Base-resolution analysis of 5-hydroxymethylcytosine in the mammalian genome.

The study of 5-hydroxylmethylcytosines (5hmC) has been hampered by the lack of a method to map it at single-base resolution on a genome-wide scale. Affinity purification-based methods cannot precisely locate 5hmC nor accurately determine its relative abundance at each modified site. We here present a genome-wide approach, Tet-assisted bisulfite sequencing (TAB-Seq), that when combined with traditional bisulfite sequencing can be used for mapping 5hmC at base resolution and quantifying the relative abundance of 5hmC as well as 5mC. Application of this method to embryonic stem cells not only confirms widespread distribution of 5hmC in the mammalian genome but also reveals sequence bias and strand asymmetry at 5hmC sites. We observe high levels of 5hmC and reciprocally low levels of 5mC near but not on transcription factor-binding sites. Additionally, the relative abundance of 5hmC varies significantly among distinct functional sequence elements, suggesting different mechanisms for 5hmC deposition and maintenance.

Genome Analysis of a Newly Sequenced B. subtilis SRCM117797 and Multiple Public B. subtilis Genomes Unveils Insights into Strain Diversification and Biased Core Gene Distribution.

The bacterium Bacillus subtilis is a widely used study model and industrial workhorse organism that belongs to the group of gram-positive bacteria. In this study, we report the analysis of a newly sequenced complete genome of B. subtilis strain SRCM117797 along with a comparative genomics of a large collection of B. subtilis strain genomes. B. subtilis strain SRCM117797 has 4,255,638 bp long chromosome with 43.4% GC content and high coding sequence association with macromolecules, metabolism, and phage genes. Genomic diversity analysis of 232 B. subtilis strains resulted in the identification of eight clusters and three singletons. Of 147 B. subtilis strains included, 89.12% had strain-specific genes, of which 6.75% encoded strain-specific insertion sequence family transposases. Our analysis showed a potential role of strain-specific insertion sequence family transposases in intra-cellular accumulation of strain-specific genes. Furthermore, the chromosomal layout of the core genes was biased: overrepresented on the upper half (closer to the origin of replication) of the chromosome, which may explain the fast-growing characteristics of B. subtilis. Overall, the study provides a complete genome sequence of B. subtilis strain SRCM117797, show an extensive genomic diversity of B. subtilis strains and insights into strain diversification mechanism and non-random chromosomal layout of core genes.

Admixture of divergent genomes facilitates hybridization across species in the family Brassicaceae.

Hybridization and polyploidization are pivotal to plant evolution. Genetic crosses between distantly related species are rare in nature due to reproductive barriers but how such hurdles can be overcome is largely unknown. Here we report the hybrid genome structure of xBrassicoraphanus, a synthetic allotetraploid of Brassica rapa and Raphanus sativus. We performed cytogenetic analysis and de novo genome assembly to examine chromosome behaviors and genome integrity in the hybrid. Transcriptome analysis was conducted to investigate expression of duplicated genes in conjunction with epigenome analysis to address whether genome admixture entails epigenetic reconfiguration. Allotetraploid xBrassicoraphanus retains both parental chromosomes without genome rearrangement. Meiotic synapsis formation and chromosome exchange are avoided between nonhomologous progenitor chromosomes. Reconfiguration of transcription network occurs, and less divergent cis-elements of duplicated genes are associated with convergent expression. Genome-wide DNA methylation asymmetry between progenitors is largely maintained but, notably, B. rapa-originated transposable elements are transcriptionally silenced in xBrassicoraphanus through gain of DNA methylation. Our results demonstrate that hybrid genome stabilization and transcription compatibility necessitate epigenome landscape adjustment and rewiring of cis-trans interactions. Overall, this study suggests that a certain extent of genome divergence facilitates hybridization across species, which may explain the great diversification and expansion of angiosperms during evolution.

Der f 38 Is a Novel TLR4-Binding Allergen Related to Allergy Pathogenesis from Dermatophagoides farinae.

It is difficult to treat allergic diseases including asthma completely because its pathogenesis remains unclear. House dust mite (HDM) is a critical allergen and Toll-like receptor (TLR) 4 is a member of the toll-like receptor family, which plays an important role in allergic diseases. The purpose of this study was to characterize a novel allergen, Der f 38 binding to TLR4, and unveil its role as an inducer of allergy. Der f 38 expression was detected in the body and feces of Dermatophagoides farinae (DF). Electron microscopy revealed that it was located in the granule layer, the epithelium layer, and microvilli of the posterior midgut. The skin prick test showed that 60% of allergic subjects were Der f 38-positive. Der f 38 enhanced surface 203c expression in basophils of Der f 38-positive allergic subjects. By analysis of the model structure of Der p 38, the expected epitope sites are exposed on the exterior side. In animal experiments, Der f 38 triggered an infiltration of inflammatory cells. Intranasal (IN) administration of Der f 38 increased neutrophils in the lung. Intraperitoneal (IP) and IN injections of Der f 38 induced both eosinophils and neutrophils. Increased total IgE level and histopathological features were found in BALB/c mice treated with Der f 38 by IP and IN injections. TLR4 knockout (KO) BALB/c mice exhibited less inflammation and IgE level in the sera compared to wild type (WT) mice. Der f 38 directly binds to TLR4 using biolayer interferometry. Der f 38 suppressed the apoptosis of neutrophils and eosinophils by downregulating proteins in the proapoptotic pathway including caspase 9, caspase 3, and BAX and upregulating proteins in the anti-apoptotic pathway including BCL-2 and MCL-1. These findings might shed light on the pathogenic mechanisms of allergy to HDM.

Genomic Survey and Biochemical Analysis of Recombinant Candidate Cyanobacteriochromes Reveals Enrichment for Near UV/Violet Sensors in the Halotolerant and Alkaliphilic Cyanobacterium Microcoleus IPPAS B353.

Cyanobacteriochromes (CBCRs), which are exclusive to and widespread among cyanobacteria, are photoproteins that sense the entire range of near-UV and visible light. CBCRs are related to the red/far-red phytochromes that utilize linear tetrapyrrole (bilin) chromophores. Best characterized from the unicellular cyanobacterium Synechocystis sp. PCC 6803 and the multicellular heterocyst forming filamentous cyanobacteria Nostoc punctiforme ATCC 29133 and Anabaena sp. PCC 7120, CBCRs have been poorly investigated in mat-forming, nonheterocystous cyanobacteria. In this study, we sequenced the genome of one of such species, Microcoleus IPPAS B353 (Microcoleus B353), and identified two phytochromes and seven CBCRs with one or more bilin-binding cGMP-specific phosphodiesterase, adenylyl cyclase and FhlA (GAF) domains. Biochemical and spectroscopic measurements of 23 purified GAF proteins from phycocyanobilin (PCB) producing recombinant Escherichia coli indicated that 13 of these proteins formed near-UV and visible light-absorbing covalent adducts: 10 GAFs contained PCB chromophores, whereas three contained the PCB isomer, phycoviolobilin (PVB). Furthermore, the complement of Microcoleus B353 CBCRs is enriched in near-UV and violet sensors, but lacks red/green and green/red CBCRs that are widely distributed in other cyanobacteria. We hypothesize that enrichment in short wavelength-absorbing CBCRs is critical for acclimation to high-light environments where this organism is found.

Development, validation and genetic analysis of a large soybean SNP genotyping array.

Cultivated soybean (Glycine max) suffers from a narrow germplasm relative to other crop species, probably because of under-use of wild soybean (Glycine soja) as a breeding resource. Use of a single nucleotide polymorphism (SNP) genotyping array is a promising method for dissecting cultivated and wild germplasms to identify important adaptive genes through high-density genetic mapping and genome-wide association studies. Here we describe a large soybean SNP array for use in diversity analyses, linkage mapping and genome-wide association analyses. More than four million high-quality SNPs identified from high-depth genome re-sequencing of 16 soybean accessions and low-depth genome re-sequencing of 31 soybean accessions were used to select 180,961 SNPs for creation of the Axiom(®) SoyaSNP array. Validation analysis for a set of 222 diverse soybean lines showed that 170,223 markers were of good quality for genotyping. Phylogenetic and allele frequency analyses of the validation set data indicated that accessions showing an intermediate morphology between cultivated and wild soybeans collected in Korea were natural hybrids. More than 90 unanchored scaffolds in the current soybean reference sequence were assigned to chromosomes using this array. Finally, dense average spacing and preferential distribution of the SNPs in gene-rich chromosomal regions suggest that this array may be suitable for genome-wide association studies of soybean germplasm. Taken together, these results suggest that use of this array may be a powerful method for soybean genetic analyses relating to many aspects of soybean breeding.

Genome-wide resequencing of KRICE_CORE reveals their potential for future breeding, as well as functional and evolutionary studies in the post-genomic era.

BACKGROUND: Rice germplasm collections continue to grow in number and size around the world. Since maintaining and screening such massive resources remains challenging, it is important to establish practical methods to manage them. A core collection, by definition, refers to a subset of the entire population that preserves the majority of genetic diversity, enhancing the efficiency of germplasm utilization. RESULTS: Here, we report whole-genome resequencing of the 137 rice mini core collection or Korean rice core set (KRICE_CORE) that represents 25,604 rice germplasms deposited in the Korean genebank of the Rural Development Administration (RDA). We implemented the Illumina HiSeq 2000 and 2500 platform to produce short reads and then assembled those with 9.8 depths using Nipponbare as a reference. Comparisons of the sequences with the reference genome yielded more than 15 million (M) single nucleotide polymorphisms (SNPs) and 1.3 M INDELs. Phylogenetic and population analyses using 2,046,529 high-quality SNPs successfully assigned rice accessions to the relevant rice subgroups, suggesting that these SNPs capture evolutionary signatures that have accumulated in rice subpopulations. Furthermore, genome-wide association studies (GWAS) for four exemplary agronomic traits in the KRIC_CORE manifest the utility of KRICE_CORE; that is, identifying previously defined genes or novel genetic factors that potentially regulate important phenotypes. CONCLUSION: This study provides strong evidence that the size of KRICE_CORE is small but contains high genetic and functional diversity across the genome. Thus, our resequencing results will be useful for future breeding, as well as functional and evolutionary studies, in the post-genomic era.

Functional analysis of three BrMYB28 transcription factors controlling the biosynthesis of glucosinolates in Brassica rapa.

Glucosinolates (GSLs) are secondary metabolites that have anticarcinogenic activity and play defense roles in plants of the Brassicaceae family. MYB28 is known as a transcription factor that regulates aliphatic GSL biosynthesis in Arabidopsis thaliana. Brassicaceae plants have three orthologous copies of AtMYB28 derived from recent genome triplication. These BrMYB28 genes have a high level of sequence homology, with 81-87% similarities in the coding DNA sequence compared to Arabidopsis. Overexpression of three paralogous BrMYB28 genes in transgenic Chinese cabbage increased the total GSL content in all T1 generation plants and in two inbred lines of homozygous T2 plants. The highest total GSL contents were detected in homozygous T2 lines overexpressing BrMYB28.1, which showed an approximate fivefold increase compared to that of nontransgenic plants. The homozygous T2 lines with overexpressed BrMYB28.1 also showed an increased content of aliphatic, indolic, and aromatic GSLs compared to that of nontransgenic plants. Furthermore, all of the three BrMYB28 genes were identified as negative regulators of BrAOP2 and positive regulators of BrGSL-OH in the homozygous T2 lines. These data indicate the regulatory mechanism of GSL biosynthesis in B. rapa is unlike that in A. thaliana. Our results will provide useful information for elucidating the regulatory mechanism of GSL biosynthesis in polyploid plants.

Elucidating the triplicated ancestral genome structure of radish based on chromosome-level comparison with the Brassica genomes.

KEYMESSAGE: This study presents a chromosome-scale draft genome sequence of radish that is assembled into nine chromosomal pseudomolecules. A comprehensive comparative genome analysis with the Brassica genomes provides genomic evidences on the evolution of the mesohexaploid radish genome. Radish (Raphanus sativus L.) is an agronomically important root vegetable crop and its origin and phylogenetic position in the tribe Brassiceae is controversial. Here we present a comprehensive analysis of the radish genome based on the chromosome sequences of R. sativus cv. WK10039. The radish genome was sequenced and assembled into 426.2 Mb spanning >98 % of the gene space, of which 344.0 Mb were integrated into nine chromosome pseudomolecules. Approximately 36 % of the genome was repetitive sequences and 46,514 protein-coding genes were predicted and annotated. Comparative mapping of the tPCK-like ancestral genome revealed that the radish genome has intermediate characteristics between the Brassica A/C and B genomes in the triplicated segments, suggesting an internal origin from the genus Brassica. The evolutionary characteristics shared between radish and other Brassica species provided genomic evidences that the current form of nine chromosomes in radish was rearranged from the chromosomes of hexaploid progenitor. Overall, this study provides a chromosome-scale draft genome sequence of radish as well as novel insight into evolution of the mesohexaploid genomes in the tribe Brassiceae.

Identification of candidate domestication regions in the radish genome based on high-depth resequencing analysis of 17 genotypes.

KEY MESSAGE: This study provides high-quality variation data of diverse radish genotypes. Genome-wide SNP comparison along with RNA-seq analysis identified candidate genes related to domestication that have potential as trait-related markers for genetics and breeding of radish. Radish (Raphanus sativus L.) is an annual root vegetable crop that also encompasses diverse wild species. Radish has a long history of domestication, but the origins and selective sweep of cultivated radishes remain controversial. Here, we present comprehensive whole-genome resequencing analysis of radish to explore genomic variation between the radish genotypes and to identify genetic bottlenecks due to domestication in Asian cultivars. High-depth resequencing and multi-sample genotyping analysis of ten cultivated and seven wild accessions obtained 4.0 million high-quality homozygous single-nucleotide polymorphisms (SNPs)/insertions or deletions. Variation analysis revealed that Asian cultivated radish types are closely related to wild Asian accessions, but are distinct from European/American cultivated radishes, supporting the notion that Asian cultivars were domesticated from wild Asian genotypes. SNP comparison between Asian genotypes identified 153 candidate domestication regions (CDRs) containing 512 genes. Network analysis of the genes in CDRs functioning in plant signaling pathways and biochemical processes identified group of genes related to root architecture, cell wall, sugar metabolism, and glucosinolate biosynthesis. Expression profiling of the genes during root development suggested that domestication-related selective advantages included a main taproot with few branched lateral roots, reduced cell wall rigidity and favorable taste. Overall, this study provides evolutionary insights into domestication-related genetic selection in radish as well as identification of gene candidates with the potential to act as trait-related markers for background selection of elite lines in molecular breeding.

Major repeat components covering one-third of the ginseng (Panax ginseng C.A. Meyer) genome and evidence for allotetraploidy.

Ginseng (Panax ginseng) is a famous medicinal herb, but the composition and structure of its genome are largely unknown. Here we characterized the major repeat components and inspected their distribution in the ginseng genome. By analyzing three repeat-rich bacterial artificial chromosome (BAC) sequences from ginseng, we identified complex insertion patterns of 34 long terminal repeat retrotransposons (LTR-RTs) and 11 LTR-RT derivatives accounting for more than 80% of the BAC sequences. The LTR-RTs were classified into three Ty3/gypsy (PgDel, PgTat and PgAthila) and two Ty1/Copia (PgTork and PgOryco) families. Mapping of 30-Gbp Illumina whole-genome shotgun reads to the BAC sequences revealed that these five LTR-RT families occupy at least 34% of the ginseng genome. The Ty3/Gypsy families were predominant, comprising 74 and 33% of the BAC sequences and the genome, respectively. In particular, the PgDel family accounted for 29% of the genome and presumably played major roles in enlargement of the size of the ginseng genome. Fluorescence in situ hybridization (FISH) revealed that the PgDel1 elements are distributed throughout the chromosomes along dispersed heterochromatic regions except for ribosomal DNA blocks. The intensity of the PgDel2 FISH signals was biased toward 24 out of 48 chromosomes. Unique gene probes showed two pairs of signals with different locations, one pair in subtelomeric regions on PgDel2-rich chromosomes and the other in interstitial regions on PgDel2-poor chromosomes, demonstrating allotetraploidy in ginseng. Our findings promote understanding of the evolution of the ginseng genome and of that of related species in the Araliaceae.

Construction of a reference genetic map of Raphanus sativus based on genotyping by whole-genome resequencing.

KEY MESSAGE: This manuscript provides a genetic map of Raphanus sativus that has been used as a reference genetic map for an ongoing genome sequencing project. The map was constructed based on genotyping by whole-genome resequencing of mapping parents and F 2 population. Raphanus sativus is an annual vegetable crop species of the Brassicaceae family and is one of the key plants in the seed industry, especially in East Asia. Assessment of the R. sativus genome provides fundamental resources for crop improvement as well as the study of crop genome structure and evolution. With the goal of anchoring genome sequence assemblies of R. sativus cv. WK10039 whose genome has been sequenced onto the chromosomes, we developed a reference genetic map based on genotyping of two parents (maternal WK10039 and paternal WK10024) and 93 individuals of the F2 mapping population by whole-genome resequencing. To develop high-confidence genetic markers, ~83 Gb of parental lines and ~591 Gb of mapping population data were generated as Illumina 100 bp paired-end reads. High stringent sequence analysis of the reads mapped to the 344 Mb of genome sequence scaffolds identified a total of 16,282 SNPs and 150 PCR-based markers. Using a subset of the markers, a high-density genetic map was constructed from the analysis of 2,637 markers spanning 1,538 cM with 1,000 unique framework loci. The genetic markers integrated 295 Mb of genome sequences to the cytogenetically defined chromosome arms. Comparative analysis of the chromosome-anchored sequences with Arabidopsis thaliana and Brassica rapa revealed that the R. sativus genome has evident triplicated sub-genome blocks and the structure of gene space is highly similar to that of B. rapa. The genetic map developed in this study will serve as fundamental genomic resources for the study of R. sativus.

Transcriptome sequencing of two parental lines of cabbage (Brassica oleracea L. var. capitata L.) and construction of an EST-based genetic map.

BACKGROUND: Expressed sequence tag (EST)-based markers are preferred because they reflect transcribed portions of the genome. We report the development of simple sequence repeat (SSR) and single nucleotide polymorphism (SNP) markers derived from transcriptome sequences in cabbage, and their utility for map construction. RESULTS: Transcriptome sequences were obtained from two cabbage parental lines, C1184 and C1234, which are susceptible and resistant to black rot disease, respectively, using the 454 platform. A total of 92,255 and 127,522 reads were generated and clustered into 34,688 and 40,947 unigenes, respectively. We identified 2,405 SSR motifs from the unigenes of the black rot-resistant parent C1234. Trinucleotide motifs were the most abundant (66.15%) among the repeat motifs. In addition, 1,167 SNPs were detected between the two parental lines. A total of 937 EST-based SSR and 97 SNP-based dCAPS markers were designed and used for detection of polymorphism between parents. Using an F2 population, we built a genetic map comprising 265 loci, and consisting of 98 EST-based SSRs, 21 SNP-based dCAPS, 55 IBP markers derived from B. rapa genome sequence and 91 public SSRs, distributed on nine linkage groups spanning a total of 1,331.88 cM with an average distance of 5.03 cM between adjacent loci. The parental lines used in this study are elite breeding lines with little genetic diversity; therefore, the markers that mapped in our genetic map will have broad spectrum utility. CONCLUSIONS: This genetic map provides additional genetic information to the existing B. oleracea map. Moreover, the new set of EST-based SSR and dCAPS markers developed herein is a valuable resource for genetic studies and will facilitate cabbage breeding. Additionally, this study demonstrates the usefulness of NGS transcriptomes for the development of genetic maps even with little genetic diversity in the mapping population.

Identification and mapping of a novel dominant resistance gene, TuRB07 to Turnip mosaic virus in Brassica rapa.

KEY MESSAGE: A novel dominant resistance gene, TuRB07, was found to confer resistance to an isolate of TuMV strain C4 in B. rapa line VC1 and mapped on the top of chromosome A06. The inheritance of resistance to Turnip mosaic virus in Brassica rapa was investigated by crossing the resistant line, VC1 with the susceptible line, SR5, and genotyping and phenotyping diverse progenies derived from this cross. Both a doubled haploid population, VCS3M-DH, an F2 and two BC1 (F1 × VC1 and F1 × SR5) populations were created. Population tests revealed that the resistance to the TuMV C4 isolate in B. rapa is controlled by a single dominant gene. This resistance gene, TuRB07 was positioned on the top of linkage group A06 of the B. rapa genome through bulk segregation analysis and fine mapping recombinants in three doubled haploid- and one backcross population using microsatellite markers developed from BAC end sequences. Within the region between the two closely linked markers flanking TuRB07, H132A24-s1, and KS10960, in the Chiifu reference genome, two genes encoding nucleotide-binding site and leucine-rich repeat proteins with a coiled-coil motif (CC-NBS-LRR), Bra018862 and Bra018863 were identified as candidate resistance genes. The gene Bra018862 is truncated, but the gene Bra018863 has all the domains to function. Furthermore, the analysis of structural variation using resequencing data of VC1 and SR5 revealed that Bra018863 might be a functional gene because the gene has no structural variation in the resistant line VC1 when compared with Chiifu, whereas at the other NBS-LRR genes large deletions were identified in the resistant line. Allelic differences of Bra018863 were found between VC1 and SR5, supporting the notion that this gene is a putative candidate gene for the virus resistance.

Comparative analysis of the radish genome based on a conserved ortholog set (COS) of Brassica.

KEY MESSAGE: This manuscript provides a Brassica conserved ortholog set (COS) that can be used as diagnostic cross-species markers as well as tools for genetic mapping and genome comparison of the Brassicaceae. A conserved ortholog set (COS) is a collection of genes that are conserved in both sequence and copy number between closely related genomes. COS is a useful resource for developing gene-based markers and is suitable for comparative genome mapping. We developed a COS for Brassica based on proteome comparisons of Arabidopsis thaliana, B. rapa, and B. oleracea to establish a basis for comparative genome analysis of crop species in the Brassicaceae. A total of 1,194 conserved orthologous single-copy genes were identified from the genomes based on whole-genome BLASTP analysis. Gene ontology analysis showed that most of them encoded proteins with unknown function and chloroplast-related genes were enriched. In addition, 152 Brassica COS primer sets were applied to 16 crop and wild species of the Brassicaceae and 57.9-92.8 % of them were successfully amplified across the species representing that a Brassica COS can be used as diagnostic cross-species markers of diverse Brassica species. We constructed a genetic map of Raphanus sativus by analyzing the segregation of 322 COS genes in an F2 population (93 individuals) of Korean cultivars (WK10039 × WK10024). Comparative genome analysis based on the COS genes showed conserved genome structures between R. sativus and B. rapa with lineage-specific rearrangement and fractionation of triplicated subgenome blocks indicating close evolutionary relationship and differentiation of the genomes. The Brassica COS developed in this study will play an important role in genetic, genomic, and breeding studies of crop Brassicaceae species.

Comparative analysis of personal protective equipment nonadherence detection: computer vision versus human observers.

OBJECTIVES: Human monitoring of personal protective equipment (PPE) adherence among healthcare providers has several limitations, including the need for additional personnel during staff shortages and decreased vigilance during prolonged tasks. To address these challenges, we developed an automated computer vision system for monitoring PPE adherence in healthcare settings. We assessed the system performance against human observers detecting nonadherence in a video surveillance experiment. MATERIALS AND METHODS: The automated system was trained to detect 15 classes of eyewear, masks, gloves, and gowns using an object detector and tracker. To assess how the system performs compared to human observers in detecting nonadherence, we designed a video surveillance experiment under 2 conditions: variations in video durations (20, 40, and 60 seconds) and the number of individuals in the videos (3 versus 6). Twelve nurses participated as human observers. Performance was assessed based on the number of detections of nonadherence. RESULTS: Human observers detected fewer instances of nonadherence than the system (parameter estimate -0.3, 95% CI -0.4 to -0.2, P < .001). Human observers detected more nonadherence during longer video durations (parameter estimate 0.7, 95% CI 0.4-1.0, P < .001). The system achieved a sensitivity of 0.86, specificity of 1, and Matthew's correlation coefficient of 0.82 for detecting PPE nonadherence. DISCUSSION: An automated system simultaneously tracks multiple objects and individuals. The system performance is also independent of observation duration, an improvement over human monitoring. CONCLUSION: The automated system presents a potential solution for scalable monitoring of hospital-wide infection control practices and improving PPE usage in healthcare settings.

Characterization of a new high copy Stowaway family MITE, BRAMI-1 in Brassica genome.

BACKGROUND: Miniature inverted-repeat transposable elements (MITEs) are expected to play important roles in evolution of genes and genome in plants, especially in the highly duplicated plant genomes. Various MITE families and their roles in plants have been characterized. However, there have been fewer studies of MITE families and their potential roles in evolution of the recently triplicated Brassica genome. RESULTS: We identified a new MITE family, BRAMI-1, belonging to the Stowaway super-family in the Brassica genome. In silico mapping revealed that 697 members are dispersed throughout the euchromatic regions of the B. rapa pseudo-chromosomes. Among them, 548 members (78.6%) are located in gene-rich regions, less than 3 kb from genes. In addition, we identified 516 and 15 members in the 470 Mb and 15 Mb genomic shotgun sequences currently available for B. oleracea and B. napus, respectively. The resulting estimated copy numbers for the entire genomes were 1440, 1464 and 2490 in B. rapa, B. oleracea and B. napus, respectively. Concurrently, only 70 members of the related Arabidopsis ATTIRTA-1 MITE family were identified in the Arabidopsis genome. Phylogenetic analysis revealed that BRAMI-1 elements proliferated in the Brassica genus after divergence from the Arabidopsis lineage. MITE insertion polymorphism (MIP) was inspected for 50 BRAMI-1 members, revealing high levels of insertion polymorphism between and within species of Brassica that clarify BRAMI-1 activation periods up to the present. Comparative analysis of the 71 genes harbouring the BRAMI-1 elements with their non-insertion paralogs (NIPs) showed that the BRAMI-1 insertions mainly reside in non-coding sequences and that the expression levels of genes with the elements differ from those of their NIPs. CONCLUSION: A Stowaway family MITE, named as BRAMI-1, was gradually amplified and remained present in over than 1400 copies in each of three Brassica species. Overall, 78% of the members were identified in gene-rich regions, and it is assumed that they may contribute to the evolution of duplicated genes in the highly duplicated Brassica genome. The resulting MIPs can serve as a good source of DNA markers for Brassica crops because the insertions are highly dispersed in the gene-rich euchromatin region and are polymorphic between or within species.

Mapping quantitative trait loci for tissue culture response in VCS3M-DH population of Brassica rapa.

KEY MESSAGE: Quantitative trait loci (QTL) controlling callus induction and plant regeneration were identified in the VCS3M-DH population of Brassica rapa. Quantitative trait loci (QTL) controlling callus induction and plant regeneration were identified in the VCS3M-DH population of Brassica rapa. The VCS3M-DH population showed wide and continuous variation in callus induction and shoot regeneration. Significant coefficient correlations were detected between these two parameters. Broad-sense heritability (h (2)) for the two traits was around 0.7, indicating genetic regulation of regeneration ability in this population. In the composite interval mapping analysis, two QTLs for callus induction ability, qCi2 and qCi7, were mapped on chromosome A02 and A07, explaining 28.6 % of phenotypic variation. For plant regeneration, four QTLs, qPr6-1 qPr6-2, qPr7, and qPr9 were identified on chromosome A06, A07, and A09, which in total explained 50.1 % of phenotypic variation. Furthermore, 15 putative candidate genes were found on the interval of the six QTLs, which were related to various plant hormones, MADS-box genes, and serine/threonine related genes. These results provide important information to identify genes related to tissue culture ability in B. rapa.

Abscess nodule of the posterior compartment of the knee.

A variety of masses in the knee must be considered in a differential diagnosis when swelling, pain, and a limitation of motion occur with no history of specific trauma. These lesions are most commonly related to the synovial lining of the joint capsule and the cruciate ligaments. The posterior compartment is an unusual location for solitary lesions of the knee joint. There have been few cases of abscess presenting as an intra-articular nodular mass. We report herein a case of a 15-year-old girl who had synovitis and an abscess nodule of the posterior compartment behind the posterior cruciate ligament in the knee joint. An arthroscopic synovectomy and mass excision along with antibiotic administration was curative in our case.

Purine nucleosidase (PNase) activity, probiotics potential, and food applicability of a newly-isolated Levilactobacillus brevis LAB42.

Hyperuricemia, a condition characterized by elevated levels of uric acid in the blood, is known as a risk factor for gout disease. In this study, we isolated a total of 72 MRS-grown colonies and evaluated their purine nucleosidase (PNase) activity. Among the isolated bacteria, Levilactobacillus (L.) brevis LAB42 displayed the highest PNase activity. Our findings also indicate that PNase activity can vary among lactic acid bacterial strains and during different growth phases. Based on the kinetics study, LAB42 consistently exhibits the highest PNase activity. Due to its ability to attach to Caco-2 cells and its resistance to acidic environments and bile exposure, L. brevis LAB42 was chosen for further studies and showed that with the right combination of additives, it has the potential to be an appropriate starter for milk fermentation.