Biomedical subjects
Publications and source records attributed to R Staden.
The long-term goal of this project is the elucidation of the complete sequence of the Caenorhabditis elegans genome. During the first year methods have been developed and a strategy implemented that is amenable to large-scale sequencing. The three cosmids sequenced in this initial phase are surprisingly rich in genes, many of which have mammalian homologues.
We describe a set of programs for creating and using indexes for the distributed forms of the major sequence libraries. The indexes conform to the specification of those distributed on cd-rom by the EMBL sequence library. The programs create entry name, accession number, author and freetext indexes and a brief directory index. If a suitable application program is given an entry name or accession number these indexes allow rapid retrieval of sequences or annotation. Similarly the author and freetext indexes provide the data for extremely fast searching on author names and "keywords". The indexing programs can create indexes for EMBL, Swiss-Prot, GenBANK, PIR and NRL3d libraries. We also describe the organisation and use of the different sequence libraries and their index files.
There are now a number of machines for determining DNA sequences. These devices are currently of two types: those such as the Applied Biosystems 373A and the Pharmacia A.L.F. which interpret the sequences of samples as they run on gels within the machine, and those, such as the Bio-Rad and Amersham readers that scan and analyse conventional autoradiographs. Both types of machine can produce their data in the form of traces which represent the band intensity of each of the four base types at each position in the sequence. At present all the machines write files in different formats. We describe a machine independent format for storing data derived from automatic sequencing machines. Files in this format can store the derived sequence, the traces and a set of confidence measures for each base. We have adopted the format as the standard for our sequence handling software.
We describe a sequence assembly and editing program for managing large and small projects. It is being used to sequence complete cosmids and has substantially reduced the time taken to process the data. In addition to handling conventionally derived sequences it can use data obtained from Applied Biosystems,Inc. 373A and Pharmacia A.L.F. fluorescent sequencing machines. Readings are assembled automatically. All editing is performed using a mouse operated contig editor that displays aligned sequences and their traces together on the screen. The editor, which can be used on single contigs or for joining contigs, permits rapid movement along the aligned sequences. Insertions, deletions and replacements can be made in individual aligned readings and global changes can be made by editing the consensus. All changes are recorded. A click on a mouse button will display the traces covering the current cursor position, hence allowing quick resolution of problems. Another function automatically moves the cursor to the next unresolved character. The editor also provides facilities for annotating the sequences. Typical annotations include flagging the positions of primers used for walking, or for marking sites, such as compressions, that have caused problems during sequencing. Graphical displays aid the assessment of progress.
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We describe programs that can screen nucleic acid and protein sequences against libraries of motifs and patterns. Such comparisons are likely to play an important role in interpreting the function of sequences determined during large scale sequencing projects. In addition we report programs for converting the Prosite protein motif library into a form that is compatible with our searching programs. The programs work on VAX and SUN computers.
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We report improvements to our sequence analysis package and adaptation to run on the Apple Macintosh range of machines. The 'standard' version of the programs, which run on a VAX, has been given a new user interface that makes the programs very much easier to work with and has facilitated the move to the Macintosh. The reorganization of the code should simplify moves to other systems that offer WIMP user interfaces. In addition to a large number of small but useful extra features, some important new analytical functions have been devised. These include sequence and contig editors; optimal alignment and comparison methods; and a new method for comparing the observed and expected frequencies of selected oligonucleotides.
With modern fast sequencing techniques and suitable computer programs it is now possible to sequence whole genomes without the need of restriction maps. This paper describes computer programs that can be used to order both sequence gel readings and clones. A method of coding for uncertainties in gel readings is described. These programs are available on request.
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The 5,577 nucleotide long sequence of bacteriophage G4 DNA has been determined using the 'plus and minus' and chain termination methods of DNA sequencing. This sequence has been compared with that of the closely related bacteriophage phiX174 (refs 1, 55). In the coding regions there is an average of 33.1% nucleotide sequence differences between the two genomes, but the distribution of these changes is not random and the sequence of some genes is more conserved than others. There is less sequence similarity between the untranslated intergenic regions of G4 and phiX174, but despite this the sequences of the J/F, F/G and H/A untranslated spaces in both genomes have similar sized hairpin loops, which may be related to their function.
A short partial sequence homology is present at all intron-exon junctions, or splice points, in the chick ovalbumin gene; it is probably a signal for a splicing enzyme. The significance of the junction sequences for splicing is discussed. We find no evidence of strong Watson-Crick base pairing between adjacent junctions.
A previous paper(1) described programs for sequence data handling and analysis by computer. The facilities of this basic set are extended by further easily used programs.
The speed of the new DNA sequencing techniques has created a need for computer programs to handle the data produced. This paper describes simple programs designed specifically for use by people with little or no computer experience. The programs are for use on small computers and provide facilities for storage, editing and analysis of both DNA and amino acid sequences. A magnetic tape containing these programs is available on request.