Large fragment Bst DNA polymerase for whole genome amplification of DNA from formalin-fixed paraffin-embedded tissues.
Author(s): Aviel-Ronen S, Qi Zhu C, Coe BP, Liu N, Watson SK, Lam WL, Tsao MS
Publication: BMC Genomics, 2006, Vol. 7, Page 312
PubMed ID: 17156491 PubMed Review Paper? No
Purpose of Paper
Conclusion of Paper
Studies
-
Study Purpose
The purpose of this study was to determine the effects of WGA on copy number detection using DNA from frozen and FFPE lung specimens. FFPE specimens were deparaffinized with toluene, digested with proteinase K overnight and purified with Phase Lock Gel. The method by which DNA was extracted from frozen specimens was unspecified.
Summary of Findings:
Using Bst DNA polymerase, slightly better WGA was possible from FFPE tissue than from snap-frozen specimens (mean amplification yields of 1035-fold versus 835-fold). Amplified products from FFPE and snap-frozen tissue were of a high molecular weight. After Bst amplification, relative gene copy numbers by real-time qPCR remained within 3-fold of unamplified specimens. Similarly, using arrays, copy number differences between matched Bst-amplified and unamplified specimens were less than 3-fold. In contrast to Bst-amplified DNA, the authors report that Phi29-amplified DNA from FFPE tissue failed to amplify during real-time qPCR.
Biospecimens
Preservative Types
- Formalin
- Frozen
Diagnoses:
- Normal
- Neoplastic - Carcinoma
Platform:
Analyte Technology Platform DNA Array CGH DNA Electrophoresis DNA Real-time qPCR DNA Whole genome amplification Pre-analytical Factors:
Classification Pre-analytical Factor Value(s) Whole genome amplification Specific Nucleic acid amplification BST DNA polymerase
Phi29 DNA polymerase
Real-time qPCR Specific Targeted nucleic acid GAPDH
Skp2
PIK3R1
NMYC
SS18L2
GHR
COPS5
LATS2
Biospecimen Preservation Type of fixation/preservation Formalin (buffered)
Snap frozen