How do the miRNA assays correspond to miRBase?

Product FAQ

Answer

Individual TaqMan™ miRNA Assays are designed to target specific mature miRNA sequences. The mature sequences correspond to those contained in miRBase. Often several precursor miRNAs with different names and sometimes across several species will produce the identical mature miRNA sequence. Thus, one particular assay may be valid for use across several species. The Assay Index File provides the cross species key for TaqMan™ MicroRNA Assays.

Answer Id: E2514

Was this answer helpful?

Yes
No
Thank you for your response

Why are miRNAs so important?

Product FAQ

Answer

MicroRNAs are believed to play an important role in post transcriptional gene regulation. A total of ~850 unique miRNAs have been discovered so far, including ~333 human miRNAs. They are relatively abundant, ranging from a few to as many as 50,000 molecules per cell. They account for approximately 1% of the predicted genes in animals and plants. The level of individual miRNAs varies dramatically across cell type and developmental stages. These differences in miRNA level are believed to be a key indicator of miRNA activity.

Recent research has implicated miRNAs in the following: cell development, differentiation, communication and cell death; DNA methylation and chromatin modification; metabolism; human cancer development; haematopoiesis; nervous system patterning; insulin secretion.

Answer Id: E2512

Was this answer helpful?

Yes
No
Thank you for your response

What does it mean when a miRNA target entry has been removed (obsoleted) from miRBase?

Product FAQ

Answer

miRBase continues to re-annotate the miRNA sequences as more information becomes available. A sequence that once mapped to a given species (e.g.: human) may no longer exist, so it is removed from miRBase. Accordingly, the original assay for that sequence no longer maps to an annotated miRNA, and as a result the annotation fields on the search results page on the Thermo Fisher Scientific portal are empty. An example of this is Assay “hsa-miR-505” (Assay ID 001049). miRBase continues to re-annotate the miRNA sequences as more information becomes available. A sequence that once mapped to a given species (e.g.: human) may no longer exist, so it is removed from miRBase. Accordingly, the original assay for that sequence no longer maps to an annotated miRNA, and as a result the annotation fields on the search results page on the Thermo Fisher Scientific portal are empty. An example of this is Assay “hsa-miR-505” (Assay ID 001049).

Answer Id: E2522

Was this answer helpful?

Yes
No
Thank you for your response

How long can I store the cDNA from my reverse transcription step?

Product FAQ

Answer

You can store your cDNA at 2-6 degrees C for up to 24 hours. For long-term storage, store the cDNA at -15 to -25 degrees C and add EDTA to a final concentration of 1 mM to prevent degradation.

Answer Id: E7479

Was this answer helpful?

Yes
No
Thank you for your response

What is mechanism of the TaqMan™ MicroRNA Assays?

Product FAQ

Answer

There are 2 steps involved in using the TaqMan™ MicroRNA Assays:

1. RT step: total RNA to cDNA using miRNA-specific RT primers from the TaqMan™ MicroRNA Assays and the TaqMan™ MicroRNA Reverse Transcription Kit (P/N 4366596, 200 reactions or 4366597, 1000 reactions)

2. PCR step: cDNA to PCR products using primers and probe from the TaqMan MicroRNA Assays together with TaqMan™ Universal PCR Master Mix No AmpErase™ UNG (P/N 4324018).

Answer Id: E2516

Was this answer helpful?

Yes
No
Thank you for your response

What are miRNAs?

Product FAQ

Answer

MicroRNAs (miRNAs) are endogenous non-coding RNAs, about 22 nucleotides in length, that play an important role in post-transcriptional gene regulation in animals and plants by targeting messenger RNA (mRNA) for degradation or translational repression. The miRNA precursors are transcribed from individual genes but are not translated into proteins. The primary transcript is processed in the nucleus to give one or more hairpin precursors that is exported to the cytoplasm. The mature miRNA is excised from the hairpin precursor. The mature miRNA is the biologically active form and regulates the expression of mRNA transcripts by binding to complementary sites on target mRNAs and inhibiting translation or, when there is perfect complementarity to the target sequence, inducing mRNA cleavage. In animals, translational repression, not transcript degradation, is the dominant miRNA mode of action.

Answer Id: E2510

Was this answer helpful?

Yes
No
Thank you for your response

What is the TaqMan™ MicroRNA Assay?

Product FAQ

Answer

A TaqMan™ MicroRNA Assay has two components: an RT reaction containing a miRNA specific stem-loop reverse-transcription primer and a separate specific TaqMan™ miRNA Assay. The RT stem-loop primer provides the specificity for the mature miRNA target; it does not detect its precursor. The formation of a RT primer/mature miRNA-chimera extends the length of the 5’ end of the miRNA. The longer RT product provides a miRNA specific cDNA template amenable to the TaqMan™ assay design.

Answer Id: E2519

Was this answer helpful?

Yes
No
Thank you for your response

Can I use any reverse transcription kit for a TaqMan™ MicroRNA RT reaction?

Product FAQ

Answer

No, only the TaqMan™ MicroRNA Reverse Transcription Kit (Cat. No. 4366596 (200 reactions)) or Cat. No. 4366597 (1,000 reactions)) should be used. This kit was developed specifically for use with TaqMan™ MicroRNA Assay reverse transcription.

Answer Id: E2768

Was this answer helpful?

Yes
No
Thank you for your response