title: Comparison of eleven methods for genomic DNA extraction suitable for large-scale whole-genome genotyping and long-term DNA banking using blood samples.
abstract: Over the recent years, next generation sequencing and microarray technologies have revolutionized scientific research with their applications to high-throughput analysis of biological systems. Isolation of high quantities of pure, intact, double stranded, highly concentrated, not contaminated genomic DNA is prerequisite for successful and reliable large scale genotyping analysis. High quantities of pure DNA are also required for the creation of DNA-banks. In the present study, eleven different DNA extraction procedures, including phenol-chloroform, silica and magnetic beads based extractions, were examined to ascertain their relative effectiveness for extracting DNA from ovine blood samples. The quality and quantity of the differentially extracted DNA was subsequently assessed by spectrophotometric measurements, Qubit measurements, real-time PCR amplifications and gel electrophoresis. Processing time, intensity of labor and cost for each method were also evaluated. Results revealed significant differences among the eleven procedures and only four of the methods yielded satisfactory outputs. These four methods, comprising three modified silica based commercial kits (Modified Blood, Modified Tissue, Modified Dx kits) and an in-house developed magnetic beads based protocol, were most appropriate for extracting high quality and quantity DNA suitable for large-scale microarray genotyping and also for long-term DNA storage as demonstrated by their successful application to 600 individuals.
authors: Animal Production Laboratory, School of Veterinary Medicine, Aristotle University of Thessaloniki, Thessaloniki, Greece; The Roslin Institute and Royal (Dick) School of Veterinary Studies, University of Edinburgh, Edinburgh, United Kingdom.
authors: Androniki
authors: A
authors: Psifidi
authors: Microbiology and Infectious Diseases Laboratory, School of Veterinary Medicine, Aristotle University of Thessaloniki, Thessaloniki, Greece.
authors: Chrysostomos I
authors: CI
authors: Dovas
authors: Animal Production Laboratory, School of Veterinary Medicine, Aristotle University of Thessaloniki, Thessaloniki, Greece.
authors: Georgios
authors: G
authors: Bramis
authors: Food safety Laboratory, School of Veterinary Medicine, Aristotle University of Thessaloniki, Thessaloniki, Greece.
authors: Thomai
authors: T
authors: Lazou
authors: Department of Clinical Veterinary Sciences, University of Bristol, Langford House, Langford, Bristol, United Kingdom.
authors: Claire L
authors: CL
authors: Russel
authors: Animal Production Laboratory, School of Veterinary Medicine, Aristotle University of Thessaloniki, Thessaloniki, Greece.
authors: Georgios
authors: G
authors: Arsenos
authors: Animal Production Laboratory, School of Veterinary Medicine, Aristotle University of Thessaloniki, Thessaloniki, Greece; The Roslin Institute and Royal (Dick) School of Veterinary Studies, University of Edinburgh, Edinburgh, United Kingdom; Scotland's Rural College, Edinburgh, United Kingdom.
authors: Georgios
authors: G
authors: Banos
mesh_terms: D000818:Animals; D003365:Costs and Cost Analysis; D015925:Cryopreservation; D004247:DNA / Q000097:blood / Q000302:isolation & purification*; D005260:Female; D060005:Genotyping Techniques; D059014:High-Throughput Nucleotide Sequencing / Q000191:economics / Q000592:standards; D020411:Oligonucleotide Array Sequence Analysis; D020641:Polymorphism, Single Nucleotide; D060888:Real-Time Polymerase Chain Reaction / Q000592:standards; D012015:Reference Standards; D017422:Sequence Analysis, DNA / Q000191:economics / Q000592:standards; D034561:Sheep, Domestic
chemical_list: D004247:DNA
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