THE PCR-SSP MANAGER COMPUTER-PROGRAM - A TOOL FOR MAINTAINING SEQUENCE ALIGNMENTS AND AUTOMATICALLY UPDATING THE SPECIFICITIES OF PCR-SSP PRIMERS AND PRIMER MIXES

An emerging problem of molecular typing methods such as PCR amplificat ion using sequence-specific primers (PCR-SSP) is that they frequently require updating as new alleles are constantly being described which p otentially affect the specificity of every PCR-SSP reaction. PCR-SSP u ses pairs of primers to detect cis-linked polymorphisms and thus each new allele described must be compared to each individual primer pair. Furthermore, sequence homology between the various loci for class I an d class II means that, for example, new HLA-A sequences have to be com pared with HLA-B and HLA-C primps mixes to rule out cross-locus amplif ication. We have developed a computer program known as SSP Manager whi ch is capable of aligning HLA class I and class II sequences obtained from Internet-accessible databases such as GenBank. The program then u pdates all individual primer specificities held in its database before updating the specificities of all primer mixes. Sets of primer mixes can then be combined from the primer mix directory to create PCR-SSP t yping trays which are subsequently analysed by the program. A report i s generated which stipulates whether all known sequences are amplified and the reason for apparent failure to test for individual alleles, e ,g, a lack of relevant sequence information. SSP Manager has the flexi bility to cope with unusual sequences (deletions and insertions), prim ers with internal mismatches and primers with a deliberate mismatch. T he program also has many tools for developing new primer mixes, such a s the facility to search for novel reactions using Boolean operators. The organisation and operational use of the SSP Manager program is des cribed and its uses are illustrated with an updated allele list for ou r previously described Phototyping PCR-SSP class I and class II typing set The SSP Manager is available on request From the authors.