What is Big dye Terminator?
BigDye is a set of dye terminators labelled with novel, high-sensitivity dyes . The donor dye is optimised to absorb the excitation energy of the argon ion laser. The linker affords extremely efficient energy transfer (quantum efficiency nearly 1.0) between the donor and acceptor dyes.
How does BigDye Terminator work?
The BigDye® Terminator kit from Applied Biosystems uses four different fluorescent dyes to label ddNTPs, which are added sequentially to the primer through a cycle sequencing reaction. Reaction products can then be run in an automated sequencer to obtain the final sequence.
What is dye terminator sequencing?
Dye terminator sequencing is the modern variant of the dideoxy chain termination sequencing method first pioneered by Dr. Frederick Sanger in 1977 (Nobel Prize in Chemistry). The modern method utilizes fluorescent-labeled dideoxy nucleotide chain terminators for use in a single sequencing reaction.
What is cycle sequencing?
Cycle sequencing is the method required to amplify and fluorescently label your DNA so that it can be detected on our automated sequencers. Cycle sequencing is very similar to PCR, but with two major differences.
What is ExoSAP?
ExoSAP-IT™ PCR Product Cleanup Reagent is used for enzymatic cleanup of amplified PCR product. It hydrolyzes excess primers and nucleotides in a single step. ExoSAP-IT-purified samples are ready for use in downstream applications such as DNA sequencing or single nucleotide polymorphism (SNP) analysis.
Is BigDye light sensitive?
BigDye® Terminator v1. 1 Ready Reaction Mix – CG (part no. 4486486) 800 µL (100 rxns) Store at -25 to -15°C. Protect from light.
What is automated DNA sequencing?
Automated DNA sequencing utilizes a fluorescent dye to label the nucleotides instead of a radioactive isotope. The system automatically identifies the nucleotide sequence and saves the information on the computer. Thus, only a review of the data is necessary to ensure no anomalies were misidentified by the computer.
What are the advantages of dye primer and dye terminator sequencing?
The major advantages of dye-labeled terminator sequencing are convenience, since only a single extension reaction is required for each template, and the synthesis of a labeled primer is unnecessary, allowing the use of preferred hybridization sites.
What are Dideoxynucleotides used for?
Dideoxynucleotides are used to terminate growing DNA chains and create the subsets of truncated fragments in a sequencing reaction.
What is the difference between PCR and cycle sequencing?
PCR is a technique used to duplicate DNA artificially. This is done to have enough quantity of it for the next process which is sequencing. DNA sequencing is a process where the sequence of the bases in DNA is determined for medical, criminal or research uses.