scholarly journals LAGAN and Multi-LAGAN: Efficient Tools for Large-Scale Multiple Alignment of Genomic DNA

2003 ◽  
Vol 13 (4) ◽  
pp. 721-731 ◽  
Author(s):  
M. Brudno
Keyword(s):  
Genomic Dna ◽  
Large Scale ◽  
Multiple Alignment

scholarly journals Homologous Recombination within Large Chromosomal Regions Facilitates Acquisition of β-Lactam and Vancomycin Resistance in Enterococcus faecium

2016 ◽  
Vol 60 (10) ◽  
pp. 5777-5786 ◽  
Author(s):  
Mónica García-Solache ◽  
Francois Lebreton ◽  
Robert E. McLaughlin ◽  
James D. Whiteaker ◽  
Michael S. Gilmore ◽  
...  
Keyword(s):  
Enterococcus Faecium ◽  
Genomic Dna ◽  
Large Scale ◽  
Vancomycin Resistance ◽  
Whole Genome ◽  
Whole Genome Analysis ◽  
Single Nucleotide ◽  
Content Type ◽  
Donor Chromosome ◽  
Vancomycin Resistant

ABSTRACTThe transfer of DNA betweenEnterococcus faeciumstrains has been characterized both by the movement of well-defined genetic elements and by the large-scale transfer of genomic DNA fragments. In this work, we report on the whole-genome analysis of transconjugants resulting from mating events between the vancomycin-resistantE. faeciumC68 strain and the vancomycin-susceptible D344RRF strain to discern the mechanism by which the transferred regions enter the recipient chromosome. Vancomycin-resistant transconjugants from five independent matings were analyzed by whole-genome sequencing. In all cases but one, the penicillin binding protein 5 (pbp5) gene and the Tn5382vancomycin resistance transposon were transferred together and replaced the correspondingpbp5region of D344RRF. In one instance, Tn5382inserted independently downstream of the D344RRFpbp5gene. Single nucleotide variant (SNV) analysis suggested that entry of donor DNA into the recipient chromosome occurred by recombination across regions of homology between donor and recipient chromosomes, rather than through insertion sequence-mediated transposition. The transfer of genomic DNA was also associated with the transfer of C68 plasmid pLRM23 and another putative plasmid. Our data are consistent with the initiation of transfer by cointegration of a transferable plasmid with the donor chromosome, with subsequent circularization of the plasmid-chromosome cointegrant in the donor prior to transfer. Entry into the recipient chromosome most commonly occurred across regions of homology between donor and recipient chromosomes.

Multiple Displacement Amplification Enables Large Scale Clonal Analysis after Retroviral Gene Therapy.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 5469-5469
Author(s):  
Stephanie Bleier ◽  
Patrick Maier ◽  
Frederik Wenz ◽  
W. Jens Zeller ◽  
Stephanie Laufs ◽  
...  
Keyword(s):  
Gene Therapy ◽  
Real Time ◽  
Real Time Pcr ◽  
Genomic Dna ◽  
Large Scale ◽  
Integration Site ◽  
Multiple Displacement Amplification ◽  
Quantitative Real Time Pcr ◽  
Integration Site Analysis ◽  
Reliable Quantification

Abstract Analysis of the fate of retrovirally transduced cells after transplantation is often hampered by the scarcity of available DNA. We evaluated a promising method for whole genome amplification named multiple displacement amplification (MDA) with respect to the even and accurate representation of retrovirally transduced genomic DNA. We were able to show that MDA is a suitable method to subsequently specify engraftment efficiencies by quantitative real-time PCR as the retroviral integrations are amplified the same way and by the same probability as all other parts of the genome. We validated the method by analyzing a dilution series containing retrovirally transduced DNA and untransduced background DNA and retroviral integrations found in primary material from a retroviral transplantation model by quantitative real-time PCR. The representation of the portion of retroviral DNA in the amplified samples was 0.9-fold (range 0.2 – 2.1-fold) of the portion determined in the original genomic DNA. Furthermore, the succession of the combination of MDA and integration site analysis by ligation-mediated PCR showed an increase in the sensitivity of the method as a specific integration site could be detected in a background of untransduced DNA, while the transduced DNA made up only 0.001%. These results show that MDA enables large scale sensitive detection and reliable quantification of retrovirally transduced human genomic DNA and therefore facilitates follow up analysis in gene therapy studies even from smallest amounts of starting material.

scholarly journals Comparison of Eleven Methods for Genomic DNA Extraction Suitable for Large-Scale Whole-Genome Genotyping and Long-Term DNA Banking Using Blood Samples

PLoS ONE ◽  
2015 ◽  
Vol 10 (1) ◽  
pp. e0115960 ◽  
Author(s):  
Androniki Psifidi ◽  
Chrysostomos I. Dovas ◽  
Georgios Bramis ◽  
Thomai Lazou ◽  
Claire L. Russel ◽  
...  
Keyword(s):  
Dna Extraction ◽  
Genomic Dna ◽  
Large Scale ◽  
Whole Genome ◽  
Blood Samples ◽  
Genomic Dna Extraction ◽  
Dna Banking

scholarly journals Optimization of extraction of genomic DNA from archived dried blood spot (DBS): potential application in epidemiological research & bio banking

2018 ◽  
Vol 2 ◽  
pp. 57 ◽  
Author(s):  
Abhinendra Kumar ◽  
Sharayu Mhatre ◽  
Sheela Godbole ◽  
Prabhat Jha ◽  
Rajesh Dikshit
Keyword(s):  
Genomic Dna ◽  
Large Scale ◽  
Venous Blood ◽  
Magnetic Bead ◽  
Epidemiological Studies ◽  
Dried Blood Spot ◽  
Future Research ◽  
Prolonged Storage ◽  
Whole Genome ◽  
Blood Spot

Background: Limited infrastructure is available to collect, store and transport venous blood in field epidemiological studies. Dried blood spot (DBS) is a robust potential alternative sample source for epidemiological studies & bio banking. A stable source of genomic DNA (gDNA) is required for long term storage in bio bank for its downstream applications. Our objective is to optimize the methods of gDNA extraction from stored DBS and with the aim of revealing its utility in large scale epidemiological studies.  Methods: The purpose of this study was to extract the maximum amount of gDNA from DBS on Whatman 903 protein saver card. gDNA was extracted through column  (Qiagen) & magnetic bead based (Invitrogen) methods. Quantification of extracted gDNA was performed with a spectrophotometer, fluorometer, and integrity analyzed by agarose gel electrophoresis.  Result: Large variation was observed in quantity & purity (260/280 ratio, 1.8-2.9) of the extracted gDNA. The intact gDNA bands on the electrophoresis gel reflect the robustness of DBS for gDNA even after prolonged storage time. The extracted gDNA amount 2.16 – 24 ng/µl is sufficient for its PCR based downstream application, but unfortunately it can’t be used for whole genome sequencing or genotyping from extracted gDNA. Sequencing or genotyping can be achieved by after increasing template copy number through whole genome amplification of extracted gDNA. The obtained results create a base for future research to develop high-throughput research and extraction methods from blood samples. Conclusion: The above results reveal, DBS can be utilized as a potential and robust sample source for bio banking in field epidemiological studies.

scholarly journals Fast and inexpensive whole genome sequencing library preparation from intact yeast cells

2020 ◽  
Author(s):  
Sibylle C Vonesch ◽  
Shengdi Li ◽  
Chelsea Szu Tu ◽  
Bianca P Hennig ◽  
Nikolay Dobrev ◽  
...  
Keyword(s):  
Whole Genome Sequencing ◽  
Genome Sequencing ◽  
Genomic Dna ◽  
Large Scale ◽  
Massively Parallel Sequencing ◽  
Yeast Cells ◽  
Whole Genome ◽  
High Quality ◽  
Rapid Preparation ◽  
Yeast Cultures

ABSTRACTThrough the increase in the capacity of sequencing machines massively parallel sequencing of thousands of samples in a single run is now possible. With the improved throughput and resulting drop in the price of sequencing, the cost and time for preparation of sequencing libraries have become the major bottleneck in large-scale experiments. Methods using a hyperactive variant of the Tn5 transposase efficiently generate libraries starting from cDNA or genomic DNA in a few hours and are highly scalable. For genome sequencing, however, the time and effort spent on genomic DNA isolation limits the practicability of sequencing large numbers of samples. Here, we describe a highly scalable method for preparing high quality whole-genome sequencing libraries directly from yeast cultures in less than three hours at 34 cents per sample. We skip the rate-limiting step of genomic DNA extraction by directly tagmenting yeast spheroplasts and add a nucleosome release step prior to enrichment PCR to improve the evenness of genomic coverage. Resulting libraries do not show any GC-bias and are comparable in quality to libraries processed from genomic DNA with a commercially available Tn5-based kit. We use our protocol to investigate CRISPR/Cas9 on- and off-target edits and reliably detect edited variants and shared polymorphisms between strains. Our protocol enables rapid preparation of unbiased and high-quality, sequencing-ready indexed libraries for hundreds of yeast strains in a single day at a low price. By adjusting individual steps of our workflow we expect that our protocol can be adapted to other organisms.

scholarly journals Large-scale screening of intracellular protein localization in living fission yeast cells by the use of a GFP-fusion genomic DNA library

2000 ◽  
Vol 5 (3) ◽  
pp. 169-190 ◽  
Author(s):  
Da-Qiao Ding ◽  
Yuki Tomita ◽  
Ayumu Yamamoto ◽  
Yuji Chikashige ◽  
Tokuko Haraguchi ◽  
...  
Keyword(s):  
Fission Yeast ◽  
Genomic Dna ◽  
Large Scale ◽  
Protein Localization ◽  
Yeast Cells ◽  
Intracellular Protein ◽  
Dna Library ◽  
Genomic Dna Library ◽  
Large Scale Screening

scholarly journals EGassembler: online bioinformatics service for large-scale processing, clustering and assembling ESTs and genomic DNA fragments

2006 ◽  
Vol 34 (Web Server) ◽  
pp. W459-W462 ◽  
Author(s):  
A. Masoudi-Nejad ◽  
K. Tonomura ◽  
S. Kawashima ◽  
Y. Moriya ◽  
M. Suzuki ◽  
...  
Keyword(s):  
Genomic Dna ◽  
Large Scale ◽  
Dna Fragments

Development of multiplex PCR assay for concurrent detection of tick borne haemoparasitic infections in bovines

2018 ◽  
Vol 63 (4) ◽  
pp. 759-765 ◽  
Author(s):  
V.R. Kundave ◽  
Hira Ram ◽  
Partha S. Banerjee ◽  
Rajat Garg ◽  
K. Mahendran ◽  
...  
Keyword(s):  
Multiplex Pcr ◽  
Genomic Dna ◽  
Large Scale ◽  
Simultaneous Detection ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Individual Species ◽  
Pcr Assay ◽  
Blood Samples ◽  
Multiplex Pcr Assay

Abstract This study describes development and evaluation of a multiplex PCR assay for simultaneous detection of Theileria annulata, Babesia bigemina and Anaplasma marginale infections in bovines. The assay was developed using parasites specific genomic DNA and three sets of PCR primers targeting the Tams1, 18S rRNA and 16S rRNA genes of T. annulata, B. bigemina and A. marginale, respectively. Blood samples collected from a total of 461 bovines, suspected for haemoparasitic infections, were examined microscopically to record the status of infection and simultaneously, genomic DNA extracted from these blood samples were utilized for the optimization and validation of multiplex PCR assay. Microscopic examination of blood samples revealed presence of single and multiple species of haemoparasites in 25.8% and 2.4% samples, respectively. Results of multiplex PCR revealed the presence of single haemoparasitic species infection in 159 cases (34.5%), whereas mixed infection was recorded in 82 (17.8%) samples. Occurrence of individual species infection detected by mPCR in the study was 26.03% (120/461) for T. annulata, 3.25% (15/461) for B. bigemina and 5.20% (24/461) for A. marginale. The detection limit of multiplex PCR assay was at the template dilutions of 10−6, 10−6 and 10−4, which corresponded to 0.1 pg, 0.1 pg and 10.0 pg of DNA for T. annulata, A. marginale, and B. bigemina, respectively. Based on the high diagnostic sensitivity and throughput, multiplex PCR assay developed in the present study could be exploited as a tool to conduct large-scale epidemiological survey for tick-borne haemoparasitic infection of bovines.

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