Effect of Different Proteinase K Digest Protocols and Deparaffinization Methods on Yield and Integrity of DNA Extracted From Formalin-fixed, Paraffin-embedded Tissue.
Base Sequence
Cell Extracts
/ chemistry
Cell Line, Tumor
DNA
/ analysis
Endopeptidase K
/ metabolism
Epithelium
/ chemistry
Formaldehyde
/ chemistry
Gene Expression Profiling
Humans
Neoplasms
/ diagnosis
Paraffin Embedding
Proteolysis
/ drug effects
Quality Control
Real-Time Polymerase Chain Reaction
Tissue Fixation
DNA Integrity Number
Illumina FFPE QC Assay
genomic screen tape
quality control
Journal
The journal of histochemistry and cytochemistry : official journal of the Histochemistry Society
ISSN: 1551-5044
Titre abrégé: J Histochem Cytochem
Pays: United States
ID NLM: 9815334
Informations de publication
Date de publication:
03 2020
03 2020
Historique:
pubmed:
12
2
2020
medline:
18
12
2020
entrez:
12
2
2020
Statut:
ppublish
Résumé
DNA extracted from formalin-fixed, paraffin-embedded tissue sections is often inadequate for sequencing, due to poor yield or degradation. We optimized the proteinase K digest by testing increased volume of enzyme and increased digest length from the manufacturer's protocol using 54 biospecimens, performing the digest in centrifuge tubes. Doubling the quantity of proteinase K resulted in a median increase in yield of 96%. Applying the optimized proteinase K protocol to sections deparaffinized on microscope slides generated a further increase in yield of 41%, but only at >50,000 epithelial tumor cells/section. DNA yield now correlated with (χ
Identifiants
pubmed: 32043912
doi: 10.1369/0022155420906234
pmc: PMC7045302
doi:
Substances chimiques
Cell Extracts
0
Formaldehyde
1HG84L3525
DNA
9007-49-2
Endopeptidase K
EC 3.4.21.64
Types de publication
Journal Article
Research Support, Non-U.S. Gov't
Langues
eng
Sous-ensembles de citation
IM
Pagination
171-184Références
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