DNA Sequence Information Research Articles

RNase H-dependent PCR enables highly specific amplification of antibody variable domains from single B-cells.

Immunization-based antibody discovery platforms require robust and effective protocols for the amplification, cloning, expression, and screening of antibodies from large numbers of B-cells in order to effectively capture the diversity of an experienced Ig-repertoire. Multiplex PCR using a series of forward and reverse primers designed to recover antibodies from a range of different germline sequences is challenging because primer design requires the recovery of full length antibody sequences, low starting template concentrations, and the need for all the primers to function under the same PCR conditions. Here we demonstrate several advantages to incorporating RNase H2-dependent PCR (rh-PCR) into a high-throughput, antibody-discovery platform. Firstly, rh-PCR eliminated primer dimer synthesis to below detectable levels, thereby eliminating clones with a false positive antibody titer. Secondly, by increasing the specificity of PCR, the rh-PCR primers increased the recovery of cognate antibody variable regions from single B-cells, as well as downstream recombinant antibody titers. Finally, we demonstrate that rh-PCR primers provide a more homogeneous sample pool and greater sequence quality in a Next Generation Sequencing-based approach to obtaining DNA sequence information from large numbers of cloned antibody cognate pairs. Furthermore, the higher specificity of the rh-PCR primers allowed for a better match between native antibody germline sequences and the VL/VH fragments amplified from single B-cells.

Pathogenicity Factors of Genomic Islands in Intestinal and Extraintestinal Escherichia coli.

Escherichia coli is a versatile bacterial species that includes both harmless commensal strains and pathogenic strains found in the gastrointestinal tract in humans and warm-blooded animals. The growing amount of DNA sequence information generated in the era of “genomics” has helped to increase our understanding of the factors and mechanisms involved in the diversification of this bacterial species. The pathogenic side of E. coli that is afforded through horizontal transfers of genes encoding virulence factors enables this bacterium to become a highly diverse and adapted pathogen that is responsible for intestinal or extraintestinal diseases in humans and animals. Many of the accessory genes acquired by horizontal transfers form syntenic blocks and are recognized as genomic islands (GIs). These genomic regions contribute to the rapid evolution, diversification and adaptation of E. coli variants because they are frequently subject to rearrangements, excision and transfer, as well as to further acquisition of additional DNA. Here, we review a subgroup of GIs from E. coli termed pathogenicity islands (PAIs), a concept defined in the late 1980s by Jörg Hacker and colleagues in Werner Goebel’s group at the University of Würzburg, Würzburg, Germany. As with other GIs, the PAIs comprise large genomic regions that differ from the rest of the genome by their G + C content, by their typical insertion within transfer RNA genes, and by their harboring of direct repeats (at their ends), integrase determinants, or other mobility loci. The hallmark of PAIs is their contribution to the emergence of virulent bacteria and to the development of intestinal and extraintestinal diseases. This review summarizes the current knowledge on the structure and functional features of PAIs, on PAI-encoded E. coli pathogenicity factors and on the role of PAIs in host–pathogen interactions.

Somatic Mutations in LRRK2 Identify a Subset of Invasive Mammary Carcinomas Associated with High Mutation Burden

Mutations in the leucine-rich repeat kinase 2 (LRRK2) gene are the most common cause of familial Parkinson disease. Although LRRK2-related Parkinson disease patients have a heightened risk of certain nonskin cancers, including breast cancer, it is unknown whether LRRK2 somatic mutations occur and are associated with breast cancer. The objective of this study was to evaluate the occurrence of LRRK2 somatic mutations in breast cancer and the clinicopathologic features associated with LRRK2-mutated tumors. Using The Cancer Genome Atlas Breast Cancer Project, somatic LRRK2 DNA sequence information was obtained for 93 cases, of which 17 cases (18%) with 18 mutations were identified. LRRK2-mutated mammary carcinomas are enriched with stop-gain, truncating mutations predicted to result in loss of function; missense mutations frequently targeted the GTPase and kinase domains. Tumors displayed predominantly high-grade morphology with abundant granular eosinophilic cytoplasm, resembling mitochondria-rich apocrine-like carcinomas. Exploration of the genomic landscape of LRRK2-mutated carcinomas yielded frequent TP53 deactivation and a remarkably high tumor mutation burden. More important, breast cancers with LRRK2 mutations are associated with reduced patient survival compared with The Cancer Genome Atlas Breast Cancer Project cohort. These findings, for the first time, show that somatic LRRK2 mutations occur frequently in breast cancer, and the high mutation burden seen in this subset of tumors suggests that LRRK2 mutations may herald benefit from immune checkpoint inhibition.

Revision of Coelastrella (Scenedesmaceae, Chlorophyta) and first register of this green coccoid microalga for continental Norway

A terrestrial green microalga was isolated at Ås, in Akershus County, Norway. The strain corresponded to a coccoid chlorophyte. Morphological characteristics by light and electron microscopy, in conjunction with DNA amplification and sequencing of the 18 s rDNA gene and ITS sequences, were used to identify the microalgae. The characteristics agree with those of the genus Coelastrella defined by Chodat, and formed a sister group with the recently described C. thermophila var. globulina. Coelastrella is a relatively small numbered genus that has not been observed in continental Norway before; there are no previous cultures available in collections of Norwegian strains. Gas chromatography analyses of the FAME-derivatives showed a high percentage of polyunsaturated fatty acids (44–45%) especially linolenic acid (C18:3n3; 30–34%). After the stationary phase, the cultures were able to accumulate several carotenoids as neoxanthin, pheophytin a, astaxanthin, canthaxanthin, lutein, and violaxanthin. Due to the scarcity of visual characters suitable for diagnostic purposes and the lack of DNA sequence information, there is a high possibility that species of this genus have been neglected in local environmental studies, even though it showed interesting properties for algal biotechnology.

DNA Data Storage in Perl

Here we report a simple and flexible method for DNA data storage based on Perl script. For this approach, the text data of the preamble of the “Universal Declaration of Human Rights” consisting of 2,046 words was encoded into the corresponding 8,148 base pairs of DNA using Perl-based encoding with a hash table. The encoded DNA sequences were then artificially synthesized for storage. The information DNA consisted of a total of 22 chemically synthesized DNA fragments with 400 nucleotides each, which were inserted into a cloning vector to multiply the plasmid DNA. The nucleotide integrity of the data-carrying DNA sequences were ensured under the accelerated aging conditions. Also, an erroneous nucleotide in the information DNA sequences was successfully corrected using the overlap extension PCR method. The stored DNA was read by sequencing, and the resulting DNA sequence information was successfully decoded to convert the DNA records back to the original document. Our results indicate that textual data can be stored in DNA using a simple, easy, and flexible Perl by running a script from the command line.

Exclusive use of digital PCR allows an absolute assay of heat-killed Lactobacilli in foods targeting multiple copies of 16S rDNA

The real-time PCR (qPCR) and digital PCR (dPCR) to amplify a single-copy of house-keeping genes (i.e., hsp60, pheS or tuf) are used for the assay of limited microbial species. In general, with a single-copy gene, there are obviously varied DNA sequences for even the same microbial species, which could cause difficulties with design of primers and probes for PCR when targeting various single copy genes. In general, for identification by dPCR (as a representative case: Lactobacillus paracasei), accumulated DNA sequence information of 16S rDNA, which is much more frequently used, should be targeted. In contrast, next-generation sequencing revealed that there are five copies of 16S rDNA in a live L. paracasei MCC1849. Therefore, we aimed to reveal, if heat-killed L. paracasei supplemented in nutritional foods that aid the host immune system have the relevant five copies per chromosomal DNA, and if the relevant copies remain unchanged on the same chromosomal DNA or remain to be different chromosomal DNA fragments. So, we revealed the actual distribution of the potential original five copies of 16S rDNA using our innovative dPCR, in which both 16S rDNA and hsp60 genes were simultaneously elongated. The molecular ratios of 16S rDNA/hsp60 dispersed in the dPCR chip were then estimated. The 16S rDNA/hsp60 molecular ratios of the heat-killed L. paracasei in foods, resultantly ranged from 5.0 to 7.2, being the same or higher than that of the five copies determined by next-generation sequencing. The 16S rDNA copy number/ratio indicated the chromosomal DNA molecular number and the associated cell number. As significance, different nutritional foods could potentially cause the loss of chromosomal DNA of supplemented beneficial microbes to a much greater degree. Our absolute dPCR does not require standard correlative samples for the estimation of final products. The estimation principle of the ratio of 16S rDNA/a house-keeping single-copy gene by our absolute dPCR could lead to a useful and accurate assay for various nutritional foods.

Enzymatic Cleavage of 3\u2019-Esterified Nucleotides Enables a Long, Continuous DNA Synthesis

The reversible dye-terminator (RDT)-based DNA sequencing-by-synthesis (SBS) chemistry has driven the advancement of the next-generation sequencing technologies for the past two decades. The RDT-based SBS chemistry relies on the DNA polymerase reaction to incorporate the RDT nucleotide (NT) for extracting DNA sequence information. The main drawback of this chemistry is the “DNA scar” issue since the removal of dye molecule from the RDT-NT after each sequencing reaction cycle leaves an extra chemical residue in the newly synthesized DNA. To circumvent this problem, we designed a novel class of reversible (2-aminoethoxy)-3-propionyl (Aep)-dNTPs by esterifying the 3’-hydroxyl group (3’-OH) of deoxyribonucleoside triphosphate (dNTP) and examined the NT-incorporation activities by A-family DNA polymerases. Using the large fragment of both Bacillus stearothermophilus (BF) and E. coli DNA polymerase I (KF) as model enzymes, we further showed that both proteins efficiently and faithfully incorporated the 3’-Aep-dNMP. Additionally, we analyzed the post-incorporation product of N + 1 primer and confirmed that the 3’-protecting group of 3’-Aep-dNMP was converted back to a normal 3’-OH after it was incorporated into the growing DNA chain by BF. By applying all four 3’-Aep-dNTPs and BF for an in vitro DNA synthesis reaction, we demonstrated that the enzyme-mediated deprotection of inserted 3’-Aep-dNMP permits a long, continuous, and scar-free DNA synthesis.

A new numerical approach for DNA representation using modified Gabor wavelet transform for the identification of protein coding regions

The fundamental step in genomic signal processing applications is to assign mathematical descriptor to nucleotides {A, T, G, C} of DNA molecule for discrete representation. The discrete representation should replicate biological information of gene when analyzed with digital signal processing tools. In this aspect, a novel binary representation of DNA sequence by combining structural and chemical information of original DNA sequence has been proposed for the identification of protein coding regions of eukaryotes. The identification model comprises two stages, mainly, numerical encoding in first stage, and analysis of biological behavior through digital signal processing algorithms in second stage. In the first stage, a new numerical encoding method based on Walsh codes of order-4 is proposed to obtain 1-D binary discrete sequence. In the second stage, the modified Gabor wavelet transform (MGWT) is employed on the discretized DNA sequence for spectrum analysis. The optimal gene numerical encoding and multiresolution approach of MGWT has readily identified the structures of coding regions of unknown gene sequences. The proposed model is validated by analyzing prediction efficiency in terms of statistical metrics such as sensitivity, specificity, accuracy on both sequence and data base level. Furthermore, the results are compared by plotting receiver operating curves (ROC) for all classification thresholds for the state-of-art encoding methods. Area under curve (AUC) value of 0.86 at sequence level and 0.84 at database level is achieved. Performance metrics indicate that the proposed encoding method exhibits relatively better performance than other numerical encoding methods.

Two SNP Markers Identified Using Genotyping-by-Sequencing Are Associated with Remontancy in a Segregating F1 Population of Syringa meyeri ‘Palibin’ × S. pubescens ‘Penda’ Bloomerang®

Lilacs (Syringa sp.) have been used as ornamental plants since the mid-16th century and remain important in modern gardens due to their attractive and fragrant flowers. However, a short flowering season is a critical drawback for their ornamental value. Breeders have identified remontancy (reblooming) in dwarf lilac (Syringa pubescens), and have tried to introgress this trait into related species by interspecific hybridization. Molecular tools for lilac breeding are limited because of the shortage of genome sequence knowledge and currently no molecular markers are available to use in breeding for remontancy. In this study, an F1 population from crossing Syringa meyeri ‘Palibin’ × S. pubescens ‘Penda’ Bloomerang® Purple was created and subjected to genotyping-by-sequencing (GBS) analysis and phenotyped for remontancy. Plants were categorized as remontant, semi-remontant, and nonremontant based on the relative quantity of inflorescences during the second flush of flowers. A total of 20,730 single-nucleotide polymorphism (SNP) markers from GBS were used in marker-trait association to find remontant-specific marker(s) without marker position information. Two SNP markers, TP70580 (A locus) and TP82604 (B locus), were correlated with remontancy. The two loci showed a partial epistasis and additive interaction effects on the level of remontancy. Accumulation of recessive alleles at the two loci was positively correlated with increased reblooming. For example, 87% of aabb plants were remontant, and only 9% were nonremontant. In contrast, 100% of AaBB plants were nonremontant. These two SNP markers associated with remontancy will be useful in developing markers for future breeding and demonstrate the feasibility of developing markers for breeding woody ornamental taxa that lack a reference genome or extensive DNA sequence information.

Taxonomic notes on Scutellaria taipeiensis (Lamiaceae) from morphological and molecular data.

The genus Scutellaria comprises eight species distributed from 50 to 2000 m in Taiwan. Amongst them, S. barbata and S. taipeiensis are very similar on the basis of morphological and plastid DNA sequence information. Therefore, a comprehensive study of the taxonomic status of S. taipeiensis is necessary. We reviewed the herbarium sheets, related literature and protologues and compared morphologies of these two species, as well as their phylogenetic relationships. All evidence, including the diagnostic characters between S. taipeiensis and S. barbata, suggest that they belonged to a single species rather than two. As a result, S. taipeiensis is treated as a synonym of S. barbata.

Integrative taxonomic consideration of the Holarctic Euconulus fulvus group of land snails (Gastropoda, Stylommatophora)

While among the most common Holarctic land snails, species of the Euconulus fulvus group have been subject to considerable recent taxonomic controversy. Based on 76 Euconulus populations collected across Eurasia and North America, we empirically evaluated these competing taxonomic hypotheses through an integration of nDNA and mtDNA phylogenetics, shell morphometrics and various qualitative traits. Our results support the existence of five taxa: Euconulus alderi (Atlantic Europe to western North America), E. fresti sp. nov. (North America), E. fulvus fulvus (Europe), E. fulvus egenus (central Asia to Atlantic North America), and E. polygyratus (north-eastern North America). Each species-level entity possessed a unique suite of observable shell features allowing for accurate identification without need of DNA sequence information. Our data did not support the recent erection of E. callopisticus, E. praticola, and E. trochiformis to species-level status. This work also helps illustrate the importance of using consensus across DNA and shell/soft body features in assessing species-level taxonomy and in determining those features which allow for accurate identification. It furthermore documents the importance of basing taxonomic work on samples drawn from across the entire geographic and ecological range of the study group.http://zoobank.org/urn:lsid:zoobank.org:pub:97985479-EDF8-4B94-A825-AC42BF31F597

A novel method SEProm for prokaryotic promoter prediction based on DNA structure and energetics.

Despite conservation in general architecture of promoters and protein-DNA interaction interface of RNA polymerases among various prokaryotes, identification of promoter regions in the whole genome sequences remains a daunting challenge. The available tools for promoter prediction do not seem to address the problem satisfactorily, apparently because the biochemical nature of promoter signals is yet to be understood fully. Using 28 structural and 3 energetic parameters, we found that prokaryotic promoter regions have a unique structural and energy state, quite distinct from that of coding regions and the information for this signature state is in-built in their sequences. We developed a novel promoter prediction tool from these 31 parameters using various statistical techniques. Here, we introduce SEProm, a novel tool that is developed by studying and utilizing the in-built structural and energy information of DNA sequences, which is applicable to all prokaryotes including archaea. Compared to five most recent, diverged and current best available tools, SEProm performs much better, predicting promoters with an 'F-value' of 82.04 and 'Precision' of 81.08. The next best 'F-value' was obtained with PromPredict (72.14) followed by BProm (68.37). On the basis of 'Precision' value, the next best 'Precision' was observed for Pepper (75.39) followed by PromPredict (72.01). SEProm maintained the lead even when comparison was done on two test organisms (not involved in training for SEProm). The software is freely available with easy to follow instructions (www.scfbio-iitd.res.in/software/TSS_Predict.jsp). Supplementary data are available at Bioinformatics online.

Large scale genome skimming from herbarium material for accurate plant identification and phylogenomics

BackgroundHerbaria are valuable sources of extensive curated plant material that are now accessible to genetic studies because of advances in high-throughput, next-generation sequencing methods. As an applied assessment of large-scale recovery of plastid and ribosomal genome sequences from herbarium material for plant identification and phylogenomics, we sequenced 672 samples covering 21 families, 142 genera and 530 named and proposed named species. We explored the impact of parameters such as sample age, DNA concentration and quality, read depth and fragment length on plastid assembly error. We also tested the efficacy of DNA sequence information for identifying plant samples using 45 specimens recently collected in the Pilbara.ResultsGenome skimming was effective at producing genomic information at large scale. Substantial sequence information on the chloroplast genome was obtained from 96.1% of samples, and complete or near-complete sequences of the nuclear ribosomal RNA gene repeat were obtained from 93.3% of samples. We were able to extract sequences for the core DNA barcode regions rbcL and matK from 96 to 93.3% of samples, respectively. Read quality and DNA fragment length had significant effects on sequencing outcomes and error correction of reads proved essential. Assembly problems were specific to certain taxa with low GC and high repeat content (Goodenia, Scaevola, Cyperus, Bulbostylis, Fimbristylis) suggesting biological rather than technical explanations. The structure of related genomes was needed to guide the assembly of repeats that exceeded the read length. DNA-based matching proved highly effective and showed that the efficacy for species identification declined in the order cpDNA >> rDNA > matK >> rbcL.ConclusionsWe showed that a large-scale approach to genome sequencing using herbarium specimens produces high-quality complete cpDNA and rDNA sequences as a source of data for DNA barcoding and phylogenomics.

DNA Barcoding Identifies Juniperus oxycedrus subsp. macrocarpa in Derna Region, East Libya

Many methods are accessible that apply diverse criteria for the reasons of identifying taxonomic specialization depending on DNA sequencing information. It is crucial for the studies of taxonomy and biodiversity using DNA barcode technology to fast and accurately make species identification in the forests. According to the Encyclopedia of Earth, many wonderful and rare plants are destroyed. In the tropics and subtropics, numerous evergreen conifers are jeopardized. These plants grow in remote places of our planet, which are inaccessible. It merits referring to that Juniperus spp. an imperative part of Mediterranean arid and semi-arid biological communities. Juniperus oxycedrus subsp. macrocarpa is a rare woody species found in Jebel Al-Akhdar, Libya in only one peripheral site north-west Derna. A robust analysis presented based on using morphological traits of needles, seeds and cones, and DNA technology. Along these lines, jeopardized plant populations could be recognized more effectively. This study universality of tree species DNA barcodes. such as the rbcL and matK plastid markers, and examined their abilities of species identification. The morphological and genetic results strongly support the recognition of J. macrocarpa at the subspecies J. oxycedrus.

Reconstructing protein-coding sequences from ancient DNA.

Obtaining information about functional details of proteins of extinct species is of critical importance for a better understanding of the real-life appearance, behavior and ecology of these lost entries in the book of life. In this chapter, we discuss the possibilities to retrieve the necessary DNA sequence information from paleogenomic data obtained from fossil specimens, which can then be used to express and subsequently analyze the protein of interest. We discuss the problems specific to ancient DNA, including miscoding lesions, short read length and incomplete paleogenome assemblies. Finally, we discuss an alternative, but currently rarely used approach, direct PCR amplification, which is especially useful for comparatively short proteins.

2D Graphical Representation of DNA Sequences Based on Variant Map

Mining and analyzing the information and structure of DNA sequences is an important method of exploring and explaining the mysteries of biology. In addition to traditional biological detection methods for DNA sequences, technological advances have allowed researchers to utilize computers to analyze DNA sequences, such as in DNA visualization. However, DNA visualization techniques have disadvantages, such as difficult operation and complex calculation. Therefore, in this paper, we propose an algorithm called VARCH based on a variant map that is a part of variant logic construction. VARCH is a high-efficiency and concise visualization algorithm for DNA sequences. The principle of this approach is that the number of base combinations in each DNA subsequence is counted, and a matrix is constructed based on the full arrangement of different base combinations. Finally, the matrix is mapped to the 2D coordinate plane, obtaining the image of DNA sequences. The experimental results demonstrate that VARCH can effectively display features of DNA sequences.

I4mC-Mouse: Improved identification of DNA N4-methylcytosine sites in the mouse genome using multiple encoding schemes

N4-methylcytosine (4mC) is one of the most important DNA modifications and involved in regulating cell differentiations and gene expressions. The accurate identification of 4mC sites is necessary to understand various biological functions. In this work, we developed a new computational predictor called i4mC-Mouse to identify 4mC sites in the mouse genome. Herein, six encoding schemes of k-space nucleotide composition (KSNC), k-mer nucleotide composition (Kmer), mono nucleotide binary encoding (MBE), dinucleotide binary encoding, electron–ion interaction pseudo potentials (EIIP) and dinucleotide physicochemical composition were explored that cover different characteristics of DNA sequence information. Subsequently, we built six RF-based encoding models and then linearly combined their probability scores to construct the final predictor. Among the six RF-based models, the Kmer, KSNC, MBE, and EIIP encodings are sufficient, which contributed to 10%, 45%, 25%, and 20% of the prediction performance, respectively. On the independent test the i4mC-Mouse predicted the 4mC sites with accuracy and MCC of 0.816 and 0.633, respectively, which were approximately 2.5% and 5% higher than those of the existing method (4mCpred-EL). For experimental biologists, a freely available web application was implemented at http://kurata14.bio.kyutech.ac.jp/i4mC-Mouse/.

Genetic Characterization of Maize Kisar Var. Kuning Genjah and Maize Var. Bisi-II-Hibrida Based On Molekular RAPD Marker

The variety of maize in Southwest Maluku (MBD) has an ability to grow in the extremely dry land, especially at Kisar island, because those areas were dominated by dry land. Dry resistance very useful to be used for genetical characterization in order to provide accurate data about the character of local maize Kisar var. Kuning Genjah. To identify the characteristics at the genome level of this local maize variety, one of the molecular methods that can be applied was RAPD molecular method. The advantage of the RAPD marker is it's technically simpler and fast in testing, does not require DNA sequence information, hence this marker becomes widely used, it only requires a small sample of DNA, the primary is commercially available and does not use radioactive compounds. The aim of this study was to find out the bands profile that has the potential to be used as a marker of dry resistance. The method applied was RAPD (Random Amplified Polymorphic DNA) using OPA-02, OPA-07, and OPC-12 primers and produces 22 DNA bands. The measurement of bands by estimating the molecular weight based on marker exponential regression. The polymorphic percentage was 90.9% between var. Kuning genjah and var. BIZI-II-Hybrid. The percentage of polymorphism showed the potential of bands that can be used as molecular markers for markers of dry resistance that can be utilized in plant breeding

Epigenética: la relación del medio ambiente con el genoma y su influencia en la salud mental.

El estudio del genoma humano, cuando efectuado solo a través de la información secuencial de ADN, no explica del todo el alto nivel de variación inter-individual usualmente observado. Es la epigenética la que permite explicar aquellas variaciones de expresión génica como un proceso reversible y hereditario en el corto plazo, bajo la influencia del medio ambiente durante diversas etapas del desarrollo y la edad adulta, sin modificar la secuencia genética. Las alteraciones epigenéticas están siendo ya incorporadas como elementos valiosos en la posible identificación de biomarcadores. Además, debido a su naturaleza reversible, pueden constituirse en factores de mejoría de síntomas de enfermedad mediante el uso de enfoques terapéuticos. El presente artículo explica los mecanismos de inhibición de la expresión génica, su relación con el medio ambiente, la dieta y su influencia en la evolución, la ocurrencia de enfermedades, la conducta humana y la salud mental.

I4mC-ROSE, a bioinformatics tool for the identification of DNA N4-methylcytosine sites in the Rosaceae genome

One of the most important epigenetic modifications is N4-methylcytosine, which regulates many biological processes including DNA replication and chromosome stability. Identification of N4-methylcytosine sites is pivotal to understand specific biological functions. Herein, we developed the first bioinformatics tool called i4mC-ROSE for identifying N4-methylcytosine sites in the genomes of Fragaria vesca and Rosa chinensis in the Rosaceae, which utilizes a random forest classifier with six encoding methods that cover various aspects of DNA sequence information. The i4mC-ROSE predictor achieves area under the curve scores of 0.883 and 0.889 for the two genomes during cross-validation. Moreover, the i4mC-ROSE outperforms other classifiers tested in this study when objectively evaluated on the independent datasets. The proposed i4mC-ROSE tool can serve users' demand for the prediction of 4mC sites in the Rosaceae genome. The i4mC-ROSE predictor and utilized datasets are publicly accessible at http://kurata14.bio.kyutech.ac.jp/i4mC-ROSE/.