scholarly journals Low-Cost Imaging of Fluorescent DNA in Agarose Gel Electrophoresis Using Raspberry Pi Cameras

2021 ◽  
Author(s):  
Hassan Ali Abid ◽  
Jian Wern Ong ◽  
Eric Shen Lin ◽  
Zhixiong Song ◽  
Oi Wah Liew ◽  
...  
Keyword(s):  
Gel Electrophoresis ◽  
Low Cost ◽  
Raspberry Pi ◽  
Agarose Gel ◽  
Agarose Gel Electrophoresis ◽  
Noise Variance ◽  
Uv Led ◽  
Resource Limited ◽  
Laboratory Investigations ◽  
Polymerase Chain

Abstract Low-cost analytical solutions built around microcomputers like the Raspberry Pi help to facilitate laboratory investigations in resource limited venues. Here, three camera modules (V1.3 with and without filter, as well as NoIR) that work with this microcomputer were assessed for their suitability in imaging fluorescent DNA following agarose gel electrophoresis. Evaluation of their utility was based on signal-to-noise (SNR) and noise variance metrics that were developed. Experiments conducted with samples were subjected to Polymerase Chain Reaction (PCR), and the amplified products were separated using gel electrophoresis and stained with Midori green. Image analysis revealed the NoIR camera performed the best with SNR and noise variance values of 21.7 and 0.222 respectively. In experiments conducted using UV LED lighting to simulate ethidium bromide (EtBr) excitation, the NoIR and V1.3 with filter removed cameras showed comparable SNR values.

scholarly journals The landscape of microsatellites in the enset (Ensete ventricosum) genome and web-based marker resource development

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Manosh Kumar Biswas ◽  
Jaypal N. Darbar ◽  
James S. Borrell ◽  
Mita Bagchi ◽  
Dhiman Biswas ◽  
...  
Keyword(s):  
Gel Electrophoresis ◽  
Low Cost ◽  
Rural Livelihoods ◽  
In Silico Analysis ◽  
Agarose Gel ◽  
Agarose Gel Electrophoresis ◽  
Web Based ◽  
Ensete Ventricosum ◽  
Link Type ◽  
Nucleotide Repeats

Abstract Ensete ventricosum (Musaceae, enset) is an Ethiopian food security crop. To realize the potential of enset for rural livelihoods, further knowledge of enset diversity, genetics and genomics is required to support breeding programs and conservation. This study was conducted to explore the enset genome to develop molecular markers, genomics resources, and characterize enset landraces while giving insight into the organization of the genome. We identified 233 microsatellites (simple sequence repeats, SSRs) per Mbp in the enset genome, representing 0.28% of the genome. Mono- and di-nucleotide repeats motifs were found in a higher proportion than other classes of SSR-motifs. In total, 154,586 non-redundant enset microsatellite markers (EMM) were identified and 40 selected for primer development. Marker validation by PCR and low-cost agarose gel electrophoresis revealed that 92.5% were polymorphic, showing a high PIC (Polymorphism Information Content; 0.87) and expected heterozygosity (He = 0.79–0.82). In silico analysis of genomes of closely related species showed 46.86% of the markers were transferable among enset species and 1.90% were transferable to Musa. The SSRs are robust (with basic PCR methods and agarose gel electrophoresis), informative, and applicable in measuring enset diversity, genotyping, selection and potentially breeding. Enset SSRs are available in a web-based database at https://enset-project.org/[email protected] (or https://enset.aau.edu.et/index.html, downloadable from Figshare).

Molecular Beacon Polymerase Chain Reaction Detection of Escherichia coli O157:H7 in Milk

2000 ◽  
Vol 63 (7) ◽  
pp. 855-859 ◽  
Author(s):  
JOHN L. McKILLIP ◽  
MARYANNE DRAKE
Keyword(s):  
Escherichia Coli ◽  
Polymerase Chain Reaction ◽  
Gel Electrophoresis ◽  
Molecular Beacon ◽  
Skim Milk ◽  
Agarose Gel ◽  
Escherichia Coli O157 ◽  
Agarose Gel Electrophoresis ◽  
Chain Reaction ◽  
Polymerase Chain

A fluorescently labeled oligonucleotide probe (molecular beacon) was applied to detect Escherichia coli O157:H7 in artificially contaminated skim milk during polymerase chain reaction (PCR) amplification of extracted DNA. The probe was designed to hybridize with a region of the slt-II gene coding for the A subunit and to fluoresce when the hairpin-stem conformation was linearized upon hybridization to the target sequence. The molecular beacon was incorporated into PCR reactions containing DNA extracted from artificially contaminated skim milk. The degree of fluorescence was monitored in PCR reactions containing 103, 105, and 107 CFU of E. coli O157:H7 per ml and was found to correlate with the amount of template in each reaction. Fluorescence significantly increased above background levels by cycle 8, 14, or 14 in reactions containing DNA from the 107-, 105-, or 103-CFU/ml template, respectively (P < 0.05). Molecular beacon PCR demonstrated positive results more rapidly than traditional agarose gel electrophoresis analysis of PCR products. Use of molecular beacons allows real-time monitoring of PCR reactions, and the closed-tube format allows simultaneous detection and confirmation of target amplicons without the need for agarose gel electrophoresis and/or Southern blotting. This is the first report of a stem-and-loop molecular beacon being applied for direct detection of a pathogen in food.

scholarly journals Polyvinylpyrrolidone-Agarose Gel Electrophoresis Purification of Polymerase Chain Reaction-Amplifiable DNA from Soils

1993 ◽  
Vol 59 (6) ◽  
pp. 1972-1974 ◽  
Author(s):  
Charles C. Young ◽  
Robert L. Burghoff ◽  
Lois G. Keim ◽  
Vera Minak-Bernero ◽  
James R. Lute ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Gel Electrophoresis ◽  
Agarose Gel ◽  
Agarose Gel Electrophoresis ◽  
Chain Reaction ◽  
Polymerase Chain

scholarly journals Detection of Acanthamoeba spp. in two major water reservoirs in the Philippines

2020 ◽  
Vol 18 (2) ◽  
pp. 118-126 ◽  
Author(s):  
Giovanni Milanez ◽  
Frederick Masangkay ◽  
Frieda Hapan ◽  
Thea Bencito ◽  
Marcus Lopez ◽  
...  
Keyword(s):  
Water Samples ◽  
Gel Electrophoresis ◽  
Water Security ◽  
The Philippines ◽  
Agarose Gel ◽  
Agarose Gel Electrophoresis ◽  
Water Reservoirs ◽  
Chain Reaction ◽  
Pcr Products ◽  
Polymerase Chain

Abstract Water reservoirs are important manmade structures providing water security to deliver clean and safe water for drinking and other purposes to the community. Eighty water samples were collected from Magat and Ipo water reservoirs using purposive sampling between November 2018 and January 2019. Water samples were collected in sterile containers for testing. The samples were cultured in non-nutrient agar and lawned with Escherichia coli and incubated at 33 °C. Twelve out of the 80 (15%) water samples were positive for amoebic growth. Light and scanning electron microscopy (SEM) revealed double-walled cystic stages and were initially identified as Acanthamoeba spp. based on morphological characteristic in reference to Page's established criteria. Their extracted DNAs were used in polymerase chain reaction using JDP1 and JDP2 primers and confirmed the presence of Acanthamoeba DNA in agarose gel electrophoresis. Aligned sequences from PCR products were deposited in GenBank under accession numbers MK886460, MK909919, MK905437, MK910997, MK911021 and MK886514. The presence of potentially pathogenic Acanthamoeba spp. in water reservoirs is considered a potential risk for public health, requiring appropriate processing of water in treatment plants.

scholarly journals Sexing as a tool for reinsertion of Amazona aestiva parrots to nature: use of less invasive technique

2021 ◽  
Vol 10 (12) ◽  
pp. e380101220601
Author(s):  
Carine Novato Ramos ◽  
Tácita Borges Barros ◽  
Edma Santos de Antonio ◽  
Bernardo Mirabal ◽  
Márcio Borba da Silva ◽  
...  
Keyword(s):  
Gel Electrophoresis ◽  
Invasive Technique ◽  
Molecular Sexing ◽  
Agarose Gel ◽  
Agarose Gel Electrophoresis ◽  
Chain Reaction ◽  
Behavioral Studies ◽  
Sample Composition ◽  
Polymerase Chain ◽  
Primer Sets

The sexing birds is considered an important tool for behavioral studies and programs for the reintroduction of animals into the wild. Several techniques are used for this purpose, such as laparoscopy, magnetic resonance and molecular sexing. The first are considered more invasive and stressful for the animal, and the last is considered the most accurate. According to it, the aim of this study was to compare the effectiveness of using three sets of primers in the molecular sexing process of true parrots (Amazona aestiva). Blood samples from 10 animals were collected at a Wildlife Screening Center (CETAS) in Bahia, Brazil. The DNA was extracted and the molecular markers amplified by Polymerase Chain Reaction (PCR) using primer pairs P2/P8, 1237L/1272H and 2250F/2718R. The amplified material was visualized with electrophoresis performed at 2% agarose and 12.5% polyacrylamide gels. Among the primer sets used, the 2250F/2718R pair showed the best results for the sexing process, including visualization of the amplified products on an agarose gel. Agarose gel electrophoresis is considered to be faster and cheaper. The results revealed a sample composition of 5 males (0.5) and 5 females (0.5).

scholarly journals TECHNICAL SHEET OF COMPARATIVE STUDY OF TWO TECHNIQUES CONTROL OF DNA FROM YEAST ISOLATES PRODUCING PECTINASE

2020 ◽  
Vol 8 (9) ◽  
pp. 979-983
Author(s):  
Coulibaly Wahauwouele Hermann ◽  
◽  
Bouatenin Koffi Maizan Jean-Paul ◽  
Kouame Kohi Alfred ◽  
Amoikon Tiemele Laurent Simon ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Molecular Biology ◽  
Spectrophotometric Method ◽  
Gel Electrophoresis ◽  
Extraction Method ◽  
Agarose Gel ◽  
Agarose Gel Electrophoresis ◽  
Chain Reaction ◽  
Polymerase Chain

Polymerase Chain Reaction (PCR) is a widely used technique in the field of molecular biology to rapidly make very large number of copies of a specific DNA sample for detailed studies. The success of the technique however is dependent on the on the quality, i.e purity of the extracted DNA specimen. The aim of this study was to evaluate the quality of extracted DNA from pectinase producing yeast to determine the suitability of the extraction method to produce pure extract without non-inhibiting substances.In this study, DNA extracts from six (06) isolates of pectinase-producing yeasts were quantitatively and qualitatively analyzed using NanoDrop spectrophotometry and agarose gel electrophoresis methods. These analyses showed that the concentration of DNA extracts from the isolates evaluated by the NanoDrop spectrophotometric method ranged from 403.8 to 1082.4 ng/µL and the purity index A 260/280 was between 2.03 and 2.11. In sum, agarose gel electrophoresis showed that the intensity of the DNA bands was irregular and not necessarily in line with the data provided by the NanoDrop spectrophotometry.

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