A method for designing all coverage of valid primer pairs, which satisfy various filtering constraints provided by users with respect to a given sequence database and has validated specificity to given sequences, is provided. By screening all suitable primer pairs present on a given DNA sequence database without omitting any one primer pair and also screening all primers having a coverage of 1 or more as well as primers having a coverage of 1, a user can be allowed to give rankings to the primers in order to easily select the primers having a high success rate in biological experiments from the resulting primers.1. A method for designing all coverage of valid primers which satisfies the conditions for specificity to large-scale DNA sequences in a large-scale DNA sequence database, comprising: receiving a given DNA sequence database to extract partial sequences for candidate primers having all possible lengths between the minimum length and the maximum length (operation 1);excluding the primers which do not satisfy input single filtering conditions when the candidate primers extracted in operation 1 are subjected to the single filtering conditions (operation 2);pair-joining Map1, which includes all the possible partial sequences obtained in operation 1, and Map2, which includes candidate primer sets satisfying the single filtering conditions obtained in operation 2, and removing the primers for Map2 when the primers for Map1 and Map2 have the same sequences other than the 5′ termini thereof (operation 3);pair-joining Map1, which includes all the possible partial sequences obtained in operation 1, and Map2, which includes candidate primer sets satisfying the single filtering conditions and 5′ cross-hybridization filtering conditions obtained in operation 3, and removing the primers for Map2 when the primers for Map1 the same sequences as the primers for Map2 except the sequences having a given mismatch number (#mismatch) (operation 4);removing false-positive primers which still remain from the results of operation 4 and do not satisfy general cross-hybridization filtering conditions (operation 5);dividing the primers remaining from the results of operation 5 into forward primer sets and reverse primer sets and excluding the primers which do not satisfy the filtering conditions for self-join calculation when the divided forward and reverse primer sets are subjected to the filtering conditions (operation 6); andcalculating penalty scores for the primer pairs passing operation 6, and sorting the primer pairs in the same sidset group according to the calculated penalty scores (operation 7)
