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Effect of PCR extension temperature on high-throughput sequencing

Authors :
López-Barragán, María José
Quiñones, Mariam
Cui, Kairong
Lemieux, Jacob
Zhao, Keji
Su, Xin-zhuan
Source :
Molecular & Biochemical Parasitology. Mar2011, Vol. 176 Issue 1, p64-67. 4p.
Publication Year :
2011

Abstract

Abstract: The DNA amplification process can be a source of bias and artifacts, especially when amplifying genomic areas with extreme AT or GC content. The human malaria parasite Plasmodium falciparum has an AT-rich genome, and some of its highly AT-rich regions have been shown to be refractory to polymerase chain reaction (PCR) amplification. Biased amplification may lead to erroneous conclusions for studies investigating genome-wide gene expression, nucleosome position, and copy number variation. Here we compare genome-wide nucleosome coverage in libraries amplified at three different extension temperatures and show that reduction in PCR extension temperature from 70°C to 60°C can greatly increase the fraction of coverage at AT-rich regions of the P. falciparum genome. Our method will improve the efficiency and coverage in sequencing an AT-rich genome. [ABSTRACT FROM AUTHOR]

Details

Language :
English
ISSN :
01666851
Volume :
176
Issue :
1
Database :
Academic Search Index
Journal :
Molecular & Biochemical Parasitology
Publication Type :
Academic Journal
Accession number :
57514999
Full Text :
https://doi.org/10.1016/j.molbiopara.2010.11.013