Oligo Quencher Labeling
Quenchers are a class of chromophores that, instead of emitting absorbed fluorescence resonance energy as light, have the useful property of transforming the light energy to heat. Such kind of probe design has simplify many fluorescence assays, Through FRET (Fluorescence Resonance Energy Transfer), it emits the energy at different wavelength, reducing the fluorescence of the fluorophore through collisional quenching when the fluorophore and quencher are in close contact.
Dye oligo quencher labelings are commonly used to monitor the kinetics of PCR amplification and other biological processes in real time. In one version of this method, the oligonucleotides are designed to hybridize to the 3' side ("downstream") of an amplification primer so that the 5'-3' exonuclease activity of a polymerase digests the 5' end of the probe, cleaving off one of the dyes. The fluorescence intensity of the sample increases and can be monitored as the probe is digested during the course of amplification.
In recent years TAMRA, as well as other fluorescent acceptor molecules in quencher dye dual-labeled probes, have been replaced with one or another of the growing family of dark quencher molecules such as our IQTM, Instant QuencherTM , BHQ1,2,3, Blackberry (BBQ-650), dabcyl, wide range IRDye QC-1 quencher and others.
Bio-Synthesis proivdes oligo quencher labeling services by following stringent standard operation procedures that are in compliant with ISO standards. All of our quencher labeled oligonucleotides are HPLC or dual HPLC purified
Quality Control
All of our oligonucleotides undergo vigorous process monitoring and strict quality control. Length and labeling are systematically controlled by PAGE and characterized by MALDI-TOP mass spectrometry analysis. Quantity is determined and validated by UV Absorbance at 260 nm.
You will receive a Quality Assurance Certificate that includes quantitated yield, melting temperature, MW and µg/OD with every oligo. You will also receive an additional label for use in your records or on an additional vial.
Purification
Desalt or cartridge (RP1) purification is acceptable for most common phosphoramidite modifiers. However, double purification with HPLC is strongly recommended for modifiers that require NHS-ester chemistry to conjugate a dye through an amine linker such as Molecular Probes dyes. Failure sequences are removed during the first HPLC; the second HPLC is performed to remove unlabeled oligonucleotides and excessive dye.
Guaranteed Yield
| Purification Options |
Synthesis Scale 1 |
100 nmol |
250 nmole |
1 umole |
| HPLC |
Guaranteed minimum yield in OD2 |
1 |
2 |
10 |
| dual HPLC 3 |
Guaranteed minimum yield in OD2 |
0.5 |
1 |
5 |
1 Synthesis scale Yield
2 Guaranteed Yield is based on 20-35 mers with single 5' or internal modification; 3' modification Yeilds may be lower
3 Dual HPLC is recommended for modification which require post-synthetic conjugation; Yields may also be lower
Delivery time
Expected shipment time for common dye modifiers is 3 - 5 working days. More exotic modifications are usually completed within 5 - 7 working days.
Packaging
Lyophilized
Shipping Conditions
Room temperature