scholarly journals Would an RRS by any other name sound as RAD?

2018 ◽  
Author(s):  
Erin O Campbell ◽  
Bryan M T Brunet ◽  
Julian R Dupuis ◽  
Felix A H Sperling
Keyword(s):  
Literature Review ◽  
Restriction Enzyme ◽  
Restriction Enzymes ◽  
Shotgun Sequencing ◽  
Sequencing Technology ◽  
Reduced Representation ◽  
Minor Elements ◽  
Scientific Innovation ◽  
A Genome ◽  
Unique Method

ABSTRACTSampling markers throughout a genome with restriction enzymes emerged in the 2000s as reduced representation shotgun sequencing (RRS). Rapid advances in sequencing technology have since spurred modifications of RRS, giving rise to many derivatives with unique names, such as RADseq. But naming conventions have often been more creative than consistent, with unclear criteria for recognition as a unique method resulting in a proliferation of names characterized by ambiguity. We conducted a literature review to assess methodological and etymological relationships among 36 restriction enzyme-based methods, as well as rates of correct referencing of commonly-used methods. We identify several instances of methodological convergence or misattribution in the literature, and note that many published derivatives have modified only minor elements of parent protocols. We urge greater restraint in naming derivative methods, to strike a better balance between clarity, recognition of scientific innovation, and correct attribution.

scholarly journals DNA methylation estimation using methylation-sensitive restriction enzyme bisulfite sequencing (MREBS)

2017 ◽  
Author(s):  
Giancarlo Bonora ◽  
Liudmilla Rubbi ◽  
Marco Morselli ◽  
Constantinos Chronis ◽  
Kathrin Plath ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Restriction Enzyme ◽  
Large Scale ◽  
Bisulfite Sequencing ◽  
Multiple Regression Model ◽  
Reduced Representation ◽  
A Genome ◽  
Genome Bisulfite Sequencing ◽  
Methylation Sensitive Restriction Enzyme ◽  
Wide Scale

ABSTRACTWhole-genome bisulfite sequencing (WGBS) and reduced representation bisulfite sequencing (RRBS) are widely used for measuring DNA methylation levels on a genome-wide scale(1). Both methods have limitations: WGBS is expensive and prohibitive for most large-scale projects; RRBS only interrogates 6-12% of the CpGs in the human genome(16,19). Here, we introduce methylation-sensitive restriction enzyme bisulfite sequencing (MREBS) which has the reduced sequencing requirements of RRBS, but significantly expands the coverage of CpG sites in the genome. We built a multiple regression model that combines the two features of MREBS: the bisulfite conversion ratios of single cytosines (as in WGBS and RRBS) as well as the number of reads that cover each locus (as in MRE-seq(12)). This combined approach allowed us to estimate differential methylation across 60% of the genome using read count data alone, and where counts were sufficiently high in both samples (about 1.5% of the genome), our estimates were significantly improved by the single CpG conversion information. We show that differential DNA methylation values based on MREBS data correlate well with those based on WGBS and RRBS. This newly developed technique combines the sequencing cost of RRBS and DNA methylation estimates on a portion of the genome similar to WGBS, making it ideal for large-scale projects of mammalian genomes.

scholarly journals Predicting genome sizes and restriction enzyme recognition-sequence probabilities across the eukaryotic tree of life

2014 ◽  
Author(s):  
Santiago Herrera ◽  
Paula H. Reyes-Herrera ◽  
Timothy M. Shank
Keyword(s):  
Restriction Enzyme ◽  
High Throughput Sequencing ◽  
Sequence Data ◽  
Restriction Enzymes ◽  
Tree Of Life ◽  
Design Element ◽  
Recognition Sequence ◽  
Reduced Representation ◽  
Common Strategy ◽  
Taxonomic Groups

High-throughput sequencing of reduced representation libraries obtained through digestion with restriction enzymes ? generically known as restriction-site associated DNA sequencing (RAD-seq) ? is a common strategy to generate genome-wide genotypic and sequence data from eukaryotes. A critical design element of any RAD-seq study is a knowledge of the approximate number of genetic markers that can be obtained for a taxon using different restriction enzymes, as this number determines the scope of a project, and ultimately defines its success. This number can only be directly determined if a reference genome sequence is available, or it can be estimated if the genome size and restriction recognition sequence probabilities are known. However, both scenarios are uncommon for non-model species. Here, we performed systematic in silico surveys of recognition sequences, for diverse and commonly used type II restriction enzymes across the eukaryotic tree of life. Our observations reveal that recognition-sequence frequencies for a given restriction enzyme are strikingly variable among broad eukaryotic taxonomic groups, being largely determined by phylogenetic relatedness. We demonstrate that genome sizes can be predicted from cleavage frequency data obtained with restriction enzymes targeting ?neutral? elements. Models based on genomic compositions are also effective tools to accurately calculate probabilities of recognition sequences across taxa, and can be applied to species for which reduced-representation data is available (including transcriptomes and ?neutral? RAD-seq datasets). The analytical pipeline developed in this study, PredRAD (https://github.com/phrh/PredRAD), and the resulting databases constitute valuable resources that will help guide the design of any study using RAD-seq or related methods.

scholarly journals HapSolo: an optimization approach for removing secondary haplotigs during diploid genome assembly and scaffolding

2021 ◽  
Vol 22 (1) ◽  
Author(s):  
Edwin A. Solares ◽  
Yuan Tao ◽  
Anthony D. Long ◽  
Brandon S. Gaut
Keyword(s):  
Cost Function ◽  
Anopheles Funestus ◽  
Hill Climbing ◽  
Optimization Approach ◽  
Sequencing Technology ◽  
Genome Data ◽  
A Genome ◽  
Long Read ◽  
Downstream Analysis ◽  
The Cost

Abstract Background Despite marked recent improvements in long-read sequencing technology, the assembly of diploid genomes remains a difficult task. A major obstacle is distinguishing between alternative contigs that represent highly heterozygous regions. If primary and secondary contigs are not properly identified, the primary assembly will overrepresent both the size and complexity of the genome, which complicates downstream analysis such as scaffolding. Results Here we illustrate a new method, which we call HapSolo, that identifies secondary contigs and defines a primary assembly based on multiple pairwise contig alignment metrics. HapSolo evaluates candidate primary assemblies using BUSCO scores and then distinguishes among candidate assemblies using a cost function. The cost function can be defined by the user but by default considers the number of missing, duplicated and single BUSCO genes within the assembly. HapSolo performs hill climbing to minimize cost over thousands of candidate assemblies. We illustrate the performance of HapSolo on genome data from three species: the Chardonnay grape (Vitis vinifera), with a genome of 490 Mb, a mosquito (Anopheles funestus; 200 Mb) and the Thorny Skate (Amblyraja radiata; 2650 Mb). Conclusions HapSolo rapidly identified candidate assemblies that yield improvements in assembly metrics, including decreased genome size and improved N50 scores. Contig N50 scores improved by 35%, 9% and 9% for Chardonnay, mosquito and the thorny skate, respectively, relative to unreduced primary assemblies. The benefits of HapSolo were amplified by down-stream analyses, which we illustrated by scaffolding with Hi-C data. We found, for example, that prior to the application of HapSolo, only 52% of the Chardonnay genome was captured in the largest 19 scaffolds, corresponding to the number of chromosomes. After the application of HapSolo, this value increased to ~ 84%. The improvements for the mosquito’s largest three scaffolds, representing the number of chromosomes, were from 61 to 86%, and the improvement was even more pronounced for thorny skate. We compared the scaffolding results to assemblies that were based on PurgeDups for identifying secondary contigs, with generally superior results for HapSolo.

scholarly journals PCR amplification and sequence analyses of reverse transcriptase-like genes in Crinipellis perniciosa isolates

2007 ◽  
Vol 32 (5) ◽  
pp. 373-380 ◽  
Author(s):  
Jorge F. Pereira ◽  
Mariana D.C. Ignacchiti ◽  
Elza F. Araújo ◽  
Sérgio H. Brommonschenkel ◽  
Júlio C.M. Cascardo ◽  
...  
Keyword(s):  
Reverse Transcriptase ◽  
Pathogenic Fungus ◽  
Pcr Amplification ◽  
Restriction Enzymes ◽  
Sequence Comparisons ◽  
Copy Numbers ◽  
A Genome ◽  
Close Relationship ◽  
Pcr Products ◽  
Broom Disease

Reverse transcriptase (RT) sequence analysis is an important technique used to detect the presence of transposable elements in a genome. Putative RT sequences were analyzed in the genome of the pathogenic fungus C. perniciosa, the causal agent of witches' broom disease of cocoa. A 394 bp fragment was amplified from genomic DNA of different isolates of C. perniciosa belonging to C-, L-, and S-biotypes and collected from various geographical areas. The cleavage of PCR products with restriction enzymes and the sequencing of various RT fragments indicated the presence of several sequences showing transition events (G:C to A:T). Southern blot analysis revealed high copy numbers of RT signals, forming different patterns among C-, S-, and L-biotype isolates. Sequence comparisons of the predicted RT peptide indicate a close relationship with the RT protein from thegypsy family of LTR-retrotransposons. The possible role of these retrotransposons in generating genetic variability in the homothallic C. perniciosa is discussed.

scholarly journals Whole-Genome Sequence of Mycoplasma bovis Strain Ningxia-1

2018 ◽  
Vol 6 (4) ◽  
Author(s):  
Peng Sun ◽  
Haifeng Luo ◽  
Xin Zhang ◽  
Jingyi Xu ◽  
Yanan Guo ◽  
...  
Keyword(s):  
Genome Sequence ◽  
Single Molecule ◽  
Whole Genome Sequence ◽  
Mycoplasma Bovis ◽  
Sequencing Technology ◽  
Circular Chromosome ◽  
A Genome ◽  
Variable Surface ◽  
Integrative Conjugative Elements ◽  
Single Circular Chromosome

ABSTRACT A genome sequence of the Mycoplasma bovis Ningxia-1 strain was tested by Pacific Biosciences (PacBio) single-molecule real-time (SMRT) sequencing technology. The strain was isolated from a lesioned calf lung in 2013 in Pengyang, Ningxia, China. The single circular chromosome of 1,033,629 bp shows differences between complete Mycoplasma bovis genome in insertion-like sequences (ISs), integrative conjugative elements (ICEs), lipoproteins (LPs), variable surface lipoproteins (VSPs), pathogenicity islands (PAIs), etc.

scholarly journals Computation of Splicing Languages from DNA Splicing System Based on Sequences of Restriction Enzymes

2019 ◽  
Vol 8 (6S3) ◽  
pp. 31-42
Keyword(s):  
Restriction Enzyme ◽  
Dna Computing ◽  
Restriction Enzymes ◽  
Visual Programming ◽  
Cleavage Pattern ◽  
Dna Molecules ◽  
Splicing Systems ◽  
Dna Splicing

In DNA splicing systems, restriction enzymes and ligases cleave and recombine DNA molecules based on the cleavage pattern of the restriction enzymes. The set of molecules resulting from the splicing system depicts a splicing language. In this research, an algorithm for DNA splicing systems is developed using C++ visual programming. The splicing languages which have been characterised through some theorems based on the crossings and sequences of the restriction enzymes, are generated as the output from this computation. In order to generate the splicing languages, the algorithm detects and calculates the number of cutting sites of the restriction enzymes found in the initial molecules, determines whether the sequence of restriction enzyme is a palindrome or not, and if the restriction enzymes have the same or different crossings. The results from this research depict the splicing languages obtained from the manual computations, which contributes to the development of computational software in DNA computing.

scholarly journals Out-Lab Therapy Approach Based on Elected A Restriction Enzyme to Transfer Target Gene

2018 ◽  
Vol 2 (2) ◽  
pp. 196-208
Author(s):  
Ayad Ismaeel
Keyword(s):  
Restriction Enzyme ◽  
Target Gene ◽  
Restriction Enzymes ◽  
Tp53 Gene ◽  
Restriction Enzyme Digestion ◽  
Therapy Approach ◽  
Software Packages ◽  
Important Approach ◽  
Effective Cost ◽  
Target Gene Sequence

An important approach of therapy the target gene sequence causes diseases via repair/recombine the mutated gene (gene transfer) using a restriction enzymes in the laboratory. This approach will cause multiple problems happening accompany to biological laboratory if ruled out problems outside of it like the digested DNA ran as a smear on an agarose gel, incomplete restriction enzyme digestion, extra bands in the gel, etc. The paper suggested new approach of therapy via repair/replacement mutated gene caused disease by detecting primers and finding restriction enzymes using bioinformatics tools, software, packages etc. then achieving the repair/ recombine of mutations before going to the biologic lab (out-lab) to avoid the problems associated these laboratories. Implement and apply this a proposed therapy approach on TP53 gene (which caused more than 50% of human cancers) and after confirming there is mutations on P53 tumor protein shows an effective cost, friendly therapy methodology and comprehensive.

scholarly journals Identification of restriction enzyme in the FSHR gene of indonesian local cattle

2021 ◽  
Vol 888 (1) ◽  
pp. 012024
Author(s):  
P W Prihandini ◽  
A Primasari ◽  
M Luthfi ◽  
D Pamungkas ◽  
A P Z N L Sari ◽  
...  
Keyword(s):  
Restriction Enzyme ◽  
Restriction Site ◽  
Follicle Stimulating Hormone ◽  
Restriction Enzymes ◽  
Future Studies ◽  
Fshr Gene ◽  
Pcr Rflp ◽  
Mapping Analysis ◽  
Pcr Products ◽  
Enzyme Mapping

Abstract The restriction enzyme is important for genotyping using the PCR-RFLP technique. Therefore, this study aims to identify the restriction enzyme mapping in the partial sequence of the follicle-stimulating hormone receptor (FSHR) gene in Indonesian local cattle. A total of 29 samples sized 306 bp, were aligned with Genbank sequence acc no. NC_032660, resulting three polymorphic sites, namely g.193G>C, g.227T>C, and g.275A>C. Furthermore, the restriction mapping analysis using the NEBcutter program V2.0 showed that no enzyme recognized the SNP g.275A>C, while the SNP g.193G>C and g.227T>C were identified by the AluI and MscI enzymes, respectively. The AluI enzyme cuts at two positions (193 bp and 243 bp) in the G allele sample producing three fragments namely 50 bp, 63 bp, and 193 bp, meanwhile, in the C allele, the AluI cuts only in position 243 bp, hence, the fragment products are 63 bp and 243 bp. In contrast, the MscI enzyme was only recognized in the T allele, producing fragments sized 77 bp and 229 bp but failed to identify the restriction site along with the PCR products in the C allele. Based on the results, the SNPs (g.193G>C and g.227T>C) and restriction enzymes (AluI and MscI) are applicable for genotyping local Indonesian cattle using the PCR-RFLP technique in future studies.

scholarly journals Computation of splicing languages from DNA splicing system with one palindromic restriction enzyme

2018 ◽  
Vol 14 (2) ◽  
pp. 188-192
Author(s):  
Nurul Izzaty Ismail ◽  
Wan Heng Fong ◽  
Nor Haniza Sarmin
Keyword(s):  
User Interface ◽  
Graphical User Interface ◽  
Restriction Enzyme ◽  
Restriction Enzymes ◽  
In Vitro Experiments ◽  
Dna Molecules ◽  
C Programming ◽  
Splicing Systems ◽  
Dna Splicing

In DNA splicing system, the potential effects of sets of restriction enzymes and a ligase that allow DNA molecules to be cleaved and reassociated to produce further molecules are studied.  A splicing language depicts the molecules resulting from a splicing system.  In this research, a C++ programming code for DNA splicing system with one palindromic restriction enzyme for one and two (non-overlapping) cutting sites is developed.  A graphical user interface, GUI is then designed to allow the user to insert the initial DNA string and restriction enzymes to generate the splicing languages which are the result of the computation of the C++ programming.  This interface displays the resulting splicing languages, which depict the results from in vitro experiments of the respective splicing system.  The results from this research simplify the lenghty manual computation of the resulting splicing languages of DNA splicing systems with one palindromic restriction enzyme.   

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