Hello, A Primer dimer is a potential by-product in PCR reaction. Primer dimer consists of primer molecules that have attached, hybridized to each other because of strings of complementary bases in the primers. As a result, the DNA polymerase amplifies the Primer dimer, leading to competition for PCR reagents, thus potentially inhibiting amplification of the DNA sequence targeted for PCR amplification. By the time you can see primer dimer a lot of primer has been removed so you will often find that if you run many samples the samples with primer dimer have amplified less well ( weaker band) than samples without primer dimer. It can often stop an amplification working completely and is often caused either by poor primer design ( homology at the 3' ends of the primers) or by leaving the reaction mixture for too long before starting cycling. Primer dimers will never be as big as your amplicons and if you running a 100 bp ladder the lowest band of ladder would be 100 bp and primer dimers would always would be lesser than that lower band of marker, to reduce primer dimers you can do touch down PCR.
Hope this info will help you.
Good luck!
Your PCR product is too low or your PCR didn't work well.
you can solve this problem by adjusting the PCR condition and DNA concentration .
Hello, A Primer dimer is a potential by-product in PCR reaction. Primer dimer consists of primer molecules that have attached, hybridized to each other because of strings of complementary bases in the primers. As a result, the DNA polymerase amplifies the Primer dimer, leading to competition for PCR reagents, thus potentially inhibiting amplification of the DNA sequence targeted for PCR amplification. By the time you can see primer dimer a lot of primer has been removed so you will often find that if you run many samples the samples with primer dimer have amplified less well ( weaker band) than samples without primer dimer. It can often stop an amplification working completely and is often caused either by poor primer design ( homology at the 3' ends of the primers) or by leaving the reaction mixture for too long before starting cycling. Primer dimers will never be as big as your amplicons and if you running a 100 bp ladder the lowest band of ladder would be 100 bp and primer dimers would always would be lesser than that lower band of marker, to reduce primer dimers you can do touch down PCR.
Hope this info will help you.
Good luck!
Mostly primer dimers are seen due to high concentration of the primer in the PCR reaction mixture. Or if there is no PCR amplification and you can see on primer dimer in the agarose gel.
If are able to see the PCR product then you can reduce the concentration of the prime and keep the other conditions same.
And you are not able to see any PCR amplification then you need standardize it by checking various primer annealing temperature and also you can play around with the DNA template concentration.
@ Dr. Fadia Al-Khayat
A Primer dimer (PD) is a potential by-product in PCR, a common biotechnological method. As its name implies, a Primer dimer consists of primer molecules that have attached (hybridized) to each other because of strings of complementary bases in the primers. As a result, the DNA polymerase amplifies the Primer dimer , leading to competition for PCR reagents, thus potentially inhibiting amplification of the DNA sequence targeted for PCR amplification. In quantitative PCR, Primer dimer may interfere with accurate quantification. The most efficient way to minimize primer dimer potential is through primer design.
Primer dimers may be visible after gel electrophoresis of the PCR product. PDs in ethidium bromide-stained gels are typically seen as a 30-50 base-pair (bp) band or smear of moderate to high intensity and distinguishable from the band of the target sequence, which is typically longer than 50 bp.
In quantitative PCR, PDs may be detected by melting curve analysis with intercalating dyes, such as SYBR Green I, a nonspecific dye for detection of double-stranded DNA. Because they usually consist of short sequences, the PDs denaturate at lower temperature than the target sequence and hence can be distinguished by their melting-curve characteristics.
For detail you can follow these references:
1.Chou, Quin; Russell, Marion; Birch, David E.; Raymond, Jonathan; Bloch, Will (1992). "Prevention of pre-PCR mis-priming and primer dimerization improves low-copy-number amplifications". Nucleic Acids Research. 20 (7): 1717–23. doi:10.1093/nar/20.7.1717. PMC 312262. PMID 1579465.
2. Brownie, Jannine; Shawcross, Susan; Theaker, Jane; Whitcombe, David; Ferrie, Richard; Newton, Clive; Little, Stephen (1997). "The elimination of primer-dimer accumulation in PCR". Nucleic Acids Research. 25 (16): 3235–41. doi:10.1093/nar/25.16.3235. PMC 146890. PMID 9241236.
3. Alberts; et al. (2017). Molecular Biology of the Cell (6th ed.). Garland Science. pp. 708–711.
4. Matheson, Hill (2009). "Review of methods of selective primer-dimer reduction in vitro". Methods in Biomolecular Research. 31 (7): 476–488.
It's mean that you have put too much primer or your sample doesn't have much DNA.
everything said above about good primer design is correct and good advice. If you are stuck with badly designed primers and a lack of time or money and a primer dimer problem then the best try to get rid of the PD is to use a hot start enzyme then the temperature never drops low enough during the pcr cycle to allow dimer formation. Hot start can be achieved either with a commercial hot start taq or just by heating the reaction mix to 80c then adding the ordinary enzyme and immediately starting cycling
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