The availability of massive amounts of DNA sequence information has begun to revolutionize the practice of biology. As a result, current large-scale sequencing output, while impressive, is not adequate to keep pace with growing demand and, in particular, is far short of what will be required to obtain the 3-billion-base human genome sequence by the target date of 2005. To reach this goal, improved automation will be essential, and it is particularly important that human involvement in sequence data processing be significantly reduced or eliminated. Progress in this respect will require both improved accuracy of the data processing software and reliable accuracy measures to reduce the need for human involvement in error correction and make human review more efficient. Here, we describe one step toward that goal: a base-calling program for automated sequencer traces, phred, with improved accuracy. phred appears to be the first base-calling program to achieve a lower error rate than the ABI software, averaging 40%–50 % fewer errors in the data sets examined independent of position in read, machine running conditions, or sequencing chemistry.

Since the advent of rapid DNA sequencing methods in 1976, scientists have had the problem of inferring DNA sequences from sequenced fragments. Shotgun sequencing is a ‘ well-established biological and computational method used in practice. Many conventional algorithms for shotgun sequencing are based on the notion.of pairwisk fragment overlap. * While shotgun sequencing infers a DNA sequence given the sequences of overlapping frag-ments, a recent and complementary method, called sequencing by hybridization (SBH), in-fers a DNA sequence given the set of oligomers that represents all subwords of some fixed length, k. In this paper,. we propose a new computer algorithm for DNA sequence assembly that combines in a novel way the techniques of both shotgun and SBH methods. Based on our preliminary investigations, the algorithm promises- to be very fast and practical for DNA sequence assembly.

Abstract. The amino acid sequence deduced from eDNA of the human placental fibronectin receptor is reported. The receptor is composed of two subunits: an 0t subunit of 1,008 amino acids which is processed into two polypeptides disulfide bonded to one another, and a 13 subunit of 778 amino acids. Each subunit has near its COOH terminus a hydrophobic segment. This and other sequence features suggest a structure for the receptor in which the hydrophobic segments serve as transmembrane domains anchoring each subunit to the membrane and dividing each into a large ectodomain and a short cytoplasmic domain. The r subunit ectodomain has five sequence elements homologous to consensus Ca2 § sites of several calciumbinding proteins, and the 13 subunit contains a fourfold repeat strikingly rich in cysteine. The r subtmit sequence is 46 % homologous to the a subunit of the vitronectin receptor. The 13 subunit is 44 % homologous to the human platelet adhesion receptor subunit IIIa and 47 % homologous to a leukocyte adhesion receptor 13 subunit. The high degree of homology (85%) of the 13 subunit with one of the polypeptides of a chicken adhesion receptor complex referred to as integrin complex strongly suggests that the latter polypeptide is the chicken homologue of the fibronectin receptor 13 subunit. These receptor subunit homologies define a superfamily of adhesion receptors. The availability of the entire protein sequence for the fibronectin receptor will facilitate studies on the functions of these receptors. T HE cell surface receptors that bind to the adhesive glycoproteins fibronectin, vitronectin, fibrinogen, and von Willebrand factor have been shown to be structurally and functionally related (Pytela et al., 1986). Furthermore, they have been shown to be members of a superfamily of receptors that include the leukocyte glycoproteins (LFA-1, Mac-l, and p150,95 (Springer et al., 1986) and another group of cell surface proteins referred to as VLA antigens (Hemler et al., 1987). Homologous forms of these receptors have been defined in various species by using antibodies that interfere with cell adhesion (Knudsen et al., 1981; Greve and Gottlieb,

the ORF-encoded polypeptides are related to sequences with unknown functions, and 496 (27%) have little or no homology to sequences in public databases. Comparisons with Eucarya-, Bacteria-, and Archaea -specific databases reveal that 1,013 of the putative gene products (54%) are most similar to polypeptide sequences described previously for other organisms in the domain Archaea. Comparisons with the Methanococcus jannaschii genome data underline the extensive divergence that has occurred between these two methanogens; only 352 (19%) of M. thermoautotrophicum ORFs encode sequences that are >50% identical to M. jannaschii polypeptides, and there is little conservation in the relative locations of orthologous genes. When the M. thermoautotrophicum ORFs are compared to sequences from only the eucaryal and bacterial domains, 786 (42%) are more similar to bacterial sequences and 241 (13%) are more similar to eucaryal sequences. The bacterial domain-like gene products include the ma

The fragment assembly problem is that of reconstructing a DNA sequence from a collection of randomly sampled fragments. Traditionally the objective of this problem has been to produce the shortest string that contains all the fragments as substrings, but in the case of repetitive target sequences this objective produces answers that are overcompressed. In this paper, the problem is reformulated as one of finding a maximum-likelihood reconstruction with respect to the 2-sided Kolmogorov-Smirnov statistic, and it is argued that this is a better formulation of the problem. Next the fragment assembly problem is recast in graph-theoretic terms as one of finding a non-cyclic subgraph with certain properties and the objectives of being shortest or maximally-likely are also recast in this framework. Finally, a series of graph reduction transformations are given that dramatically reduce the size of the graph to be explored in practical instances of the problem. This reduction is ...

Abstract. Dynamin is a 100-kD microtubule-activated GTPase. Recent evidence has revealed a high degree of sequence homology with the product of the Drosophila gene shibire, mutations in which block the recycling of synaptic vesicles and, more generally, the formation of coated and non-coated vesicles at the plasma membrane. We have now transfected cultured mammalian COS-7 cells with both wild-type and mutant dynamin cDNAs. Point mutations in the GTPbinding consensus sequence elements of dynamin equivalent to dominant negative mutations in ras, and an NH2-terminal deletion of the entire GTP-binding domain of dynamin, block transferrin uptake and alter the distribution of clathrin heavy chain and a-, but not 3 % adaptin. COOH-terminal deletions reverse these

Abstract. The Saccharomyces cerevisiae CDC42 gene product is involved in the morphogenetic events of the cell division cycle; temperature-sensitive cdc42 mutants are unable to form buds and display delocalized cell-surface deposition at the restrictive temperature

Abstract. Northern blot analysis of rat heart mRNA probed with a eDNA coding for the principal polypeptide of rat liver gap junctions demonstrated a 3.0-kb band. This band was observed only after hybridization and washing using low stringency conditions; high stringency conditions abolished the hybridization. A rat heart eDNA library was screened with the same eDNA probe under the permissive hybridization conditions, and a single positive clone identified and purified. The clone contained a 220-bp insert, which showed 55 % homology to the original eDNA probe near the 5 ' end. The 220-bp eDNA was used to rescreen a heart eDNA library under high stringency conditions, and three additional cDNAs that together spanned 2,768 bp were isolated. This composite eDNA

Abstract. Cdc31 mutants of Saccharomyces cerevisiae arrest at the nonpermissive temperature with large buds, G2 DNA content and, a single, abnormally large spindle pole body (SPB) (Byers, B. 1981. Molecular Genetics in Yeast. Alfred Benzon Symposium. 16:119-133). In this report, we show that the CDC31 gene product is essential for cell viability. We demonstrate that purified CDC31 protein binds Ca 2 ÷ and that this binding is highly specific. Taken together, three lines of evidence indicate that CDC31 is a component of the SPB. First, CDC31 cofractionates with enriched preparations of SPBs. Second, immunofluorescence staining indicates that CDC31 colocalizes with a known SPB component. Third, immunoelectron microscopy with whole cells and with isolated SPBs reveals that CDC31 is localized to the half bridge of the SPB, which lies immediately adjacent to the SPB plaques. CDC31 was detected mainly at the cytoplasmic side of the half bridge and, therefore, defines a further substructure of the SPB. We suggest that CDC31 is a member of a family of calcium-binding, centrosome-associated proteins from a phylogenetically diverse group of organisms. I N the budding yeast Saccharomyces cerevisiae, centrosomal functions are provided by the spindle pole body (SPB), t a cylindrical multilaminated organelle which is embedded in the nuclear envelope (Byers and Goetsch, 1975; see Fig. 8). An outer, central, and inner plaque can be discriminated by EM. The central plaque serves to anchor the SPB in the nuclear envelope. Outer and inner plaques function as nucleation sites for cytoplasmic and nuclear microtubules, respectively (Byers, 1981a). The inner plaque is not as electron dense as the outer and central plaques and is only clearly detectable after extraction of isolated SPBs with DEAE-dextran (Rout and Kilmartin, 1990). An additional substructure of the SPB, the half bridge, appears in the electron microscope as a darkly stained strip along the cytoplasmic margin of the nuclear envelope (Byers and Goetsch,

ABSTRACT Mutations have been introduced into the cloned DNA sequences coding for influenza virus hemagglutinin (HA), and the resulting mutant genes have been expressed in simian cells by the use of SV40-HA recombinant viral vectors. In this study we analyzed the effect of specific alterations in the cytoplasmic domain of the HA molecule on its rate of biosynthesis and transport, cellular localization, and biological activity. Several of the mutants displayed abnormalities in the pathway of transport from the endoplasmic reticulum to the cell surface. One mutant HA remained within the endoplasmic reticulum; others were delayed in reaching the Golgi apparatus after core glycosylation had been completed in the endoplasmic reticulum, but then progressed at a normal rate from the Golgi apparatus to the cell surface; another was delayed in transport from the Golgi apparatus to the plasma membrane. However, two mutants were indistinguishable from wild-type HA in their rate of movement from the endoplasmic reticulum through the Golgi apparatus to the cell surface. We conclude that changes in the cytoplasmic domain can powerfully influence the rate of intracellular transport and the efficiency with which HA reaches the cell surface. Nevertheless, absolute conservation of this region of the molecule is not required for maturation and efficient