A single-container procedure for multiple genomic DNA samples preparation for pulsed field gel electrophoresis. 2022

Karen León Arcia, and Heidi Quintero Álvarez, and Laura María Pantoja-Echevarría, and Yainelis Garrido Nicot
Molecular Genetics Department, Cuban Neuroscience Center, 190 St and 25 St, No. 2503. Cubanacan, Playa, Havana 11600, Cuba. Electronic address: kla@cneuro.edu.cu.

Genomic DNA preparation is a critical step for successful fingerprinting analysis by Pulsed Field Gel Electrophoresis (PFGE). This paper presents a simple and rapid protocol to prepare DNA samples from up to 24 bacterial isolates simultaneously. It involves performing the conventional PFGE sample preparation steps (cell growth and harvest, agarose-immobilization of intact cells, and DNA release and purification) into a 24-wells culture plate, without subjecting the biological material to repeated transference of containers. The single-container protocol rendered high quality genomic DNA from E. coli clinical isolates. The DNA yields obtained from samples prepared with the single-container protocol showed no differences to those obtained using the conventional DNA sample preparation for PFGE. This procedure is a cost-effective alternative that provides a larger capacity of analysis to the PFGE technique, which could be advantageous in a context of infectious disease outbreaks for pathogens molecular subtyping.

UI MeSH Term Description Entries
D004196 Disease Outbreaks Sudden increase in the incidence of a disease. The concept includes EPIDEMICS and PANDEMICS. Outbreaks,Infectious Disease Outbreaks,Disease Outbreak,Disease Outbreak, Infectious,Disease Outbreaks, Infectious,Infectious Disease Outbreak,Outbreak, Disease,Outbreak, Infectious Disease,Outbreaks, Disease,Outbreaks, Infectious Disease
D004247 DNA A deoxyribonucleotide polymer that is the primary genetic material of all cells. Eukaryotic and prokaryotic organisms normally contain DNA in a double-stranded state, yet several important biological processes transiently involve single-stranded regions. DNA, which consists of a polysugar-phosphate backbone possessing projections of purines (adenine and guanine) and pyrimidines (thymine and cytosine), forms a double helix that is held together by hydrogen bonds between these purines and pyrimidines (adenine to thymine and guanine to cytosine). DNA, Double-Stranded,Deoxyribonucleic Acid,ds-DNA,DNA, Double Stranded,Double-Stranded DNA,ds DNA
D004269 DNA, Bacterial Deoxyribonucleic acid that makes up the genetic material of bacteria. Bacterial DNA
D004926 Escherichia coli A species of gram-negative, facultatively anaerobic, rod-shaped bacteria (GRAM-NEGATIVE FACULTATIVELY ANAEROBIC RODS) commonly found in the lower part of the intestine of warm-blooded animals. It is usually nonpathogenic, but some strains are known to produce DIARRHEA and pyogenic infections. Pathogenic strains (virotypes) are classified by their specific pathogenic mechanisms such as toxins (ENTEROTOXIGENIC ESCHERICHIA COLI), etc. Alkalescens-Dispar Group,Bacillus coli,Bacterium coli,Bacterium coli commune,Diffusely Adherent Escherichia coli,E coli,EAggEC,Enteroaggregative Escherichia coli,Enterococcus coli,Diffusely Adherent E. coli,Enteroaggregative E. coli,Enteroinvasive E. coli,Enteroinvasive Escherichia coli
D015373 Bacterial Typing Techniques Procedures for identifying types and strains of bacteria. The most frequently employed typing systems are BACTERIOPHAGE TYPING and SEROTYPING as well as bacteriocin typing and biotyping. Bacteriocin Typing,Biotyping, Bacterial,Typing, Bacterial,Bacterial Biotyping,Bacterial Typing,Bacterial Typing Technic,Bacterial Typing Technics,Bacterial Typing Technique,Technic, Bacterial Typing,Technics, Bacterial Typing,Technique, Bacterial Typing,Techniques, Bacterial Typing,Typing Technic, Bacterial,Typing Technics, Bacterial,Typing Technique, Bacterial,Typing Techniques, Bacterial,Typing, Bacteriocin
D016172 DNA Fingerprinting A technique for identifying individuals of a species that is based on the uniqueness of their DNA sequence. Uniqueness is determined by identifying which combination of allelic variations occur in the individual at a statistically relevant number of different loci. In forensic studies, RESTRICTION FRAGMENT LENGTH POLYMORPHISM of multiple, highly polymorphic VNTR LOCI or MICROSATELLITE REPEAT loci are analyzed. The number of loci used for the profile depends on the ALLELE FREQUENCY in the population. DNA Fingerprints,DNA Profiling,DNA Typing,Genetic Fingerprinting,DNA Fingerprint,DNA Fingerprintings,DNA Profilings,DNA Typings,Fingerprint, DNA,Fingerprinting, DNA,Fingerprinting, Genetic,Fingerprintings, DNA,Fingerprintings, Genetic,Fingerprints, DNA,Genetic Fingerprintings,Profiling, DNA,Typing, DNA,Typings, DNA
D016521 Electrophoresis, Gel, Pulsed-Field Gel electrophoresis in which the direction of the electric field is changed periodically. This technique is similar to other electrophoretic methods normally used to separate double-stranded DNA molecules ranging in size up to tens of thousands of base-pairs. However, by alternating the electric field direction one is able to separate DNA molecules up to several million base-pairs in length. Electrophoresis, Gel, Pulsed-Field Gradient,Gel Electrophoresis, Pulsed-Field,Contour-Clamped Homogeneous-Field Gel Electrophoresis,Electrophoresis, Gel, Pulsed Field,Electrophoresis, Pulsed Field Gel,Field Inversion Gel Electrophoresis,Orthogonal Field Alternation Gel Electrophoresis,Orthogonal-Field Alternation-Gel Electrophoresis,Pulsed Field Gradient Gel Electrophoresis,Pulsed-Field Gel Electrophoresis,Pulsed-Field Gradient Gel Electrophoresis,Alternation-Gel Electrophoresis, Orthogonal-Field,Contour Clamped Homogeneous Field Gel Electrophoresis,Electrophoresis, Orthogonal-Field Alternation-Gel,Electrophoresis, Pulsed-Field Gel,Gel Electrophoresis, Pulsed Field,Pulsed Field Gel Electrophoresis

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