THE JOURNAL OF BIOLOGICAL CHEMISTRY VOL. 284, NO. 43, pp. 29596–29604, October 23, 2009.

Lu TX, Munitz A, Rothenberg ME. MicroRNA-21 is up-regulated in allergic airway inflammation and regulates IL-12p35 expression. J Immunol. 2009 Apr 15;182(8):4994-5002. (PDF) »

Sachdeva M, Zhu S, Wu F, Wu H, Walia V, Kumar S, Elble R, Watabe K, Mo YY. p53 represses c-Myc through induction of the tumor suppressor miR-145. Proc Natl Acad Sci U S A. 2009 Mar 3; 106(9):3207-12. (PDF) »

Godlewski J, Nowicki MO, Bronisz A, Williams S, Otsuki A, Nuovo G, Raychaudhury A, Newton HB, Chiocca EA, Lawler S. Targeting of the Bmi-1 oncogene/stem cell renewal factor by microRNA-128 inhibits glioma proliferation and self-renewal. Cancer Res. 2008 Nov 15;68(22):9125-30. (PDF) »

Qing Ji, Xinbao Hao, Yang Meng, Min Zhang, Jeffrey DeSano,Daiming Fan and Liang Xu. Restoration of tumor suppressor miR-34 inhibits human p53-mutant gastric cancer tumorspheres. BMC Cancer 2008, 8:266 doi:10.1186/1471-2407-8-266. (PDF) »

Corney DC, Flesken-Nikitin A, Godwin AK, Wang W, Nikitin AY. MicroRNA-34b and MicroRNA-34c are targets of p53 and cooperate in control of cell proliferation and adhesion-independent growth. Cancer Res. 2007 Sep 15;67(18):8433-8. (PDF) »

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