Single gene retrieval from thermally degraded DNA.
To simulate single gene retrieval from ancient DNA, several related factors have been investigated. By monitoring a 889 bp polymerase chain reaction (PCR) product and genomic DNA degradation, we find that heat and oxygen (especially heat) are both crucial factors influencing DNA degradation. The heat influence, mainly represented by temperature and heating time, affects the DNA degradation via DNA depurination followed by cleavage of nearby phosphodiesters. The heating time influence is temperature-dependent. By reactive oxygen species (ROS) scavenging and 1,3-diphenyl-isobenzofuran (DPBF) bleaching experiments the influence of oxygen on DNA thermal degradation was shown to occur via a singlet oxygen pathway. A comparative study of the thermal degradation of cellular DNA and isolated DNA showed that cellular lipids can aggravate DNA thermal degradation. These results confirm the possibility of gene amplification from thermally degraded DNA. They can be used to evaluate the feasibility of the retrieval of single gene from ancient remains.
Zhang
Lianwen
L
Department of Biological Sciences and Biotechnology, Tsinghua University, Beijing 100084, People's Republic of China.
Wu
Qingyu
Q
D000388:Air; D004247:DNA / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D004587:Electrophoresis, Agar Gel / Q000379:methods; D005796:Genes; D006358:Hot Temperature / Q000009:adverse effects*; D009584:Nitrogen; D016133:Polymerase Chain Reaction / Q000379:methods; D017382:Reactive Oxygen Species / Q000378:metabolism; D046940:Synechococcus / Q000737:chemistry / Q000235:genetics; D013997:Time Factors
D017382:Reactive Oxygen Species; D004247:DNA; D009584:Nitrogen