Simple ‘universal’ DNA extraction procedure compatible with direct PCR amplification

1996 ◽  
Vol 52 (4) ◽  
pp. 295-295 ◽  
Author(s):  
D. Goldenberger ◽  
I. Perschil ◽  
M. Ritzler ◽  
M. Altwegg
Keyword(s):  
Dna Extraction ◽  
Extraction Procedure ◽  
Pcr Amplification ◽  
Direct Pcr

scholarly journals A simple "universal" DNA extraction procedure using SDS and proteinase K is compatible with direct PCR amplification.

1995 ◽  
Vol 4 (6) ◽  
pp. 368-370 ◽  
Author(s):  
D Goldenberger ◽  
I Perschil ◽  
M Ritzler ◽  
M Altwegg
Keyword(s):  
Dna Extraction ◽  
Extraction Procedure ◽  
Pcr Amplification ◽  
Proteinase K ◽  
Direct Pcr

scholarly journals A Syringe-Based and Centrifugation-Free DNA Extraction Procedure for the Rapid Detection of Bacteria

Chemosensors ◽  
2021 ◽  
Vol 9 (7) ◽  
pp. 167
Author(s):  
Taehwi Yoon ◽  
Seokjoon Kim ◽  
Jung Ho Kim ◽  
Ki Soo Park
Keyword(s):  
Nucleic Acid ◽  
Dna Extraction ◽  
Extraction Procedure ◽  
Food Poisoning ◽  
Pcr Amplification ◽  
High Sensitivity ◽  
Diagnostic Systems ◽  
Extraction Device ◽  
One Step ◽  
Polymerase Chain

Several bacteria are known to cause food poisoning; therefore, diagnostic systems that detect bacteria are essential. Nucleic acid-based testing methods that involve polymerase chain reaction (PCR) amplification are of great interest due to their high sensitivity and specificity. Herein, we developed a syringe-based one-step DNA extraction device that streamlines the extraction of genomic DNA (gDNA) from bacteria within 2 min, enabling versatile application of nucleic acid-based testing in the field. Notably, the bolt-nut structured case coupled with the syringe enables control of the volume of solution dispensed for enabling DNA extraction without the need for bulky centrifuge equipment. Using the proposed system, the gDNA of a model bacterium, Escherichia coli, was extracted at a good quantity and quality and amplified via PCR. The DNA extracted was comparable to that extracted via a centrifugation-based procedure. In addition, bacteria that were artificially spiked in common samples, including a work cloth, a work bench, and meat, were successfully detected with high accuracy.

scholarly journals Comparison and Validation of Ichthyoplankton DNA Extraction Methods

2021 ◽  
Vol 4 (4) ◽  
pp. 87
Author(s):  
Diouri Lamia ◽  
Uwiringiyeyezu Théophile ◽  
Abdelouahab Hinde ◽  
Malki Mohamed ◽  
Baibai Tarik ◽  
...  
Keyword(s):  
Nucleic Acids ◽  
Dna Extraction ◽  
Classical Method ◽  
Pcr Amplification ◽  
Temporal Distribution ◽  
Extraction Methods ◽  
Direct Pcr ◽  
Fisheries Resources ◽  
Morphological Criteria

Ichthyoplankton is the cluster of planktonic organisms that consists of fish eggs and larvae. These planktonic stages belong to the temporary zooplankton, representing future exploitable stocks. The study of the early ontogenesis of fish plays a key role in the understanding and evaluation of these populations through the study of their abundance and their spatio-temporal distribution. To better understand and protect these fisheries resources, it is essential to identify the different stages of fish embryonic development. This identification is usually performed using the classical method, based on morphological criteria under a binocular magnifying glass; however, this methodology is not always sufficient and is time consuming and, therefore, it is necessary to rely increasingly on molecular tools. The major problem with these tools is the yield and quality of the nucleic acids extracted from ichthyoplankton, especially in the case of eggs, which are small. Several methods have been used for DNA extraction from ichthyoplankton, either automated or manual, but very often from larvae or adults. In the present work, five fish egg DNA extraction protocols were compared based on their DNA yield and extraction quality, verified by agarose gel electrophoresis and quantitative PCR amplification. The results showed that extraction by our heat-protocol for direct PCR (Hp-dPCR) presents the simplest and cheapest protocol of all the kits used in this study, providing a sufficient quantity and quality of nucleic acids to be used for PCR amplification, and being within the reach of third world laboratories that often do not have sufficiently large budgets to obtain automated kits.

scholarly journals Improved direct lysis PCR amplification method of microalgal culture for sequencing and species identification

2021 ◽  
Vol 948 (1) ◽  
pp. 012013
Author(s):  
F Fitriyah ◽  
Y Faramitha ◽  
D A Sari ◽  
I Kresnawaty ◽  
T Panji ◽  
...  
Keyword(s):  
Dna Extraction ◽  
Species Identification ◽  
Dna Barcode ◽  
Pcr Amplification ◽  
Extraction Methods ◽  
Rbcl Gene ◽  
Direct Pcr ◽  
Nannochloropsis Gaditana ◽  
Amplification Method ◽  
Microalgal Culture

Abstract Molecular approach plays important role in species identification for microalgae which involves sequencing of specific DNA barcode present in the genome. This approach involved preparation of template DNA for polymerase chain reaction (PCR) which is time consuming and requires large amounts of algal cells. Microalgal direct PCR have been used frequently for species identification, which simplified the DNA isolation procedure. However, the recent attempts to amplify the rbcL gene of microalga using the previously reported protocol led to poor repeatability. In this study, Nannochloropsis gaditana NIES-2587 was cultured in f/2 liquid medium. The culture growth was estimated on optical density value and the lysis process was improved using gradual temperature procedure during the PCR process. The same culture was extracted using manual DNA extraction method for comparison. The DNA obtained from both methods were amplified using RbclN primer pair to amplify 1486 bp partial sequence of Nannochloropsis rbcL gene, followed by the sequencing of the PCR product. Molecular identification based on the sequence result and BLAST analysis indicated that direct PCR and manual DNA extraction methods successfully produced high sequences result and confirmed the identity of microalgae species into N. gaditana strain CCMP527 with a genetic similarity of >99%.

Quality Improvement of the DNA extracted by boiling method in Gram negative bacteria

2017 ◽  
Vol 6 (04) ◽  
pp. 5347 ◽  
Author(s):  
Omar B. Ahmed* ◽  
Anas S. Dablool
Keyword(s):  
Dna Extraction ◽  
Control Method ◽  
Extraction Procedure ◽  
Cost Effective ◽  
High Yield ◽  
Gram Negative Bacteria ◽  
Bacterial Dna ◽  
Gram Negative ◽  
E Coli

Several methods of Deoxyribonucleic acid (DNA) extraction have been applied to extract bacterial DNA. The amount and the quality of the DNA obtained for each one of those methods are variable. The study aimed to evaluate bacterial DNA extraction using conventional boiling method followed by alcohol precipitation. DNA extraction from Gram negative bacilli was extracted and precipitated using boiling method with further precipitation by ethanol. The extraction procedure performed using the boiling method resulted in high DNA yields for both E. coli and K. pneumoniae bacteria in (199.7 and 285.7μg/ml, respectively) which was close to control method (229.3 and 440.3μg/ml). It was concluded that after alcohol precipitation boiling procedure was easy, cost-effective, and applicable for high-yield quality of DNA in Gram-negative bacteria.

scholarly journals DMSO Improves the Ski-Slope Effect in Direct PCR

2021 ◽  
Vol 11 (4) ◽  
pp. 1943
Author(s):  
Joo-Young Kim ◽  
Ju Yeon Jung ◽  
Da-Hye Kim ◽  
Seohyun Moon ◽  
Won-Hae Lee ◽  
...  
Keyword(s):  
Dna Sequences ◽  
Pcr Amplification ◽  
Analytical Techniques ◽  
Direct Pcr ◽  
Dna Profile ◽  
Novel Technologies ◽  
Specific Amplification ◽  
Polymerase Chain ◽  
Dna Profiles

Analytical techniques such as DNA profiling are widely used in various fields, including forensic science, and novel technologies such as direct polymerase chain reaction (PCR) amplification are continuously being developed in order to acquire DNA profiles efficiently. However, non-specific amplification may occur depending on the quality of the crime scene evidence and amplification methods employed. In particular, the ski-slope effect observed in direct PCR amplification has led to inaccurate interpretations of the DNA profile results. In this study, we aimed to reduce the ski-slope effect by using dimethyl sulfoxide (DMSO) in direct PCR. We confirmed that DMSO (3.75%, v/v) increased the amplification yield of large-sized DNA sequences more than that of small-sized ones. Using 50 Korean buccal samples, we further demonstrated that DMSO reduced the ski-slope effect in direct PCR. These results suggest that the experimental method developed in this study is suitable for direct PCR and may help to successfully obtain DNA profiles from various types of evidence at crime scenes.

Direct PCR amplification of forensic touch and other challenging DNA samples: A review

2018 ◽  
Vol 32 ◽  
pp. 40-49 ◽  
Author(s):  
Sarah E. Cavanaugh ◽  
Abigail S. Bathrick
Keyword(s):  
Pcr Amplification ◽  
Direct Pcr

scholarly journals Direct PCR amplification of DNA from human bloodstains, saliva, and touch samples collected with microFLOQ ® swabs

2018 ◽  
Vol 32 ◽  
pp. 80-87 ◽  
Author(s):  
Angie Ambers ◽  
Rachel Wiley ◽  
Nicole Novroski ◽  
Bruce Budowle
Keyword(s):  
Pcr Amplification ◽  
Direct Pcr

scholarly journals Direct PCR from whole blood, without DNA extraction

1990 ◽  
Vol 18 (19) ◽  
pp. 5908-5909 ◽  
Author(s):  
B. Mercier ◽  
C. Gaucher ◽  
O. Feugeas ◽  
C. Mazurier
Keyword(s):  
Dna Extraction ◽  
Whole Blood ◽  
Direct Pcr
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