miRNA qPCR assay FAQs

qSTAR miRNA First-strand cDNA synthesis kit is for synthesizing miRNA cDNA using Ecoli poly A polymerase and MMLV reverse transcriptase.

qSTAR qPCR Primer Pairs are pre-designed, qPCR tested and ready-to-use primer sets for SYBR Green based qPCR experiments. The primer pairs are also available in a 96-well panel or 384-well panel.

No. The primer pairs are only compatible with the miRNA cDNA synthesis kit provided by OriGene because of the unique linker sequence in the oligo dT.

No. This kit is only compatible with SYBR Green based qPCR and has not been designed to accommodate any commercial probe-based qPCR.

The Tm of a qSTAR qPCR miRNA Primer is around 55°C, and the amplicon is around 80 bp.

Yes, we can provide our primer panels in other plates within approximately one week of receiving the custom order. Please email the catalog numbers of the qPCR primer panels needed plus the catalog number and manufacturer of the plates to techsupport@origene.com. Our technical support scientists will prepare a custom quote that can be used to order custom panels using a special TSC1001 code.

Yes, OriGene can design a custom qPCR primer panel. Custom panels are offered for human and mouse miRNA sequences in 10 or 200 reaction-format. Please fill out the custom request form on our website at http://www.origene.com to generate custom-mixed panel of miRNAs.

Each panel is delivered with a USB drive that contains a spreadsheet of primer sequences and locations.

Each well in the 200-reaction format contains 1 nmol of each lyophilized primer (2 nmol total). Please re-dissolve in 200ul of water so the final concentration is 10uM total. Each well in the PCR plate contains 10pmol of the primer pairs. The primers will be re-suspended once the PCR reagents are added to the plate.

A sequence specific qPCR template standard is a tube of linear DNA made from a miRNA cDNA cloned in a plasmid. The copy number of a template standard solution is determined by the SYBR Green method and calculation based on MW.

No. qSTAR miRNA template standards are uniquely designed to function only with OriGene's qPCR detection system.

Yes, as long as it is diluted in the qPCR detection range and each dilution is mixed thoroughly.

Yes, as long as the buffer has no negative effects on qPCR. Use appropriate controls.

Since the miRNA cDNA templates in your samples are single-stranded, the actual copy number of your gene of interest is two times the number you would infer in comparison to the qPCR standards. For example, if the copy number of your gene is 5 copy/ul from the standard curve of a qPCR program, the actual number is 10 copy/ul. The reason is that the first cycle for a single-stranded sample is to make the complementary strand; therefore, there is one cycle delay in the PCR reaction compared to the double-stranded cDNA template standard's reaction.

OriGene's miRNA primers are guaranteed to work in SYBR Green qPCR experiments. In order to determine the functionality of miRNA primers, the U6 control primers from the cDNA synthesis kit must be used along side each amplification experiment. If no U6 transcript is detected by qPCR from your cDNA sample, our scientists will work with you to pinpoint the problem and, if it is determined that our primers are at fault, OriGene will replace them free of charge or credit your account.

Here is our guideline:

Full Product Name, OriGene Technologies Inc., Rockville, MD, USA, Catalog #, Lot #.

In addition, we would love to hear from you when the paper is published. Email us a copy of the accepted manuscript and receive a special gift.