Molecular Beacons

• Structure and Function of Molecular Beacons
• Molecular Beacon Design and Function
• Benefits of Using Molecular Beacons
• Related Product Categories

Molecular Beacons are a type of oligonucleotide probe (short singled-stranded nucleic acid sequences between 30–50 bases) used in molecular biology and genetics research. They are designed to have a unique sequence flanked by indirect repeats so that a stem-loop structure is formed, allowing the 5′ and 3′ ends to be maintained in proximity. Molecular Beacon probes fluoresce upon hybridization to the target. These structured probes are highly sensitive, sequence specific, and are used for sequence detection in qPCR for in vitro studies.

STRUCTURE AND FUNCTION OF MOLECULAR BEACONS

A Molecular Beacon is a single-stranded bi-labeled fluorescent probe held in a hairpin-loop conformation (20 to 25 nt is most common) by complementary stem sequences (around 4 to 6 nt) at both ends of the probe. The 5’ and 3’ ends of the probe contain a reporter and a quencher molecule, respectively. The loop is a single-stranded DNA sequence complementary to the target sequence. The proximity of the reporter (fluorophore, or fluorescent dye) and quencher causes the quenching of the natural fluorescence emission of the reporter by bringing about energy transfer. The structure and mechanism of a molecular beacon is shown below (Figure 1).

MOLECULAR BEACON DESIGN AND FUNCTION

• Molecular Beacon probes are designed with four essential parts: loop, stem, quencher, and reporter.
• The stem and loop regions of the Molecular Beacon form a stable hairpin structure, bringing the fluorophore and quencher in close proximity. This close proximity results in the quencher absorbing the fluorescence emitted by the fluorophore, effectively suppressing the signal.
• When Molecular Beacons encounter the complementary target sequence, the hairpin-loop structure opens and separates the 5’-end reporter from the 3’-end quencher. As the quencher is no longer in proximity to the reporter, the fluorescence of the reporter is no longer quenched, thereby allowing fluorescence emission. 
• The intensity of the fluorescence signal is directly proportional to the amount of target nucleic acid present in the sample.
• In real-time PCR, when added prior to amplification, Molecular Beacons undergo disruption of base-pairing in the stem. This allows the probe to hybridize to the complementary target upon denaturation, which separates the reporter dye and quencher, thereby inducing reporter dye fluorescence.

Figure 1. How Molecular Beacons Work

Benefits of Using Molecular Beacons

• Increased specificity and sensitivity
• Real-time detection capabilities
• Ideal for single nucleotide polymorphism (SNP) profiling

ADD LOCKED NUCLEIC ACID TO YOUR BEACON PROBE

• Increase thermal stability and hybridization specificity
• Obtain greater accuracy in SNP detection, allele discrimination and in vitro quantification or detection
  
• Achieve easier and more sensitive probe designs for problematic target sequences

Product Features

• Amounts: 1, 3, 5, and 10 OD
• Purification: HPLC
• Sequence Lengths: 15 - 40 bases
• Quality Control: 100% mass spectrometry
• Format: Supplied dry in amber tubes
• Custom formats available (normalizations, special plates, etc.)

Our probes are provided in a format to simplify your experimental planning.

Applications of Molecular Beacons

• SNP detection
• Allele discrimination
• Pathogen detection
• Multiplexing
• Viral load quantification
• Gene expression analysis
• Gene copy determination
• Endpoint genotyping
• in vitro quantification or detection

Guaranteed Yields

Guaranteed OD Yield	Approx. nmol	Approx. µg	Approx. No. of Reactions*
1	3	32	600
3	9	96	1,800
5	15	160	3,000
10	30	320	6,000

*Estimate is based on 3 nmol or 32 µg for 1 OD and 200 nM in 25 µL reaction (5.0 pmol/reaction). Estimate is based on an average sequence length of 30 bases.

The most common fluorophore and quencher combinations are listed below:

Spectral Properties Table

Dye	Max. EX (nm)	Max. EM (nm)	Compatible Quencher
6-FAM™	495	520	BHQ™-1, DABCYL
JOE™	529	555	BHQ™-1, DABCYL
TET™	521	536	BHQ™-1, DABCYL
HEX™	535	556	BHQ™-1, DABCYL
Cyanine 3	549	566	BHQ™-2, DABCYL
ROX™	586	610	BHQ™-2, DABCYL
TxRd (Sulforhodamine 101-X)	597	616	BHQ™-2, DABCYL
Cyanine 5	646	669	BHQ™-3, DABCYL
Cyanine 5.5	675	694	BHQ™-3, DABCYL

Shipping Schedule

Quantity (OD)	1	3	5	10
Molecular Beacons with Standard Dyes	5 days	5 days	5 days	5 days
Locked Nucleic Acid Molecular Beacons with Standard Dyes	7 days	7 days	7 days	7 days
Molecular Beacons with Post-Synthesis Dyes, with/without Locked Nucleic Acid	7-10 days	7-10 days	7-10 days	7-10 days

*Delivery times may be longer due to international transit, customs clearance delays, etc. Large projects will be placed on a delivery schedule based upon project needs.

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