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== Size and other limitations ==
PCR works readily with a DNA template of up to two to three thousand base pairs in length. However, above this size, product yields often decrease, as with increasing length [[stochastic]] effects such as premature termination by the polymerase begin to affect the efficiency of the PCR. It is possible to amplify larger pieces of up to 50,000 base pairs with a slower heating cycle and special polymerases. These are polymerases [[fusion protein|fused]] to a processivity-enhancing DNA-binding protein, enhancing adherence of the polymerase to the DNA.<ref>{{cite journal |vauthors=Pavlov AR, Belova GI, Kozyavkin SA, Slesarev AI |year= 2002|title=Helix-hairpin-helix motifs confer salt resistance and processivity on chimeric DNA polymerases|journal=Proc. Natl. Acad. Sci. U.S.A.|volume=99|pages=3510–13515|pmid=12368475|doi=10.1073/pnas.202127199 |issue=21 |pmc=129704|bibcode= 2002PNAS...9913510P|doi-access= free}}</ref><ref>{{cite journal |author=Demidov VV|year=2002|title= A happy marriage: advancing DNA polymerases with DNA topoisomerase supplements|journal=Trends Biotechnol.|volume=20|pages=491|doi=10.1016/S0167-7799(02)02101-7 |issue=12}}</ref>
Other valuable properties of the chimeric polymerases [http://www.fidelitysystems.com/TopoTaq.html TopoTaq] and PfuC2 include enhanced thermostability, specificity and resistance to contaminants and [[polymerase chain reaction inhibitors|inhibitors]].<ref>{{cite journal |vauthors=Pavlov AR, Pavlova NV, Kozyavkin SA, Slesarev AI |year= 2004|title=Recent developments in the optimization of thermostable DNA polymerases for efficient applications|journal= Trends Biotechnol.|volume=22|pages= 253–260|pmid= 15109812|doi=10.1016/j.tibtech.2004.02.011 |issue=5}}</ref><ref>{{cite book |chapter-url=http://www.horizonpress.com/hsp/abs/absdna.html|vauthors=Pavlov AR, Pavlova NV, Kozyavkin SA, Slesarev AI |year=2004|chapter=Thermostable Chimeric DNA Polymerases with High Resistance to Inhibitors|title=DNA Amplification: Current Technologies and Applications|publisher=Horizon Bioscience|pages=3–20|isbn=0-9545232-9-6}}</ref> They were engineered using the unique [[helix-hairpin-helix]] (HhH) DNA binding domains of [[topoisomerase]] V<ref>{{cite journal |author=Forterre P|year= 2006|title=DNA topoisomerase V: a new fold of mysterious origin|journal= Trends Biotechnol.|volume=24|pages= 245–247|pmid=16650908|doi=10.1016/j.tibtech.2006.04.006 |issue=6}}</ref> from hyperthermophile ''[[Methanopyrus]] kandleri''. Chimeric polymerases overcome many limitations of native enzymes and are used in direct PCR amplification from cell cultures and even [[food sampling|food samples]], thus by-passing laborious DNA isolation steps. A robust strand-displacement activity of the hybrid TopoTaq polymerase helps solve PCR problems that can be caused by [[Stem-loop|hairpins]] and [[Gyromagnetic ratio|G-loaded]] double helices. Helices with a high G-C content possess a higher melting temperature, often impairing PCR, depending on the conditions.<ref name=Hybrid>{{cite book|chapter-url=http://bioscience.jbpub.com/catalog/0763733830/table_of_contents.htm |vauthors=Pavlov AR, Pavlova NV, Kozyavkin SA, Slesarev AI |year= 2006|chapter=Thermostable DNA Polymerases for a Wide Spectrum of Applications: Comparison of a Robust Hybrid TopoTaq to other enzymes|title=DNA Sequencing II: Optimizing Preparation and Cleanup|editor=Kieleczawa J|publisher=Jones and Bartlett|pages=241–257|isbn=0-7637-3383-0}}</ref>
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