What is GC-rich PCR?
The GC-RICH PCR System is a blend of Taq DNA Polymerase and a proofreading polymerase for amplifying longer nucleic acid fragments. The GC-Rich Solution provides an solution to enable amplification of all kind of difficult PCR products.
What is considered GC-rich?
When we say “GC-rich” we mean? 60% of the bases are either cytosine (C) or guanine (G.) There are several options available, which alone or in combination may help you to deal with this problem, but first let’s look at why GC-rich sequences are more difficult to amplify.
What is considered high GC content for PCR?
Here are some guidelines for designing your PCR primers: Aim for the GC content to be between 40 and 60% with the 3′ of a primer ending in G or C to promote binding. This is known as a GC Clamp. The G and C bases have stronger hydrogen bonding and help with the stability of the primer.
What does GC content tell you?
Molecular biology In polymerase chain reaction (PCR) experiments, the GC-content of short oligonucleotides known as primers is often used to predict their annealing temperature to the template DNA. A higher GC-content level indicates a relatively higher melting temperature.
Why is GC rich DNA more stable?
From the base-pairing diagram, we can see that the G-C pair has 3 hydrogen bonds, while the A-T pair has only 2. Therefore, the G-C pairing is more stable than the A-T pairing. Thus, strands with more G-C content have more hydrogen bonding, are more stable, and have a greater resistance to denaturation.
What does higher GC content tell you?
A higher GC-content level indicates a relatively higher melting temperature.
Why is it recommended to have a 40% 60% GC content?
GC bonds contribute more to the stability—i.e., increased melting temperatures—of primer and template, binding more than AT bonds. Primers with 40% to 60% GC content ensure stable binding of primer and template.
What does GC content tell us?
This measure indicates the proportion of G and C bases out of an implied four total bases, also including adenine and thymine in DNA and adenine and uracil in RNA. GC-content may be given for a certain fragment of DNA or RNA or for an entire genome.
Why is GC stronger than at?
Adenine pairs with thymine by two hydrogen bonds and cytosine pairs with guanine by three hydrogen bonds (Berg et. Between the G-C base pairs there are 3 hydrogen bonds which makes this bond pair stronger than the A-T base pair.
Why euchromatin is GC rich?
chromosome and arm on Y chromosome which show dark band because of more condensation of that region and trypsin unable to digest that protein so it take more Geimsa stain than GC rich region which is less condensed and having mostly housekeeping genes and called euchromatic region so AT rich take more geimsa stain than …
What kind of polymerase is used in GC 2?
The Advantage GC 2 Polymerase Mix contains Titanium Taq DNA Polymerase and a minor amount of a proofreading polymerase for a balance of high-yield and high-fidelity PCR. Separate tubes of GC 2 PCR buffer and GC-Melt are supplied with the polymerase mix.
Which is the best polymerase mix for hot start PCR?
A hot-start PCR polymerase mix optimized for efficient and accurate amplification of GC-rich (up to 90%) templates. The Advantage GC 2 Polymerase Mix contains Titanium Taq DNA Polymerase and a minor amount of a proofreading polymerase for a balance of high-yield and high-fidelity PCR.
How is PCR amplification of GC-rich templates influenced?
Polymerase Chain Reaction (PCR) Amplification of GC-Rich Templates The efficiency of polymerase chain reaction (PCR) amplification is influenced by the nucleotide composition and sequence of the template DNA.
Which is the best GC melt reagent for PCR?
The combination of DMSO and GC-Melt Reagent allow the amplification of virtually all GC-rich sequences that are resistant to PCR amplification by standard techniques. For a complete kit, use the Advantage GC 2 PCR Kit (Cat. # 639120, 639119) which includes the enzyme blend, GC buffers, and dNTPs, plus GC-rich control template and primers.