Abstract
A new approach to monitoring UV damage and repair in the human genome has been developed. The proposed approach is based on a combination of features unique to interspersed repetitive Alu elements, and the ability of certain DNA lesions to block Taq polymerase-mediated DNA synthesis: namely, the extraordinary abundance of Alu repeats throughout the human genome in conjunction with distinct sequence motifs comprising long runs of T residues which are likely targets for formation of UV lesions. Hence, Taq polymerase- mediated extension synthesis with Alu specific primers was employed to visualize formation of discrete predicted adducts within the element. Several variations of the Alu-primer driven amplification protocol were developed to monitor the following aspects of damage: (i) induction of UV-photoproducts at predicted sites within the Alu sequence, (ii) modification of extension synthesis profiles, (iii) UV dose dependent, quantitative inhibition of Alu- primer driven amplification. The assays reveal sites of predicted Taq polymerase blockage within the Alu sequence, a global decrease in the mean length of extension products, and a measurable reduction in the quantity of extension products that is inversely proportional to UV dose. Thus, the exceptional abundance of Alu repeats and their primary sequence features, in combination with the ability of UV lesions to block elongation by Taq polymerase, provide a novel and sensitive system for detecting UV damage in the human genome. The system detects UV damage at levels that are compatible with cellular DNA repair, and provides a unique amplification-based protocol for probing the overall integrity of human DNA.
| Original language | English (US) |
|---|---|
| Pages (from-to) | 31-39 |
| Number of pages | 9 |
| Journal | Mutation Research - DNA Repair |
| Volume | 385 |
| Issue number | 1 |
| DOIs | |
| State | Published - Oct 1997 |
| Externally published | Yes |
Fingerprint
Keywords
- Alu repeat
- DNA damage
- Genomic integrity
- PCR
- UV-photoproduct
ASJC Scopus subject areas
- Toxicology
- Genetics
- Molecular Biology
Cite this
Alu-mediated detection of DNA damage in the human genome. / Englander, Ella; Howard, Bruce H.
In: Mutation Research - DNA Repair, Vol. 385, No. 1, 10.1997, p. 31-39.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Alu-mediated detection of DNA damage in the human genome
AU - Englander, Ella
AU - Howard, Bruce H.
PY - 1997/10
Y1 - 1997/10
N2 - A new approach to monitoring UV damage and repair in the human genome has been developed. The proposed approach is based on a combination of features unique to interspersed repetitive Alu elements, and the ability of certain DNA lesions to block Taq polymerase-mediated DNA synthesis: namely, the extraordinary abundance of Alu repeats throughout the human genome in conjunction with distinct sequence motifs comprising long runs of T residues which are likely targets for formation of UV lesions. Hence, Taq polymerase- mediated extension synthesis with Alu specific primers was employed to visualize formation of discrete predicted adducts within the element. Several variations of the Alu-primer driven amplification protocol were developed to monitor the following aspects of damage: (i) induction of UV-photoproducts at predicted sites within the Alu sequence, (ii) modification of extension synthesis profiles, (iii) UV dose dependent, quantitative inhibition of Alu- primer driven amplification. The assays reveal sites of predicted Taq polymerase blockage within the Alu sequence, a global decrease in the mean length of extension products, and a measurable reduction in the quantity of extension products that is inversely proportional to UV dose. Thus, the exceptional abundance of Alu repeats and their primary sequence features, in combination with the ability of UV lesions to block elongation by Taq polymerase, provide a novel and sensitive system for detecting UV damage in the human genome. The system detects UV damage at levels that are compatible with cellular DNA repair, and provides a unique amplification-based protocol for probing the overall integrity of human DNA.
AB - A new approach to monitoring UV damage and repair in the human genome has been developed. The proposed approach is based on a combination of features unique to interspersed repetitive Alu elements, and the ability of certain DNA lesions to block Taq polymerase-mediated DNA synthesis: namely, the extraordinary abundance of Alu repeats throughout the human genome in conjunction with distinct sequence motifs comprising long runs of T residues which are likely targets for formation of UV lesions. Hence, Taq polymerase- mediated extension synthesis with Alu specific primers was employed to visualize formation of discrete predicted adducts within the element. Several variations of the Alu-primer driven amplification protocol were developed to monitor the following aspects of damage: (i) induction of UV-photoproducts at predicted sites within the Alu sequence, (ii) modification of extension synthesis profiles, (iii) UV dose dependent, quantitative inhibition of Alu- primer driven amplification. The assays reveal sites of predicted Taq polymerase blockage within the Alu sequence, a global decrease in the mean length of extension products, and a measurable reduction in the quantity of extension products that is inversely proportional to UV dose. Thus, the exceptional abundance of Alu repeats and their primary sequence features, in combination with the ability of UV lesions to block elongation by Taq polymerase, provide a novel and sensitive system for detecting UV damage in the human genome. The system detects UV damage at levels that are compatible with cellular DNA repair, and provides a unique amplification-based protocol for probing the overall integrity of human DNA.
KW - Alu repeat
KW - DNA damage
KW - Genomic integrity
KW - PCR
KW - UV-photoproduct
UR - http://www.scopus.com/inward/record.url?scp=0030723214&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=0030723214&partnerID=8YFLogxK
U2 - 10.1016/S0921-8777(97)00036-0
DO - 10.1016/S0921-8777(97)00036-0
M3 - Article
C2 - 9372846
AN - SCOPUS:0030723214
VL - 385
SP - 31
EP - 39
JO - Mutation Research - DNA Repair
JF - Mutation Research - DNA Repair
SN - 0921-8777
IS - 1
ER -