Reference sequences are used in gap4 (see section Gap4 introduction). Here they can be used to define a numbering system independent of gaps introduced to produce alignments. The numbering can start at any point in the reference sequence. If the reference sequence is entered with a feature table the features are converted to tags and can be used to control translation of the sequence in the contig editor. For mutation detection work the reference sequence and feature table enable mutations to be reported using positions defined by the reference sequence, and also allows the effect of the mutations to be noted. Gap4 is able to store entries from the EMBL sequence library complete with their feature tables. These feature tables are converted to gap4 database annotations (tags), which means that they can be selectively displayed in the template display and editor, and used to translate only the exons (in the correct reading frame). Obviously it may be useful to augment the feature tables with the sites of known polymorphisms or deleterious mutations so that they can be displayed in gap4 as landmarks. When it comes to producing a report of the observed mutations the feature table is used to work out if a mutation is expressed and if so what the amino acid change is. Additional tags can be created to specify the positions of the primers or restriction sites used to obtain data covering segments of the sequence. For any project the reference sequence need only be set up once. Either project databases can be started with the reference sequence already configured or the reference can be assembled along with the reading data. The reference sequence can be designated (or reassigned) as follows. In pregap4 (see section Pregap4 introduction) it can be named in the module "Reference Traces". In the gap4 editor it can be set by right clicking on its name. Once set it should appear labelled "S" at the left edge of the editor.