Caracterización química y cuantificación del rendimiento de extracción de pigmento en siete accesiones mexicanas de Bixa orellana

Achiote (Bixa orellana) is a plant used for obtaining a natural dye rich on carotenoids (mainly bixin and norbixin); it is also the plant species with the highest content of tocotrienols in nature. In the present work, the pigment extraction yield of seven Mexican accessions of Bixa orellana was quantified. Also color parameters and content of tocotrienols, tocopherols, norbixin, bixin, total phenolic compounds and antioxidant capacity were evaluated in the corresponding annatto extracts. The highest percentage of pigment extraction yield was obtained with KOH (4.84%). Accessions 43 (L*= 4.01 ± 0.79, C*= 7.33 ± 1.07, h= 25.76 ± 6.35) and 50 (L*= 3.17 ± 0.64, C*= 6.81 ± 0.53, h= 26.41 ± 4.41) had the lowest color values, meaning these accessions had a darker and redder color. Four accessions showed the highest content of bixin: accession 48 (3.1%), 45 (2.6%) 43 (2.4%) and 47 (2.2%). Accession 50 had showed the highest content of total phenolic compounds and of tocotrienols (T3), mainly the isoform δ-T3 (5.03 ± 0.64 mg g−1 Seed Dry Weight), as well as the highest antioxidant capacity.El achiote (Bixa orellana) es una planta utilizada para obtener un colorante natural rico en carotenoides (principalmente bixina y norbixina); además, es la especie vegetal con el mayor contenido de tocotrienoles. En este trabajo, se determinó el rendimiento de extracción de pigmento de siete accesiones mexicanas de Bixa orellana. También se evaluaron los parámetros de color y el contenido de tocotrienoles, tocoferoles, norbixina, bixina, compuestos fenólicos totales y la capacidad antioxidante en extractos de annato. El mayor porcentaje de rendimiento de extracción de pigmento fue obtenido con KOH (4.847905%). Las accesiones 43 (L*= 4.01 ± 0.79, C*= 7.33 ± 1.07, h= 25.76 ± 6.35) y 50 (L*= 3.17 ± 0.64, C*= 6.81 ± 0.53, h= 26.41 ± 4.41) presentaron los valores más bajos de los párametros de color, lo que significa que estas accesiones tuvieron un color más oscuro y más rojo. Cuatro accesiones mostraron el mayor contenido de bixina: accesión 48 (3.1%), 45 (2.6%) 43 (2.4%) and 47 (2.2%). La accession 50 mostró el mayor contenido de compuestos fenólicos totales y de tocotrienoles (T3), principalmente la isoforma δ-T3 (5.03 ± 0.64 mg g-1 Peso Seco), así como también la mayor capacidad antioxidante.Fil: Raddatz Mota, D.. Universidad Autónoma Metropolitana; MéxicoFil: Pérez Flores, L. J.. Universidad Autónoma Metropolitana; MéxicoFil: Carrari, Fernando Oscar. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigación en Ciencias Veterinarias y Agronómicas. Instituto de Biotecnología; ArgentinaFil: Insani, Ester Marina. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigación en Ciencias Veterinarias y Agronómicas. Instituto de Biotecnología; ArgentinaFil: Asis, Ramón. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Córdoba. Centro de Investigaciones en Bioquímica Clínica e Inmunología; ArgentinaFil: Mendoza Espinoza, J. A.. Universidad Nacional Autónoma de México; MéxicoFil: Díaz de León Sánchez, F.. Universidad Autónoma Metropolitana; MéxicoFil: Rivera Cabrera, F.. Universidad Autónoma Metropolitana; Méxic

Dnmt2-dependent methylomes lack defined DNA methylation patterns

Several organisms have retained methyltransferase 2 (Dnmt2) as their only candidate DNA methyltransferase gene. However, information about Dnmt2-dependent methylation patterns has been limited to a few isolated loci and the results have been discussed controversially. In addition, recent studies have shown that Dnmt2 functions as a tRNA methyltransferase, which raised the possibility that Dnmt2-only genomes might be unmethylated. We have now used whole-genome bisulfite sequencing to analyze the methylomes of Dnmt2-only organisms at single-base resolution. Our results show that the genomes of Schistosoma mansoni and Drosophila melanogaster lack detectable DNA methylation patterns. Residual unconverted cytosine residues shared many attributes with bisulfite deamination artifacts and were observed at comparable levels in Dnmt2-deficient flies. Furthermore, genetically modified Dnmt2-only mouse embryonic stem cells lost the DNA methylation patterns found in wild-type cells. Our results thus uncover fundamental differences among animal methylomes and suggest that DNA methylation is dispensable for a considerable number of eukaryotic organisms

NIPTeR:an R package for fast and accurate trisomy prediction in non-invasive prenatal testing

BACKGROUND: Various algorithms have been developed to predict fetal trisomies using cell-free DNA in non-invasive prenatal testing (NIPT). As basis for prediction, a control group of non-trisomy samples is needed. Prediction accuracy is dependent on the characteristics of this group and can be improved by reducing variability between samples and by ensuring the control group is representative for the sample analyzed.RESULTS: NIPTeR is an open-source R Package that enables fast NIPT analysis and simple but flexible workflow creation, including variation reduction, trisomy prediction algorithms and quality control. This broad range of functions allows users to account for variability in NIPT data, calculate control group statistics and predict the presence of trisomies.CONCLUSION: NIPTeR supports laboratories processing next-generation sequencing data for NIPT in assessing data quality and determining whether a fetal trisomy is present. NIPTeR is available under the GNU LGPL v3 license and can be freely downloaded from https://github.com/molgenis/NIPTeR or CRAN.</p

Targeted RNA-Sequencing Enables Detection of Relevant Translocations and Single Nucleotide Variants and Provides a Method for Classification of Hematological Malignancies-RANKING

BACKGROUND: Patients with hematological malignancies (HMs) carry a wide range of chromosomal and molecular abnormalities that impact their prognosis and treatment. Since no current technique can detect all relevant abnormalities, technique(s) are chosen depending on the reason for referral, and abnormalities can be missed. We tested targeted transcriptome sequencing as a single platform to detect all relevant abnormalities and compared it to current techniques. MATERIAL AND METHODS: We performed RNA-sequencing of 1385 genes (TruSight RNA Pan-Cancer, Illumina) in bone marrow from 136 patients with a primary diagnosis of HM. We then applied machine learning to expression profile data to perform leukemia classification, a method we named RANKING. Gene fusions for all the genes in the panel were detected, and overexpression of the genes EVI1, CCND1, and BCL2 was quantified. Single nucleotide variants/indels were analyzed in acute myeloid leukemia (AML), myelodysplastic syndrome and patients with acute lymphoblastic leukemia (ALL) using a virtual myeloid (54 genes) or lymphoid panel (72 genes). RESULTS: RANKING correctly predicted the leukemia classification of all AML and ALL samples and improved classification in 3 patients. Compared to current methods, only one variant was missed, c.2447A>T in KIT (RT-PCR at 10(-4)), and BCL2 overexpression was not seen due to a t(14; 18)(q32; q21) in 2% of the cells. Our RNA-sequencing method also identified 6 additional fusion genes and overexpression of CCND1 due to a t(11; 14)(q13; q32) in 2 samples. CONCLUSIONS: Our combination of targeted RNA-sequencing and data analysis workflow can improve the detection of relevant variants, and expression patterns can assist in establishing HM classification

CoNVaDING:Single Exon Variation Detection in Targeted NGS Data

We have developed a tool for detecting single exon copy number variations (CNVs) in targeted next-generation sequencing data: CoNVaDING (Copy Number Variation Detection In Next-generation sequencing Gene panels). CoNVaDING includes a stringent quality control metric, that excludes or flags low quality exons. Since this quality control shows exactly which exons can be reliably analysed and which exons are in need of an alternative analysis method, CoNVaDING is not only useful for CNV detection in a research setting, but also in clinical diagnostics. During the validation phase, CoNVaDING detected all known CNVs in high quality targets in 320 samples analysed, giving 100% sensitivity and 99.998% specificity for 308,574 exons. CoNVaDING outperforms existing tools by exhibiting a higher sensitivity and specificity and by precisely identifying low-quality samples and regions. This article is protected by copyright. All rights reserved.</p

Feasibility of Follow-Up Studies and Reclassification in Spinocerebellar Ataxia Gene Variants of Unknown Significance

Spinocerebellar ataxia (SCA) is a heterogeneous group of neurodegenerative disorders with autosomal dominant inheritance. Genetic testing for SCA leads to diagnosis, prognosis and risk assessment for patients and their family members. While advances in sequencing and computing technologies have provided researchers with a rapid expansion in the genetic test content that can be used to unravel the genetic causes that underlie diseases, the large number of variants with unknown significance (VUSes) detected represent challenges. To minimize the proportion of VUSes, follow-up studies are needed to aid in their reclassification as either (likely) pathogenic or (likely) benign variants. In this study, we addressed the challenge of prioritizing VUSes for follow-up using (a combination of) variant segregation studies, 3D protein modeling, in vitro splicing assays and functional assays. Of the 39 VUSes prioritized for further analysis, 13 were eligible for follow up. We were able to reclassify 4 of these VUSes to LP, increasing the molecular diagnostic yield by 1.1%. Reclassification of VUSes remains difficult due to limited possibilities for performing variant segregation studies in the classification process and the limited availability of routine functional tests

The effect of cigarette smoke exposure on the development of inflammation in lungs, gut and joints of TNFΔARE mice

The inflammatory cytokine TNF-alpha is a central mediator in many immune-mediated diseases, such as Crohn's disease (CD), spondyloarthritis (SpA) and chronic obstructive pulmonary disease (COPD). Epidemiologic studies have shown that cigarette smoking (CS) is a prominent common risk factor in these TNF-dependent diseases. We exposed TNF Delta ARE mice; in which a systemic TNF-alpha overexpression leads to the development of inflammation; to 2 or 4 weeks of air or CS. We investigated the effect of deregulated TNF expression on CS-induced pulmonary inflammation and the effect of CS exposure on the initiation and progression of gut and joint inflammation. Upon 2 weeks of CS exposure, inflammation in lungs of TNF Delta ARE mice was significantly aggravated. However, upon 4 weeks of CS-exposure, this aggravation was no longer observed. TNF Delta ARE mice have no increases in CD4+ and CD8+ T cells and a diminished neutrophil response in the lungs after 4 weeks of CS exposure. In the gut and joints of TNF Delta ARE mice, 2 or 4 weeks of CS exposure did not modulate the development of inflammation. In conclusion, CS exposure does not modulate gut and joint inflammation in TNF Delta ARE mice. The lung responses towards CS in TNF Delta ARE mice however depend on the duration of CS exposure