SEAL: a distributed short read mapping and duplicate removal tool

Summary: SEAL is a scalable tool for short read pair mapping and duplicate removal. It computes mappings that are consistent with those produced by BWA and removes duplicates according to the same criteria employed by Picard MarkDuplicates. On a 16-node Hadoop cluster, it is capable of processing about 13 GB per hour in map+rmdup mode, while reaching a throughput of 19 GB per hour in mapping-only mode

Circadian clock components control daily growth activities by modulating cytokinin levels and cell division-associated gene expression in <i>Populus</i> trees

Trees are carbon dioxide sinks and major producers of terrestrial biomass with distinct seasonal growth patterns. Circadian clocks enable the coordination of physiological and biochemical temporal activities, optimally regulating multiple traits including growth. To dissect the clock's role in growth, we analysed Populus tremula x P. tremuloides trees with impaired clock function due to down-regulation of central clock components. late elongated hypocotyl (lhy-10) trees, in which expression of LHY1 and LHY2 is reduced by RNAi, have a short free-running period and show disrupted temporal regulation of gene expression and reduced growth, producing 30-40% less biomass than wild-type trees. Genes important in growth regulation were expressed with an earlier phase in lhy-10, and CYCLIN D3 expression was misaligned and arrhythmic. Levels of cytokinins were lower in lhy-10 trees, which also showed a change in the time of peak expression of genes associated with cell division and growth. However, auxin levels were not altered in lhy-10 trees, and the size of the lignification zone in the stem showed a relative increase. The reduced growth rate and anatomical features of lhy-10 trees were mainly caused by misregulation of cell division, which may have resulted from impaired clock function

Iterative Correction of Reference Nucleotides (iCORN) using second generation sequencing technology

Motivation: The accuracy of reference genomes is important for downstream analysis but a low error rate requires expensive manual interrogation of the sequence. Here, we describe a novel algorithm (Iterative Correction of Reference Nucleotides) that iteratively aligns deep coverage of short sequencing reads to correct errors in reference genome sequences and evaluate their accuracy

Summarizing and correcting the GC content bias in high-throughput sequencing

GC content bias describes the dependence between fragment count (read coverage) and GC content found in Illumina sequencing data. This bias can dominate the signal of interest for analyses that focus on measuring fragment abundance within a genome, such as copy number estimation (DNA-seq). The bias is not consistent between samples; and there is no consensus as to the best methods to remove it in a single sample. We analyze regularities in the GC bias patterns, and find a compact description for this unimodal curve family. It is the GC content of the full DNA fragment, not only the sequenced read, that most influences fragment count. This GC effect is unimodal: both GC-rich fragments and AT-rich fragments are underrepresented in the sequencing results. This empirical evidence strengthens the hypothesis that PCR is the most important cause of the GC bias. We propose a model that produces predictions at the base pair level, allowing strand-specific GC-effect correction regardless of the downstream smoothing or binning. These GC modeling considerations can inform other high-throughput sequencing analyses such as ChIP-seq and RNA-seq

Utility of early circulating tumour DNA dynamics as a surrogate for progression free survival in the BEECH phase I/II trial in metastatic breast cancer

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Hubungan Asupan Protein, Seng, Zat Besi, Dan Riwayat Penyakit Infeksi Dengan Z-score Tb/u Pada Balita

Latar Belakang : Masalah gizi yang paling banyak ditemukan pada anak di Indonesia adalah stunting, Indikator untuk menilai stunting berdasarkan pada Indeks Tinggi Badan menurut Umur (TB/U) dengan ambang batas (Z-score) &lt;-2 Standart Deviasi (SD). Several micronutrients are required for adequate growth among children. However, it has been unclear as to which nutrient deficiencies contribute most often to growth faltering in populations at risk for poor nutrition and poor growth. Inadequate intakes of dietary energy and protein and frequent infections are well-known causes of growth retardation (3–5). However, the role of specific micronutrient deficiencies in the etiology of growth retardation has gained attention more recently (6–8). Tujuan : Mengetahui hubungan antara asupan protein, seng, zat besi, dan penyakit infeksi terhadap indeks z-score TB/U pada Balita usia 24-59 bulan.Metode : Penelitian observasional dengan pendekatan cross sectional di Kelurahan Jangli Semarang, jumlah sampel 61 Balita usia 24-59 bulan, dipilih dengan simple random sampling. Data yang dikumpulkan meliputi: identitas sampel, berat badan, tinggi badan, riwayat asupan makan, dan riwayat penyakit infeksi. Berat badan diukur menggunakan timbangan digital dan tinggi badan diukur menggunakan microtoise. Asupan protein, seng, zat besi, dan riwayat penyakit infeksi diperoleh dari food frequency questionairre semi-kuantitatif. Data dianalisis dengan uji analisis depskripsi, analisis bivariate menggunakan uji Chi Square, Pearson, dan Spearman.Hasil : Sebanyak 36,1 subjek mengalami stunting. Rerata z-score TB/U -1,25 ± 1,2. Rerata asupan protein, seng, dan zat besi subjek berturut-turut 34.8 ± 13 g, 5.2 ± 2.5 mg, 8.2 ± 6.5 mg dengan sebagian besar tingkat kecukupan protein, seng, dan zat besi subjek adalah cukup. Sebanyak 29.1% subjek memiliki riwayat infeksi. Terdapat hubungan antara protein dan penyakit infeksi dengan z-score TB/U pada Balita. Tidak terdapat hubungan antara asupan seng, dan zat besi dengan z-score TB/U pada Balita. Simpulan : Terdapat hubungan antara asupan protein dan riwayat penyakit infeksi terhadap indeks z-score TB/U pada Balita

A Modified Method for Whole Exome Resequencing from Minimal Amounts of Starting DNA

Next generation DNA sequencing (NGS) technologies have revolutionized the pace at which whole genome and exome sequences can be generated. However, despite these advances, many of the methods for targeted resequencing, such as the generation of high-depth exome sequences, are somewhat limited by the relatively large amounts of starting DNA that are normally required. In the case of tumour analysis this is particularly pertinent as many tumour biopsies often return submicrogram quantities of DNA, especially when tumours are microdissected prior to analysis. Here, we present a method for exome capture and resequencing using as little as 50 ng of starting DNA. The sequencing libraries generated by this minimal starting amount (MSA-Cap) method generate datasets that are comparable to standard amount (SA) whole exome libraries that use three micrograms of starting DNA. This method, which can be performed in most laboratories using commonly available reagents, has the potential to enhance large scale profiling efforts such as the resequencing of tumour exomes

Efficient Genotyping of KRAS Mutant Non-Small Cell Lung Cancer Using a Multiplexed Droplet Digital PCR Approach

© 2015 Pender et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.Droplet digital PCR (ddPCR) can be used to detect low frequency mutations in oncogenedriven lung cancer. The range of KRAS point mutations observed in NSCLC necessitates a multiplex approach to efficient mutation detection in circulating DNA. Here we report the design and optimisation of three discriminatory ddPCR multiplex assays investigating nine different KRAS mutations using PrimePCRddPCRMutation Assays and the Bio-Rad QX100 system. Together these mutations account for 95% of the nucleotide changes found in KRAS in human cancer. Multiplex reactions were optimised on genomic DNA extracted from KRAS mutant cell lines and tested on DNA extracted from fixed tumour tissue from a cohort of lung cancer patients without prior knowledge of the specific KRAS genotype. The multiplex ddPCR assays had a limit of detection of better than 1 mutant KRAS molecule in 2,000 wild-type KRAS molecules, which compared favourably with a limit of detection of 1 in 50 for next generation sequencing and 1 in 10 for Sanger sequencing. Multiplex ddPCR assays thus provide a highly efficient methodology to identify KRAS mutations in lung adenocarcinoma

A method for counting PCR template molecules with application to next-generation sequencing

Amplification by polymerase chain reaction is often used in the preparation of template DNA molecules for next-generation sequencing. Amplification increases the number of available molecules for sequencing but changes the representation of the template molecules in the amplified product and introduces random errors. Such changes in representation hinder applications requiring accurate quantification of template molecules, such as allele calling or estimation of microbial diversity. We present a simple method to count the number of template molecules using degenerate bases and show that it improves genotyping accuracy and removes noise from PCR amplification. This method can be easily added to existing DNA library preparation techniques and can improve the accuracy of variant calling