DNA extraction protocols

Bedbugs are a very recurrent pest all over the world which is often difficult to eradicate. This is the case for Cimex lectularius, a species whose bites can be manifested in a wide variety of ways. Thus, it seems necessary to develop a mechanism that confirms the presence of this insect in order to eliminate the pest as soon as possible. For this reason, the Basque Institute of Forensic Sciences in Sanfran (BIFSS) has been working on this research field and has developed 12 new autosomal STR markers that allow the identification of the Cimicidae family of insects.

A goal of breeding programs for resistance to white pine blister rust is the development of multigenic resistance, even if the genetics and mechanisms of resistance may be imperfectly understood. The goal of multigenic resistance has prompted efforts to categorize host resistance reactions at increasingly finer scales, to identify heritable traits that may confer quantitative resistance. PCR amplification of Cronartium ribicola DNA presents a sensitive and highly specific method for detection of C. ribicola in host tissues, and is well suited to screening of large numbers of samples for which other methods of pathogen detection (e.g., microscopy) may be unsuitable. PCR amplification can be used to detect presence of the pathogen in different host tissues, and so can provide useful information on putatative resistance responses that may be localized in specific tissue types.

Cyprus traditional sausages from the Troodos mountainous region of Pitsilia gained the protected geographical indication (PGI) designation from the European Committee (EU 2020/C 203/06). Still, we lack authentication protocols for the distinction of "Pitsilia" from industrially produced Cyprus sausages. Microbial activity is an essential contributor to traditional sausages' sensorial characteristics, but whether the microbial patterns might be associated with the area of production is unclear. In the present research, we applied high-throughput sequencing (HTS) to provide a linkage between the area of production and Cyprus sausages' bacterial diversity. To strengthen our findings, we used three different DNA extraction commercial kits: (i) the DNeasy PowerFood Microbial Kit (QIAGEN); (ii) the NucleoSpin Food Kit (MACHEREY-NAGEL); and (iii) the blackPREP Food DNA I Kit (Analytik Jena), in which we applied three different microbial cell wall lysis modifications. The modifications included heat treatment, bead beating, and enzymatic treatment. Results regarding metagenomic sequencing were evaluated in terms of number of reads, alpha diversity indexes, and taxonomic composition. The efficacy of each method of DNA isolation was assessed quantitatively based on the extracted DNA yield and the obtained copy number of (a) the 16S rRNA gene, (b) the internal transcribed spacer (ITS) region, and (c) three Gram-positive bacteria that belong to the genera Latilactobacillus (formerly Lactobacillus), Bacillus, and Enterococcus via absolute quantification using qPCR. Compared with some examined industrial sausages, Pitsilia sausages had significantly higher bacterial alpha diversity (Shannon and Simpson indexes). Principal coordinates analysis separated the total bacterial community composition (beta diversity) of the three Pitsilia sausages from the industrial sausages, with the exception of one industrial sausage produced in Pitsilia, according

Cyprus traditional sausages from the Troodos mountainous region of Pitsilia gained the protected geographical indication (PGI) designation from the European Committee (EU 2020/C 203/06). Still, we lack authentication protocols for the distinction of “Pitsilia” from industrially produced Cyprus sausages. Microbial activity is an essential contributor to traditional sausages’ sensorial characteristics, but whether the microbial patterns might be associated with the area of production is unclear. In the present research, we applied high-throughput sequencing (HTS) to provide a linkage between the area of production and Cyprus sausages’ bacterial diversity. To strengthen our findings, we used three different DNA extraction commercial kits: (i) the DNeasy PowerFood Microbial Kit (QIAGEN); (ii) the NucleoSpin Food Kit (MACHEREY-NAGEL); and (iii) the blackPREP Food DNA I Kit (Analytik Jena), in which we applied three different microbial cell wall lysis modifications. The modifications incl...

The potential for co-infection with COVID-19 and other respiratory infections raises the possibility that a new severe acute respiratorysyndrome coronavirus 2 (SARS-CoV-2) mimics the influenza A virus regarding methods and modes of transmission, clinical features, relatedimmune responses, and seasonal coincidence. This study aimed to investigate the presence of SARS-CoV-2 and influenza A virus coinfections inadmitted patients of a tertiary care hospital. In this study, the total included 589 admitted patients in our tertiary care hospital. The detectionof co-infection between SARS-CoV-2 and influenza A virus by real-time reverse-transcriptase polymerase chain reaction (rRT-PCR) in a patientduring the second wave of the COVID-19 pandemic. There were 207 (35.1%) patients infected with SARS-CoV-2 and 43 (7.3%) patientsinfected with the influenza A virus. Only 6 (1.0%) patients were infected with SARS-CoV-2 and influenza A viruses. The females were morelikely to be infected with SARS-CoV-2 than non-infected SARS-CoV-2 case-patients (60.9% (n = 126) vs. 31.4% (n = 120), and also with theinfluenza A virus compared with influenza-negative patients (40.9% (n = 223) vs. 55.9% (n = 24). In conclusion, our results strongly suggestthat influenza A co-infects with SARS-CoV-2. The patients with SARS-CoV-2 and influenza A co-infection had similar clinical characteristics asthose with SARS-CoV-2 infection alone. Comorbidities, like hypertension and diabetes, and increasing age make patients more susceptible toSARS-CoV-2 and influenza A coinfections

(PDF) The Demographic and Clinical Characteristics of the Co-infection for the Detection of SARS- COV-2 and Influenza A Virus by rRT-PCR. Available from: https://www.researchgate.net/publication/378076755_The_Demographic_and_Clinical_Characteristics_of_the_Co-infection_for_the_Detection_of_SARS-_COV-2_and_Influenza_A_Virus_by_rRT-PCR [accessed Sep 04 2024].

To explore the natural microbial community of any ecosystems by high-resolution molecular approaches including next generation sequencing, it is extremely important to develop a sensitive and reproducible DNA extraction method that facilitate isolation of microbial DNA of sufficient purity and quantity from culturable and uncultured microbial species living in that environment. Proper lysis of heterogeneous community microbial cells without damaging their genomes is a major challenge. In this study, we have developed an improved method for extraction of community DNA from different environmental and human origin samples. We introduced a combination of physical, chemical and mechanical lysis methods for proper lysis of microbial inhabitants. The community microbial DNA was precipitated by using salt and organic solvent. Both the quality and quantity of isolated DNA was compared with the existing methodologies and the supremacy of our method was confirmed. Maximum recovery of genomic ...

Introduction: The quality of the extracted genomic DNA is largely reduced in patients who are exposed to chemotherapeutic treatment, which is usually encountered in the commonly used DNA extraction methods that are not designed to isolate DNA from those patients. In this study, a special non-organic protocol was developed to overcome the negative effects of chemotherapeutic drugs on the quality of extracted DNA. Materials and Methods: Numerous modulations were applied in the washing, suspension, and lysis approaches to compensate for the harmful impacts of the chemotherapeutic drugs on the quality of genomic DNA. Obvious purity and adequate yields of the extracted DNA were demonstrated in the suggested protocol. The validity of this protocol for digestion with restriction endonucleases, conventio nal PCR, and realtime PCR was confirmed. Results: This protocol proved satisfactory values of absorbance ratio (1.8 ± 0.02 and 2.1 ± 1.2, for A260/280 and A260/230, respectively) and adequate yields of DNA (10 ± 2.24) µg/100 µl. The validation experiments proved the efficacy of extracted DNA for downstream applications of molecular biology. Conclusions: The utilization of this method entails a useful approach for extracting molecular biology-grade DNA without having inhibitors against common enzymes used in molecular biology even after exposing patients to several sessions of chemotherapy .

Here, we describe a rapid, efficient and noncomplicated method for extracting poultry genomic DNA. This simple method obtains excellent qualities of molecular biology grade DNA in a matter of only about 15 min. A straightforward protocol is followed in this extraction procedure, in which, when the blood cells are placed in a distilled water alone, water rapidly disrupts red blood cells (RBCs) membranes and alleviate chickens' undesired high blood viscosity without being aided by any other chemicals. Moreover, the time, cost and efforts were hugely reduced since the step of leukocytes lysis was successfully mixed with the protein precipitation. The isolated genomic DNA, in terms of its quantity and quality, is satisfying and is proved to be suitable for restriction endonucleases digestion, polymerase chain reaction (PCR), and restriction fragment length polymorphism (RFLP). Thus, instead of extracting a limited amount of DNA using expensive or relatively expensive kits, this guaranteed procedure can be utilized alternatively to accurately extract genomic DNA (gDNA) from minimal starter quantities of the chicken's blood that do not exceed 50 µL. By relying on this technique, all the practical handover steps are reduced to the extreme limits. This pilot study may provide a high yield gDNA extraction that minimizes labour, expense, and steps in very high competency and reproducibility.

Introduction: The identification of perpetrator involved in crime becomes difficult due to lack of evidence. Biological evidence plays an integral role in establishing link between survivor, suspect, and the scene of crime. The era of eye witness is almost diminished and DNA analysis is necessary for the identification of perpetrator. The loss or degradation of biological traces due to urinating, defecating, douching of genitals, showering and delay in medico-legal examination may yield negative results.
Objective: The objective of study was to link DNA profiles generated from biological samples to associate suspect with survivor and other parameters affecting the results of DNA profiling.
Materials and Methods: This present study was conducted in the Biology & Serology Division of Regional Forensic Science Laboratory, Northern Range, Dharamshala and DNA Division of State Forensic Science Laboratory, Himachal Pradesh, Junga, India. A total of 142 sexual assault cases received for examination during the year 2018 and 2019 were studied.
Results: Human spermatozoa were detected in 39 (27.46%) cases and genetic profiles were generated. Spermatozoa were detected even after taking bath and washing of clothes in 3 (2.11%) cases. 10 (7.04%) survivors were menstruating at the time of assault. Hymen was absent in 43 (30.28%) cases followed by old healed tags in 37 (26.06%), recent tears in 15 (10.56%), intact in 9 (6.34%) and partially ruptured in 4 (2.82%) cases. Out of 39 cases, DNA profile resulted as an inclusion of assailant in 27 (19.01%) cases and exclusion in 12 (8.45%) cases. The persistence of spermatozoa on vaginal swabs was found up to 4 days. Only 18 (12.68%) survivors underwent medico-legal examination on first day of assault.