scholarly journals Double Digestion of Insert DNA v1 (protocols.io.76qhrdw)

protocols.io ◽  
2019 ◽  
Author(s):  
iGEM Dusseldorf
Keyword(s):  
Double Digestion

scholarly journals An approach using ddRADseq and machine learning for understanding speciation in Antarctic Antarctophilinidae gastropods

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Juan Moles ◽  
Shahan Derkarabetian ◽  
Stefano Schiaparelli ◽  
Michael Schrödl ◽  
Jesús S. Troncoso ◽  
...  
Keyword(s):  
Machine Learning ◽  
Deep Sea ◽  
Weddell Sea ◽  
Nucleotide Polymorphisms ◽  
Sea Levels ◽  
Suitable Material ◽  
Genetic Lineages ◽  
Genome Wide ◽  
Gastropod Mollusks ◽  
Double Digestion

AbstractSampling impediments and paucity of suitable material for molecular analyses have precluded the study of speciation and radiation of deep-sea species in Antarctica. We analyzed barcodes together with genome-wide single nucleotide polymorphisms obtained from double digestion restriction site-associated DNA sequencing (ddRADseq) for species in the family Antarctophilinidae. We also reevaluated the fossil record associated with this taxon to provide further insights into the origin of the group. Novel approaches to identify distinctive genetic lineages, including unsupervised machine learning variational autoencoder plots, were used to establish species hypothesis frameworks. In this sense, three undescribed species and a complex of cryptic species were identified, suggesting allopatric speciation connected to geographic or bathymetric isolation. We further observed that the shallow waters around the Scotia Arc and on the continental shelf in the Weddell Sea present high endemism and diversity. In contrast, likely due to the glacial pressure during the Cenozoic, a deep-sea group with fewer species emerged expanding over great areas in the South-Atlantic Antarctic Ridge. Our study agrees on how diachronic paleoclimatic and current environmental factors shaped Antarctic communities both at the shallow and deep-sea levels, promoting Antarctica as the center of origin for numerous taxa such as gastropod mollusks.

scholarly journals De Novo SNP Discovery and Genotyping of Iranian Pimpinella Species Using ddRAD Sequencing

Agronomy ◽  
2021 ◽  
Vol 11 (7) ◽  
pp. 1342
Author(s):  
Shaghayegh Mehravi ◽  
Gholam Ali Ranjbar ◽  
Ghader Mirzaghaderi ◽  
Anita Alice Severn-Ellis ◽  
Armin Scheben ◽  
...  
Keyword(s):  
De Novo ◽  
Genetic Relationships ◽  
Nucleotide Polymorphisms ◽  
High Quality ◽  
Genomic Resources ◽  
High Quality Snps ◽  
The Family ◽  
Double Digestion ◽  
Flanking Sequences ◽  
Downstream Analysis

The species of Pimpinella, one of the largest genera of the family Apiaceae, are traditionally cultivated for medicinal purposes. In this study, high-throughput double digest restriction-site associated DNA sequencing technology (ddRAD-seq) was used to identify single nucleotide polymorphisms (SNPs) in eight Pimpinella species from Iran. After double-digestion with the enzymes HpyCH4IV and HinfI, a total of 334,702,966 paired-end reads were de novo assembled into 1,270,791 loci with an average of 28.8 reads per locus. After stringent filtering, 2440 high-quality SNPs were identified for downstream analysis. Analysis of genetic relationships and population structure, based on these retained SNPs, indicated the presence of three major groups. Gene ontology and pathway analysis were determined by using comparison SNP-associated flanking sequences with a public non-redundant database. Due to the lack of genomic resources in this genus, our present study is the first report to provide high-quality SNPs in Pimpinella based on a de novo analysis pipeline using ddRAD-seq. This data will enhance the molecular knowledge of the genus Pimpinella and will provide an important source of information for breeders and the research community to enhance breeding programs and support the management of Pimpinella genomic resources.

Restriction Enzyme Double Digestion v1

2021 ◽  
Author(s):  
Zhenfeng Fu
Keyword(s):  
Restriction Enzyme ◽  
Enzyme Digestion ◽  
Dna Molecule ◽  
Double Digestion ◽  
Carrier Molecule ◽  
Sticky End

Enzyme digestion is to cut the DNA molecule and the carrier molecule at the sticky end to obtain the corresponding sticky end connection.

scholarly journals Methylation and rearrangement of mouse intracisternal a particle genes in development, aging, and myeloma.

1983 ◽  
Vol 3 (8) ◽  
pp. 1371-1380 ◽  
Author(s):  
L L Mays-Hoopes ◽  
A Brown ◽  
R C Huang
Keyword(s):  
Mouse Liver ◽  
Adult Life ◽  
Mouse Strains ◽  
Adult Liver ◽  
Myeloma Cells ◽  
Adult Mice ◽  
Mouse Tissues ◽  
Double Digestion ◽  
Genomic Dnas ◽  
Intracisternal A Particle

Sequences of DNA that hybridize on Southern blots with cloned intracisternal A-particle (IAP) sequences have been examined in genomic DNAs of neonatal mice, livers of adult mice (3, 6, 12, 18, 24, and 26 months old), and the solid myeloma tumor MOPC-315. The isoschizomers HpaII (CCGG or mCCGG) and MspI (CCGG or CmCGG) were used to assess methylation. All the DNAs produced a major 0.5-kilobase MspI fragment that hybridizes with IAP probe. Only the myeloma DNA, and to a much lesser degree DNA from senescent mouse liver, produced this fragment in HpaII digest; the other DNAs all had IAP sequences resistant to HpaII digestion. These sequences thus become fully methylated to CmCGG early and remain so in adult life, except in the myeloma cells that are expressing the IAP genes. An increase in MspI-sensitive sites in IAP gene-containing DNA was observed in aging mice. The probe used to assess methylation, a 0.8-kilobase fragment produced by BamHI-HindIII double digestion, is common to several cloned IAP genes and is part of a region of DNA which is conserved in genomes of all mouse tissues. The probe hybridized to 1.5- and 1.4-kilobase doublet bands produced by BamHI, HindIII, and EcoRI triple digestions of neonatal DNA. These two bands were found in neonatal livers of Swiss Webster, BALB/c, and C57BL/6J mouse strains, showed less in adult liver, and were barely detectable in senescent livers from C57BL/6J mice.

scholarly journals Differentiation of Pathogenic BartonellaSpecies by Infrequent Restriction Site PCR

2000 ◽  
Vol 38 (8) ◽  
pp. 3010-3015 ◽  
Author(s):  
Scott A. Handley ◽  
Russell L. Regnery
Keyword(s):  
Capillary Electrophoresis ◽  
Restriction Endonuclease ◽  
Restriction Site ◽  
Polyacrylamide Gel Electrophoresis ◽  
Ease Of Use ◽  
Dna Templates ◽  
Pattern Complexity ◽  
Double Digestion ◽  
Pcr Method ◽  
High Level

Infrequent restriction site PCR (IRS-PCR) is a recently described DNA fingerprinting technique based on selective amplification of restriction endonuclease-cleaved fragments. Bartonellaisolates associated with human disease and related nonhuman isolates were analyzed by IRS-PCR genomic fingerprinting. Preparation of DNA templates began with double digestion using three different restriction endonuclease combinations. Combinations included the frequently cutting endonuclease HhaI in conjunction with an infrequently cutting endonuclease, EagI, SmaI, orXbaI. Digestion was followed by ligation of oligonucleotide adapters designed with ends complementary to the restriction endonuclease sites. Amplification of fragments flanked with anEagI, SmaI, or XbaI site in combination with an HhaI site produced a series of different-sized amplicons resolvable into patterns by polyacrylamide gel electrophoresis (PAGE). The pattern complexity was varied by the addition of selective nucleotides to the 3′ ends of theEagI-, SmaI-, or XbaI-specific primers. Amplicons were also generated with fluorescently labeled primers and were subsequently resolved and detected by capillary electrophoresis. Analysis by traditional slab PAGE and capillary electrophoresis provided suitable resolution of patterns produced with the enzyme combinations EagI-HhaI andSmaI-HhaI. However, the combination ofXbaI-HhaI produced too many fragments for sufficient resolution by traditional PAGE, thus requiring the better resolving properties of capillary electrophoresis. Due to the flexibility in modulating the pattern complexity and electrophoresis methods, these techniques allow for a high level of experimental optimization. The results provide evidence of the discriminatory power, ease of use, and flexibility of the IRS-PCR method as it applies to the identification of human-pathogenic Bartonella species.

scholarly journals Principles and applications of Ligation Mediated PCR methods for DNA-based typing of microbial organisms.

2016 ◽  
Vol 63 (1) ◽  
Author(s):  
Beata Krawczyk ◽  
Józef Kur ◽  
Karolina Stojowska-Swędrzyńska ◽  
Marta Śpibida
Keyword(s):  
Pcr Amplification ◽  
Dna Amplification ◽  
Restriction Endonuclease Analysis ◽  
Site Amplification ◽  
Genomic Fingerprinting ◽  
Restriction Sites ◽  
Double Digestion ◽  
Pcr Method ◽  
Alternative Techniques ◽  
Endonuclease Analysis

A significant number of DNA-based techniques has been introduced into the field of microorganisms' characterization and taxonomy. These genomic fingerprinting methods were developed to detect DNA sequence polymorphisms by using general principles, such as restriction endonuclease analysis, molecular hybridization, and PCR amplification. In recent years, some alternative techniques based on ligation of oligonucleotide adapters before DNA amplification by PCR, known as Ligation-Mediated PCR methods (LM PCR), have been successfully applied for the typing of microorganisms below the species level. These molecular methods include: Amplified Fragment Length Polymorphism (AFLP), Amplification of DNA fragments Surrounding Rare Restriction Sites (ADSRRS), PCR Melting Profiles (PCR MP), Ligation Mediated PCR/Shifter (LM PCR/Shifter), Infrequent-Restriction-Site Amplification (IRS PCR), double digestion Ligation Mediated Suppression PCR (ddLMS PCR). These techniques are now applied more and more often because they involve less time, are comparably inexpensive, and require only standard lab equipment. Here, we present a general review of this group of methods showing their possibilities and limitations. We also identify questions and propose solutions which may be helpful in choosing a particular LM PCR method for the achievement of the required goal.

scholarly journals A CYTOCHEMICAL AND RADIOAUTOGRAPHIC STUDY OF HUMAN TISSUE CULTURE CELL NUCLEOLI

1970 ◽  
Vol 45 (3) ◽  
pp. 479-492 ◽  
Author(s):  
L. Recher ◽  
J. Whitescarver ◽  
L. Briggs
Keyword(s):  
Tissue Culture ◽  
Human Tissue ◽  
Culture Cell ◽  
Tissue Culture Cell ◽  
Active Chromatin ◽  
Pepsin Digestion ◽  
Basic Proteins ◽  
Double Digestion ◽  
Silver Grains ◽  
Structural Aspects

Fine structural aspects of human tissue culture cell nucleoli were studied by cytochemical and radioautographic methods. Ribonuclease and pepsin digestions were carried out on glutaraldehyde-fixed cells that, in some instances, were labeled with thymidine-3H prior to digestion. Double digestion by ribonuclease and pepsin revealed a fine fibrillar reticulum that appears to be the supportive structure of nucleolonemal threads. The nature of the reticulum remains to be determined. The question of whether it may represent a dispersed form of chromatin was raised. Structural findings suggested such an hypothesis but the results of radioautographic studies do not support it. The reticulum showed a striking absence of radioactive labeling following a 3 hr incorporation of thymidine-3H. Only few silver grains were observed occasionally in the fibrillar nucleolonema that may or may not be significant. The radioautographic results are believed to be inconclusive for the various reasons discussed. The possibility that the reticulum is composed of proteins has to be considered. It appears that basic proteins can resist pepsin digestion in aldehyde-fixed cells. Individual chromatin fibrils were found to be associated with the nucleolar reticulum. It is possible that these alone represent the dispersed genetically active chromatin of nucleoli.

scholarly journals Identification and Mapping of Eastern Filbert Blight Resistance Quantitative Trait Loci in European Hazelnut Using Double Digestion Restriction Site Associated DNA Sequencing

2019 ◽  
Vol 144 (5) ◽  
pp. 295-304 ◽  
Author(s):  
Josh A. Honig ◽  
Megan F. Muehlbauer ◽  
John M. Capik ◽  
Christine Kubik ◽  
Jennifer N. Vaiciunas ◽  
...  
Keyword(s):  
United States ◽  
Genetic Diversity ◽  
Quantitative Trait ◽  
Genetic Linkage ◽  
Genetic Linkage Map ◽  
Restriction Site ◽  
Eastern United States ◽  
Double Digestion ◽  
Disease Pressure ◽  
Resistance Source

European hazelnut (Corylus avellana L.) is an economically important edible nut producing species, which ranked sixth in world tree nut production in 2016. European hazelnut production in the United States is primarily limited to the Willamette Valley of Oregon, and currently nonexistent in the eastern United States because of the presence of a devastating endemic disease, eastern filbert blight (EFB) caused by Anisogramma anomala (Peck) E. Muller. The primary commercial means of control of EFB to date is through the development and planting of genetically resistant european hazelnut cultivars, with an R-gene introduced from the obsolete, late-shedding pollinizer ‘Gasaway’. Although the ‘Gasaway’ resistance source provides protection against EFB in the Pacific northwestern United States (PNW), recent reports have shown that it is not effective in parts of the eastern United States. This may be in part because the identification and selection of ‘Gasaway’ and ‘Gasaway’-derived cultivars occurred in an environment (PNW) with limited genetic diversity of A. anomala. The objectives of the current research were to develop a genetic linkage map using double digestion restriction site associated DNA sequencing (ddRADseq) and identify quantitative trait loci (QTL) markers associated with EFB resistance from the resistant selection Rutgers H3R07P25 from southern Russia. A mapping population composed of 119 seedling trees was evaluated in a geographic location (New Jersey) where the EFB fungus is endemic, exhibits high disease pressure, and has a high level of genetic diversity. The completed genetic linkage map included a total of 2217 markers and spanned a total genetic distance of 1383.4 cM, with an average marker spacing of 0.65 cM. A single QTL region associated with EFB resistance from H3R07P25 was located on european hazelnut linkage group (LG) 2 and was responsible for 72.8% of the phenotypic variation observed in the study. Based on its LG placement, origin, and disease response in the field, this resistance source is different from the ‘Gasaway’ source located on LG6. The current results, in combination with results from previous research, indicate that the H3R07P25 source is likely exhibiting resistance to a broader range of naturally occurring A. anomala isolates. As such, H3R07P25 will be important for the development of new european hazelnut germplasm that combines EFB resistance from multiple sources in a gene pyramiding approach. Identification of EFB resistance in high disease pressure environments representing a diversity of A. anomala populations is likely a requirement for identifying plants expressing durable EFB resistance, which is a precursor to the development of a commercially viable european hazelnut industry in the eastern United States.

Sign in / Sign up

Export Citation Format

Share Document