Biomedical subjects

Torstein Tengs

Publications and source records attributed to Torstein Tengs.

9 recordsLinked to original sources

PubMed2006

Equal performance of TaqMan, MGB, molecular beacon, and SYBR green-based detection assays in detection and quantification of roundup ready soybean.

We have tested and compared the performance of 12 different assays representing four different real-time polymerase chain reaction (PCR) chemistries in the context of genetically modified organism detection. Several different molecular beacon, SYBR Green, TaqMan, and MGB assays were designed for the event specific detection and quantification of the 3' integration junction of GTS 40-3-2 (Roundup Ready) soybean. Sensitivity as well as robustness in the presence of background DNA were tested. None of the PCR-based approaches appeared to be significantly better than any of the other, but the molecular beacon assays had the lowest efficiency and also seemed more sensitive to changes in experimental setup.

DNA, Plant↗

PubMed2006

Sensitive mutation detection in heterogeneous cancer specimens by massively parallel picoliter reactor sequencing.

The sensitivity of conventional DNA sequencing in tumor biopsies is limited by stromal contamination and by genetic heterogeneity within the cancer. Here, we show that microreactor-based pyrosequencing can detect rare cancer-associated sequence variations by independent and parallel sampling of multiple representatives of a given DNA fragment. This technology can thereby facilitate accurate molecular diagnosis of heterogeneous cancer specimens and enable patient selection for targeted cancer therapies.

Base Sequence↗

PubMed2006

Heterotachy processes in rhodophyte-derived secondhand plastid genes: Implications for addressing the origin and evolution of dinoflagellate plastids.

Serial transfer of plastids from one eukaryotic host to another is the key process involved in evolution of secondhand plastids. Such transfers drastically change the environment of the plastids and hence the selection regimes, presumably leading to changes over time in the characteristics of plastid gene evolution and to misleading phylogenetic inferences. About half of the dinoflagellate protists species are photosynthetic and unique in harboring a diversity of plastids acquired from a wide range of eukaryotic algae. They are therefore ideal for studying evolutionary processes of plastids gained through secondary and tertiary endosymbioses. In the light of these processes, we have evaluated the origin of 2 types of dinoflagellate plastids, containing the peridinin or 19'-hexanoyloxyfucoxanthin (19'-HNOF) pigments, by inferring the phylogeny using "covarion" evolutionary models allowing the pattern of among-site rate variation to change over time. Our investigations of genes from secondary and tertiary plastids derived from the rhodophyte plastid lineage clearly reveal "heterotachy" processes characterized as stationary covarion substitution patterns and changes in proportion of variable sites across sequences. Failure to accommodate covarion-like substitution patterns can have strong effects on the plastid tree topology. Importantly, multigene analyses performed with probabilistic methods using among-site rate and covarion models of evolution conflict with proposed single origin of the peridinin- and 19'-HNOF-containing plastids, suggesting that analysis of secondhand plastids can be hampered by convergence in the evolutionary signature of the plastid DNA sequences. Another type of sequence convergence was detected at protein level involving the psaA gene. Excluding the psaA sequence from a concatenated protein alignment grouped the peridinin plastid with haptophytes, congruent with all DNA trees. Altogether, taking account of complex processes involved in the evolution of dinoflagellate plastid sequences (both at the DNA and amino acid level), we demonstrate the difficulty of excluding independent, tertiary origin for both the peridinin and 19'-HNOF plastids involving engulfment of haptophyte-like algae. In addition, the refined topologies suggest the red algal order, Porphyridales, as the endosymbiont ancestor of the secondary plastids in cryptophytes, haptophytes, and heterokonts.

Animals↗

PubMed2005

A transforming MET mutation discovered in non-small cell lung cancer using microarray-based resequencing.

We have designed resequencing microarrays to test the performance of this platform when interrogating a large number of exons (164 total) from genes associated with cancer. To evaluate false positive and negative rates, dideoxy sequencing was done for 335,420 bases interrogated by the arrays. From the array data, calls could be made for approximately 97.5% of the bases, and false positive rates were very low with only a single mutation reported from the array dataset for which the corresponding dideoxy trace had a clean wildtype sequence. For the nucleotide positions where array calls were made, false negative rates were 1.41% for heterozygous mutations. All the homozygous mutations were detected, but 8.11% were erroneously reported as heterozygous changes from the reference sequence by the array analysis software. In addition, 20 non-small cell lung cancer (NSCLC) samples were analyzed using the arrays, and both somatic and germline mutations were found. The most interesting findings were two MET mutations that have recently been implemented in NSCLC. Large scale MALDI-TOF genotyping indicated that one of these mutations (T1010I) might represent a true cancer-causing genotype, whereas the other (N375S) appears to be a common germline polymorphism.

Carcinoma, Non-Small-Cell Lung↗

PubMed2004

Statistical analysis of genomic tag data.

We present a series of statistical solutions to challenges that commonly arise in the production and analysis of genomic tag libraries. Tag libraries are collections of fragments of DNA or RNA, with each unique fragment often present in millions or billions of copies. Inferences can be made from data obtained by sequencing a subset of the library. The statistical approaches outlined in this paper are divided into three parts. First, we demonstrate the application of classical capture-recapture theory to the question of library complexity, i.e. the number of unique fragments in the library. Simulation studies verify the accuracy, for sample sizes of magnitudes typical in genomic studies, of the formulas we use to make our estimates. Second, we present a straightforward statistical cost analysis of tag experiments designed to uncover either disease-causing pathogens or new genes. Third, we develop a hidden Markov model approach to karyotyping a sample using a tag library derived from the sample's genomic DNA. While the resolution of the approach depends upon the number of tags sequenced from the library, we show via simulation that copy number alterations can be reliably detected for lengths as small as 1 Mb, even when a moderate number of tags are sequenced. Simulations predict very good specificity as well. Finally, all three of our approaches are applied to data from real tag library experiments. The hidden Markov model results are in line with what was expected from simulation, and genomic alterations found by applying the method to a cancer cell line library are confirmed using PCR. The methods and data described in this paper are contained in an R package, tagAnalysis, freely available at http://meyerson.dfci.harvard.edu/~tl974/tagAnalysis.

Journal Article↗

PubMed2004

Genomic representations using concatenates of Type IIB restriction endonuclease digestion fragments.

We have developed a method for genomic representation using Type IIB restriction endonucleases. Representation by concatenation of restriction digests, or RECORD, is an approach to sample the fragments generated by cleavage with these enzymes. Here, we show that the RECORD libraries may be used for digital karyotyping and for pathogen identification by computational subtraction.

Bacteria↗

PubMed2004

Highly divergent SSU rRNA genes found in the marine ciliates Myrionecta rubra and Mesodinium pulex.

Myrionecta rubra and Mesodinium pulex are among the most commonly encountered planktonic ciliates in coastal marine and estuarine regions throughout the world. Despite their widespread distribution, both ciliates have received little attention by taxonomists. In order to better understand the phylogenetic position of these ciliates, we determined the SSU rRNA gene sequence from cultures of M. rubra and M. pulex. Partial sequence data were also generated from isolated cells of M. rubra from Chesapeake Bay. The M. rubra and M. pulex sequences were very divergent from all other ciliates, but shared a branch with 100% bootstrap support. Both species had numerous deletions and substitutions in their SSU rRNA gene, resulting in a long branch for the clade. This made the sequences prone to spurious phylogenetic affiliations when using simple phylogenetic methods. Maximum likelihood analysis placed M. rubra and M. pulex on the basal ciliate branch, following removal of ambiguously aligned regions. Fluorescent in situ hybridization probes were used with confocal laser scanning microscopy to confirm that these divergent sequences were both expressed in the cytoplasm and nucleolus of M. ruisra and M. pulex. We found that our sequence data matched several recently discovered unidentified eukaryotes in Genbank from diverse marine habitats, all of which had apparently been misattributed to highly divergent amoeboid organisms.

Animals↗

PubMed2003

Identical ribosomal DNA sequence data from Pfiesteria piscicida (Dinophyceae) isolates with different toxicity phenotypes.

Complete small subunit ribosomal RNA, internal transcribed spacer 1 and 2, 5.8S, and partial large subunit ribosomal RNA gene sequences were generated from multiple isolates of Pfiesteria piscicida. Sequences were derived from isolates that have been shown to be ichthyotoxic as well as isolates that have no history of toxic behavior. All of the sequences generated were identical for the different cultures, and we therefore conclude that differences in toxicity seen between isolates of P. piscicida are linked to factors other than genetic strain variation detectable by ribosomal gene sequence analyses.

Animals↗

PubMed2002

Discovery of the toxic dinoflagellate Pfiesteria in northern European waters.

Several dinoflagellate strains of the genus Pfiesteria were isolated by culturing techniques from sediment samples taken in the Oslofjord region of Norway. Pfiesteria piscicida, well known as a fish killer from the Atlantic coast of America, was identified by genetic methods and light microscopy. The related species Pfiesteria shumwayae was attracted from the sediment by the presence of fish, and has proved toxic. This present survey demonstrates the wide distribution of these potentially harmful species, but so far they have not been connected with fish kills in Europe.

Animals↗