Total Sequence Length Research Articles

The complete mitochondrial genome of Rapana venosa (Gastropoda, Muricidae)

The complete mitochondrial (mt) genome of the veined rapa whelk, Rapana venosa, was determined using genome walking techniques in this study. The total length of the mt genome sequence of R. venosa was 15,271 bp, which is comparable to the reported Muricidae mitogenomes to date. It contained 13 protein-coding genes, 21 transfer RNA genes, and two ribosomal RNA genes. A bias towards a higher representation of nucleotides A and T (69%) was detected in the mt genome of R. venosa. A small number of non-coding nucleotides (302 bp) was detected, and the largest non-coding region was 74 bp in length.

The complete mitochondrial genome of cultivated radish WK10039 (Raphanus sativus L.)

We determined the complete nucleotide sequence of the mitochondrial genome of radish cultivar WK10039 (Raphanus sativus L.). The total length of the mtDNA sequence is 244,054 bp, with GC content of 45.3%. The radish mtDNA contains 82 protein-coding genes, 17 tRNA genes, and 3 rRNA genes. Among the protein-coding genes, 34 encode proteins with known functions. There are two 5529 bp repeats in the radish mitochondrial genome that may contribute to DNA recombination resulting in at least three different forms of mtDNA in radish.

English

In the present study, molecular phylogeny of bivalve family Veneridae (Mollusca:Bivalvia) was analysed using internal transcribed spacer (ITS) region of 21 species belonging to different subfamilies of Veneridae. ITS of ribosomal DNA can be utilised for delineating evolu- tionary and genetic relationships between closely related taxa. ITS region of Paphia malabarica belonging to subfamily Tapetinae and Meretrix casta belonging to meretricinae was sequenced. Total genomic DNA was extracted from the adductor muscle using CTAB protocol and the internal transcribed spacer region of nuclear ribosomal DNA was PCR amplified and sequenced using ITS (ITS1 and ITS2) forward and re- verse primers. Total length of sequence was found to be 895 bp in Paphia malabarica and 785 bp in Meretrix casta. GC contents in the se- quences were found to be 58.99% and 64.68% respectively in Paphia malabarica and Meretrix casta. ITS1 region of Paphia malabarica con- sisted of 393 bp with GC content 58.12% and 309 bp with 63.75% GC content in Meretrix casta. ITS2 region generated 248bp with 63% GC content in Paphia malabarica and 240bp with 72.06% GC content in Meretrix casta. 5.8S region rendered 157bp with 60% GC content in Pa- phia malabarica and 138bp with 61.32% GC content in Meretrix casta. Phylogenetic trees were constructed using Neighbor-Joining and Maxi- mum-Parsimony methods. The analysis produced trees with similar topology. The tree topologies indicated two clades with three well resolved groups. Phylogenetic analysis supported the morphological classification and monophyly of the family Veneridae.

Complete mitochondrial genome of Ostrea denselamellosa (Bivalvia, Ostreidae)

The complete mitochondrial (mt) genome of the flat oyster, Ostrea denselamellosa, was determined using Long-PCR and genome walking techniques in this study. The total length of the mt genome sequence of O. denselamellosa was 16,227 bp, which is the smallest reported Ostreidae mt genome to date. It contained 12 protein-coding genes (lacking of ATP8), 23 transfer RNA genes, and two ribosomal RNA genes. A bias towards a higher representation of nucleotides A and T (60.7%) was detected in the mt genome of O. denselamellosa. The rrnL was split into two fragments (3′ half, 711 bp; 5′ half, 509 bp), which seems to be the unique characteristics of Ostreidae mt genomes.

First Report of Watermelon mosaic virus Naturally Infecting Cucumis anguria.

Watermelon mosaic virus (WMV, Potyvirus) is an important virus of cucurbit crops worldwide, although it is rarely found in tropical regions. Cucumis anguria L. is a native species from tropical regions of Africa and can be found as a volunteer or cultivated plant in the Neotropic (1). In Venezuela, C. anguria is a ubiquitous volunteer cucurbit that grows in crop fields year round and may serve as reservoir for viruses. In 2009 and 2010, during surveys for viruses in cultivated and wild cucurbits, WMV was found at a low frequency (3 out of 284 samples) and two out of three positive samples were C. anguria. One of these samples was used to mechanically inoculate WMV to melon (Cucumis melo L.) and zucchini squash (Cucurbita pepo L.) plants. Typical symptoms of WMV were observed 2 weeks after inoculation and viral identification was confirmed by double antibody sandwich (DAS)-ELISA. Based on whole genome sequences, three phylogenetic groups of WMV, namely groups G1 to G3, are defined (2). Since molecular characterization of this virus is scarce in South America, the complete sequence of a WMV isolate recovered from C. anguria (hereafter, VE10-99) was obtained. RNA extractions, amplification procedures, and sequencing analyses were performed according to Desbiez and Lecoq (2). The total sequence length of VE10-99 isolate was 10,039 nucleotides, excluding the polyadenylated tail (GenBank Accession No. KC292915). This isolate had the typical potyviral genome organization, exhibiting a unique large ORF with 9 putative cleavage sites. In a BLAST analysis, the isolate most closely related to VE10-99 was the Chilean isolate CHI87-620 (EU660580), sharing 96% nucleotide identity. Phylogenetic analyses showed that VE10-99 belongs to group G2 of WMV. No evidence for recombination was found in the genome of VE10-99. Although recombination events have been noticed between members of G1 and G3, recombinant isolates between members of G2 and G1 are more frequent (2). In fact, the isolate CHI87-620 had been the only bona fide member of G2 until the sequencing of VE10-99. G2/G1 recombinants seem to have almost completely replaced the parental isolates throughout the world, probably due to a better fitness (2). To our knowledge, WMV natural infections in C. anguria had not been described before. The finding of a bona fide G2 member in Venezuela raises the question about the origin of G2 group. Currently, the prevalence of WMV appears reduced compared to the previous survey performed in Venezuela in 1966 (16 WMV-positive plants out of 95 samples) (3).

An Analysis of the Athetis lepigone Transcriptome from Four Developmental Stages

Athetis lepigone Möschler (Lepidoptera: Noctuidae) has recently become an important insect pest of maize (Zea mays) crops in China. In order to understand the characteristics of the different developmental stages of this pest, we used Illumina short-read sequences to perform de novo transcriptome assembly and gene expression analysis for egg, larva, pupa and adult developmental stages. We obtained 10.08 Gb of raw data from Illumina sequencing and recovered 81,356 unigenes longer than 100 bp through a de novo assembly. The total sequence length reached 49.75 Mb with 858 bp of N50 and an average unigene length of 612 bp. Annotation analysis of predicted proteins indicate that 33,736 unigenes (41.47% of total unigenes) are matches to genes in the Genbank Nr database. The unigene sequences were subjected to GO, COG and KEGG functional classification. A large number of differentially expressed genes were recovered by pairwise comparison of the four developmental stages. The most dramatic differences in gene expression were found in the transitions from one stage to another stage. Some of these differentially expressed genes are related to cuticle and wing formation as well as the growth and development. We identified more than 2,500 microsatellite markers that may be used for population studies of A. lepigone. This study lays the foundation for further research on population genetics and gene function analysis in A. lepigone.

Complete mitochondrial genome of the spotted scat Scatophagus argus (Teleostei, Scatophagidae)

The complete mitochondrial sequence of the spotted scat Scatophagus argus has been determined using long amplification polymerase chain reaction (LA-PCR). The total length of sequence is 16,778 bp, and includes 13 protein-coding genes, 2 ribosomal RNA genes, 22 transfer RNA genes and 1 control region. The base composition of H-strand is 27.23% (A), 27.54% (T), 16.22% (G) and 28.81% (C), with an AT content of 55.08%. The arrangement of genes in S. argus is identical to that of other fish species. All genes are encoded on the heavy strand with the exception of ND6 and eight tRNA genes. The mitochondrial genome of S. argus presented here will contribute to resolve phylogenetic relationships within the family Scatophagidae and the Perciformes.

Interaction of YOYO-1 with guanine-rich DNA

The oxazole homodimer YOYO-1 has served as a valuable tool for the detection and quantification of nucleic acids. While the base specificity and selectivity of binding of YOYO-1 has been researched to some extent, the effect of unorthodox nucleic acid conformations on dye binding has received relatively less attention. In this work, we attempt to correlate the quadruplex-forming ability of G-rich sequences with binding of YOYO-1. Oligonucleotides differing in the number of tandem G repeats, total length, and length of loop sequence were evaluated for their ability to form quadruplexes in presence of sodium (Na+) or potassium (K+) ions. The fluorescence behavior of YOYO-1 upon binding such G-rich sequences was also ascertained. A distinct correlation was observed between the strength and propensity of quadruplex formation, and the affinity of YOYO-1 to bind such sequences. Specifically, as exemplified by the oligonucleotides 5′-G4T2G4-3′ and 5′-G3TG3TG3-3′, sequences possessing longer G-rich regions and shorter loop sequences formed stronger quadruplexes in presence of K+ which translated to weaker binding of YOYO-1. The dependence of binding of YOYO-1 on sequence and structural features of G-rich DNA has not been explored previously and such studies are expected to aid in more effective interpretation of applications involving the fluorophore.

Broadband homodecoupled NMR spectroscopy with enhanced sensitivity

A new type of broadband homodecoupling technique is described, which is based on the original version of the Zangger–Sterk experiment, but results in a spectrum with higher sensitivity. The homodecoupling is performed by a combination of selective and non-selective 180° RF pulses in the presence of weak rectangular pulsed field gradients in a pseudo 2D experiment. The proposed experiment uses a fast pulsing approach to increase the signal-to-noise ratio per unit time. The recycle delay is significantly shortened typically to about 100ms. After each scan, the offset of the selective shaped pulse is changed to access fresh magnetisation from adjacent frequency/spatial regions. The physical acquisition time was limited to 40ms to keep the total length of the pulse sequence as short as possible. Broadband inversion BIP pulses are used instead of 180° hard pulses. They are used pairwise to cancel out unwanted phase shifts over the bandwidth. Reconstruction of the homodecoupled spectrum was done by concatenating the first 10ms of the FID from each single increment to obtain the final homodecoupled proton FID followed by Fourier transformation.The new method can either be used to acquire broadband homodecoupled spectra in a shorter time or to increase the signal-to-noise ratio compared to the original Zangger–Sterk experiment. Using eight different frequencies can thus lead to a signal to noise gain of a factor √8 or a factor of eight in time.

The complete mitochondrial genome of the Bufo tibetanus (Anura: Bufonidae)

The total length of the Bufo tibetanus mitochondrial genome sequence is 17,405 bp. It consists of 13 protein-coding genes, 2 rRNA genes, 22 tRNA genes and 1 displacement loop (D-loop). Except for eight tRNAs and nd6 genes, all other mitochondrial genes are encoded on the heavy strand (H strand). Most of the 22 tRNA genes can fold into a typical cloverleaf secondary structure by tRNAscan-SE, except for tRNASer. Molecular data presented here provide a useful toll for helping set the stage for further studies and the molecular evolution of the mitochondrial genome.

Prediction of peptides binding to MHC class I and II alleles by temporal motif mining

BackgroundMHC (Major Histocompatibility Complex) is a key player in the immune response of most vertebrates. The computational prediction of whether a given antigenic peptide will bind to a specific MHC allele is important in the development of vaccines for emerging pathogens, the creation of possibilities for controlling immune response, and for the applications of immunotherapy. One of the problems that make this computational prediction difficult is the detection of the binding core region in peptides, coupled with the presence of bulges and loops causing variations in the total sequence length. Most machine learning methods require the sequences to be of the same length to successfully discover the binding motifs, ignoring the length variance in both motif mining and prediction steps. In order to overcome this limitation, we propose the use of time-based motif mining methods that work position-independently.ResultsThe prediction method was tested on a benchmark set of 28 different alleles for MHC class I and 27 different alleles for MHC class II. The obtained results are comparable to the state of the art methods for both MHC classes, surpassing the published results for some alleles. The average prediction AUC values are 0.897 for class I, and 0.858 for class II.ConclusionsTemporal motif mining using partial periodic patterns can capture information about the sequences well enough to predict the binding of the peptides and is comparable to state of the art methods in the literature. Unlike neural networks or matrix based predictors, our proposed method does not depend on peptide length and can work with both short and long fragments. This advantage allows better use of the available training data and the prediction of peptides of uncommon lengths.

Characterization of SNPs in strawberry cultivars in China

Single nucleotide polymorphisms (SNPs) occur at high frequencies in both plant and animal genomes and can provide broad genome coverage and reliable estimates of genetic relationships. The availability of expressed sequence tag (EST) data has made it feasible to discover SNPs. DNA analysis is crucial in genetic studies not only for strawberry breeding programs but also for characterization of hybrids and species. We cloned 96 EST sequences, and 116 SNPs were discovered by comparing 16 strawberry cultivars grown in the region of Nanjing, China. Sequence alignment of 6 group sequences derived from 16 sample cultivars yielded 116 SNPs, within a total genomic sequence length of 1755 bp. The SNPs were discovered with a mean frequency of one SNP per 15 bp. These SNPs were comprised of 57% transitions, 32.7% transversions, 8.6% InDels, and 1.7% others, based on which a phylogenetic tree was constructed. Among the 116 SNPs, 75% were located within the open reading frame (ORF), while 25% were located outside the ORF. All 16 cultivars scattered well in dendrogram derived from the SNP data, demonstrating that SNPs can be a powerful tool for cultivar identification and genetic diversity analysis in strawberries.

DNA Barcode Sequencing from Old Type Specimens as a Tool in Taxonomy: A Case Study in the Diverse Genus Eois (Lepidoptera: Geometridae)

In this study we report on the sequencing of the COI barcode region from 96 historical specimens (92 type specimens +4 non-types) of Eois. Eois is a diverse clade of tropical geometrid moths and is the target of a number of ongoing studies on life-histories, phylogeny, co-evolution with host plants or parasitoids, and diversity patterns across temporal and spatial dimensions. The unequivocal application of valid names is crucial for all aspects of biodiversity research as well as monitoring and conservation efforts. The availability of barcodes from historical type specimens has the potential to facilitate the much-needed acceleration of species description. We performed non-destructive DNA extraction on the abdomens of Eois specimens between 79 and 157 years of age. We used six primer combinations (recovering between 109 and 130 bp each) to target the full-length barcode sequence of each specimen. We were able to obtain sequences for 91 of 96 specimens (success rate 94.8%). Sequence length ranged from 121 bp to full barcode sequences (658 bp), the average sequence length was ∼500 bp. We detected a moderately strong and statistically significant negative correlation between specimen age and total sequence length, which is in agreement with expectations. The abdomen proved to be an exceedingly valuable source of DNA in old specimens of Lepidoptera. Barcode sequences obtained in this study are currently being used in an effort towards a step-wise taxonomic revision of Eois. We encourage that DNA barcodes obtained from types specimens should be included in all species descriptions and revisions whenever feasible.

Modeling judgment of sequentially presented categories using weighting and sampling without replacement

In a series of experiments, Kusev et al. (Journal of Experimental Psychology: Human Perception and Performance 37:1874-1886, 2011) studied relative-frequency judgments of items drawn from two distinct categories. The experiments showed that the judged frequencies of categories of sequentially encountered stimuli are affected by the properties of the experienced sequences. Specifically, a first-run effect was observed, whereby people overestimated the frequency of a given category when that category was the first repeated category to occur in the sequence. Here, we (1) interpret these findings as reflecting the operation of a judgment heuristic sensitive to sequential patterns, (2) present mathematical definitions of the sequences used in Kusev et al. (Journal of Experimental Psychology: Human Perception and Performance 37:1874-1886, 2011), and (3) present a mathematical formalization of the first-run effect-the judgments-relative-to-patterns model-to account for the judged frequencies of sequentially encountered stimuli. The model parameter w accounts for the effect of the length of the first run on frequency estimates, given the total sequence length. We fitted data from Kusev et al. (Journal of Experimental Psychology: Human Perception and Performance 37:1874-1886, 2011) to the model parameters, so that with increasing values of w, subsequent items in the first run have less influence on judgments. We see the role of the model as essential for advancing knowledge in the psychology of judgments, as well as in other disciplines, such as computer science, cognitive neuroscience, artificial intelligence, and human-computer interaction.

Effect of robustness on selection of a mutation-rate regulating gene

We study the evolution of a population of sequences, where each sequence is divided into a reproduction-rate (fitness) encoding part and a mutation-rate regulating part. Evolutionary selection acts on the sequence both by a direct fitness landscape and by indirect selection on a mutation landscape through which the sequence's mutation rate is determined, thereby providing a model of a mutation-rate-regulating gene. Coupling of the fitness landscape and mutation landscape leads to adaptive evolution of the sequence. We investigate the effects of robustness in the mutation landscape and fitness landscapes on selection of the sequence. We find that the effects of robustness in both the mutation and the fitness landscape can be described by an effective sequence length, defined as the mutational load divided by the per-base mutation rate, and we give expressions for the effective sequence length for various fitness and mutation landscapes. The probability that the sequence with a reduced mutation rate evolves is increased by increasing the robustness of the mutation landscape, and decreased by increasing the robustness of the fitness landscape. However, in the case of the mutation-rate-regulating part making up only a very small part of the total sequence length, we show that selection for a more robust sequence with less-reduced mutation rate is very weak, and therefore we conjecture that robust sequences play little role in selection of error-reducing mechanisms in real populations.

Flanking sequence effects on oligonucleotide hybridization

Microarray platforms measure both sequence content and prevalence of sequence by leveraging oligonucleotide hybridization. Computational probe design centers on short regions of high sequence similarity, most often between 25 and 50 base pairs in length. A variety of pattern-matching algorithms specific to the probe length are employed to determine the uniqueness of a target against its background, and a subset of these consider internal structure of the probe as a contra-indicator of successful hybridization. Since it is assumed that the target structure will mirror that of the probe, no consideration is given to the flanking sequences. In this study, we start to examine on the effects of total target sequence length in the context of common hybridization assay conditions. There are two considerations: due to the flexible nature of the single-stranded backbone, loops and bubbles can still yield stable hybridization between species with less than perfect sequence homology; the flanking sequences may loop back and occupy or obstruct the intended duplex region. Selecting a small subset of 33mer probes from the Affymetrix SNP6.0 Array we generated a pool of potentially cross-hybridizing sequence regions found by using the SeqNFindTM platform. Pools of length variant targets centered around the potentially cross hybridizing sequence were generated from human reference genome 36.3 and computationally examined using OMPTM to calculate the affinity constants. To demonstrate that the modeled properties affect measurements a small subset of these targets and probes were tested on microarrays.

Analysis of complete mitochondrial genome of <I>Cristaria plicata</I>

The complete mitochondrial genome of Cristaria plicata was obtained using long amplification polymerase chain reaction (LA-PCR). Analysis showed that the total length of sequence was 15 712 bp, including 13 protein-coding genes, 2 ribosomal RNA genes, 22 transfer RNA genes, and 26 non-coding regions ranged from 2 bp to 328 bp in size. The nucleotide composition of A, T, C, G were 36.54% 27.22% 23.22% and 13.02%, respectively. Most genes were encoded on the L strand, While ND3-ND5, ND4L, COI-COIII, ATP6, ATP8, tRNAAsp, and tRNAHis were encoded on the H strand. The arrangement of genes in C. plicata was identical to that of Lampsilis ornata, but was different from that of Hyriopsis cum-ingii between COII and 12S rRNA in the same family. Thirteen protein genes contained 3 initiation codons, i.e., I (AUU, AUC), V (GUG), and M (AUA, AUG), all of which had complete stop codons (UAA or UAG), except for the stop codon in ND2 that had an incomplete T. Fifteen tRNAs had typical cloverleaf structure, except for tRNAThr, tRNALys, tRNASer(UCN), tRNAAsp, tRNAArg, tRNATyr, and tRNAMet. Like other freshwater bivalvia, C. plicata contained ATP8 gene, which might be related to the balance of osmotic pressure in the cytoplasm.

Potential use of barcoding to identify aquatic hyphomycetes

Tetracladium is a common aquatic hyphomycete genus, whose taxonomy has been based on the morphology and development of asexual spores. Ecological surveys have relied almost exclusively on spore morphology. Since selective pressures have resulted in convergent shapes, misidentifications are a concern. To supplement morphological information, we determined COX1, ITS and D1/D2 sequences as potential barcodes on 21 strains belonging to 7 described Tetracladium species and an unidentified strain. Attempts to amplify the IGS region were unsuccessful. The ratio of intraspecific to interspecific variability was optimal with ITS, which also provided the intuitively most acceptable cladogram. Typical conidia and their variability for the seven described species are illustrated. Internal node reliability depended less on total sequence length and more on the mixture of conserved and variable regions used to build cladograms. This finding can be exploited for quickly increasing phylogenetic accuracy without greatly increasing the amount of amplification and sequencing. The results have important implications for identifying freshwater hyphomycetes in the field but further work is required to establish if this method works with plant pathogens.

Cymobase - the Reference Database for Cytoskeletal and Motor Proteins

Motor proteins are involved in processes like cellular transport, muscle contraction, and cell division. Three motors are myosin, dynein, and kinesin. They convert chemical energy (ATP) into mechanical work (movement). Protein sequences are the bases for many biochemical and cell biological experiments, as well as bioinformatical analyses.We implemented a web application (CyMoBase) to represent all sequence related information. Since the fist publication of Pfarao (www.cymobase.org), many changes have been integrated in the database scheme and the web application. Now, it is also possible to derive information about structures and genes. Over the years, the number of data has increased considerable. Today, there are 37 proteins, 16500 sequences, 132 domains, 819 species, 599 publications and 1392 projects, with a total sequence length of over 16 million amino acids. The web application has got a new central search page including nine search modules (species names, protein classes, taxonomy, species groups, domains, sequence meta data, publications, sequence names, and genes) and eleven result tabs (sequences, publications, downloadable FASTA files, alignment viewer, phylogenetic trees, sequence stats, domain composition, complex inventory, protein inventory, molecular weights, and class composition). All search modules can be combined to filter the results.Furthermore, we provide the gene structure of all sequences as computed by WebScipio. An other viewing option is the “Complex Inventory”, which presents the existence or absence of sequence homologs of certain complexes. The aligned protein sequences are available via the “FASTA Files” result tab.The size of the database, the kind of annotation, the possibility to use and combining different search modules, and the number of information and options offered by the web interface makes Pfarao the reference database number one for cytoskeletal and motor proteins.

Next-Generation Sequencing (NGS) in CMML, MDS and AML Detects Molecular Mutations in Oncogenes and Allows the Identification of Balanced Chromosomal Abnormalities with Extraordinary Sensitivity and Specificity.

Abstract 144PicoTiterPlate (PTP) pyrosequencing allows the detection of low-abundance oncogene aberrations in complex samples even with low tumor content. Here, we compared deep sequencing data of two Next-Generation Sequencing (NGS) assays to detect molecular mutations using a PCR-based strategy and, in addition, to uncover inversions, translocations, and insertions in a targeted sequence enrichment workflow (454 Life Sciences, Roche Diagnostics Corporation, Branford, CT). First, we studied 95 patients (CMML, n=81; AML, n=6; MDS, n=3; MPS, n=3; ET, n=2) using the amplicon approach and investigated seven candidate genes with relevance in oncogenesis of myeloid malignancies: TET2, RUNX1, JAK2, MPL, KRAS, NRAS, and CBL. 43 primer pairs were designed to cover the complete coding regions of TET2, RUNX1 (beta isoform), and hotspot regions of the latter genes. In total, 4128 individual PCR reactions were performed with DNA isolated from bone marrow mononuclear cells, followed by product purification, fluorometric quantitation, and equimolar pooling of the corresponding 43 amplicon products to generate one single sequence library per patient. For sequencing, a 454 8-lane PTP was used applying standard FLX chemistry and representing one patient per lane. The median number of base pairs sequenced per patient was 9.23 Mb. For each amplicon a median of 840 reads was generated (coverage range: 485–1929 reads). As initial proof-of-concept analysis 27 of the 95 patients with known mutations (n=32) as detected by conventional sequencing or melting curve analyses were investigated (range of cells carrying the respective mutation: 1.1% for JAK2 V617F to 98.14% for TET2 C1464X). In all cases, 454 NGS confirmed results from routine diagnostic methods (GS Amplicon Variant Analyzer software version 2.0.01). We then investigated the remaining 69 CMML patients: In median, 2 variances (range 1–8 variances), i.e. differences in comparison to the reference sequence, per patient were detected. These variances included both point mutations in all candidate genes and large deletions (12-19 bp) in CBL, RUNX1, and TET2. Only 20/81 patients of the CMML-cohort (24.69%) were without any detectable mutation. Secondly, in a cohort of six AML bone marrow specimens a custom NimbleGen array (385K format; Madison, WI) was used to perform a targeted DNA sequence enrichment procedure. In total, capture probes spanning 1.91 Mb were designed to represent all coding regions of 92 target genes (1559 exons) with relevance in hematological malignancies (e.g. KIT, NF1, TP53, BCR, ABL1, NPM1, or FLT3). In addition, the complete genomic regions were targeted for RUNX1, CBFB, and MLL. For sequencing, 454 Titanium chemistry was applied, loading three patients per lane on a 2-lane PTP including three molecular identifiers (MIDs) each. Data analysis was performed using the GS Reference Mapper software version 2.0.01. For the enrichment assay, the median enrichment of the targeted genomic loci was 207-fold, as assessed by ligation-mediated LM-PCR. Overall, 1,098,132 reads were generated in the two lanes, yielding a total sequence length of 386,097,740 bases. In median, 96.52% of the sequenced bases mapped against the human genome, and 66.0% were derived from the customized NimbleGen array capture probes, resulting in a median coverage of 18.7-fold . With this method it was possible to detect and confirm point mutations (KIT, FLT3-TKD, and KRAS) and insertions (FLT3-ITD). Moreover, by capturing chimeric DNA fragments and generating reads mapping to both fusion partners this approach detected balanced aberrations, i.e. inv(16)(p13q22) and the translocations t(8;21)(q22;q22) or t(9;11)(p22;q23). In conclusion, both assays to specifically sequence targeted regions with oncogenic relevance on a NGS platform demonstrated promising results and are feasible. The amplicon approach is more suitable for detection of mutations in a routine setting and is ideally suited for large genes such as TET2, ATM, and NF1, which are labor-intensive to sequence conventionally. The array-based capturing assay is characterized by a complex and time-consuming workflow with low-throughput. However, the ability to detect balanced genomic aberrations which are detectable thus far only by cytogenetics and FISH has the potential to become an important diagnostic assay, especially in tumors in which cytogenetics can not be applied successfully. Disclosures:Grossmann:MLL Munich Leukemia Laboratory: Employment. Kohlmann:MLL Munich Leukemia Laboratory: Employment. Haferlach:MLL Munich Leukemia Laboratory: Equity Ownership. Dicker:MLL Munich Leukemia Laboratory: Employment. Kazak:MLL Munich Leukemia Laboratory: Employment. Schindela:MLL Munich Leukemia Laboratory: Employment. Schnittger:MLL Munich Leukemia Laboratory: Equity Ownership. Kern:MLL Munich Leukemia Laboratory: Equity Ownership. Haferlach:MLL Munich Leukemia Laboratory: Equity Ownership.