The invention discloses a method and a device for quick contrast and analysis of a short sequence for second-generation sequencing, which can solve the problems of low contrast efficiency and high memory occupation ratio of sequencing data. The method comprises the following steps of obtaining a DNA (deoxyribonucleic acid) short sequence obtained by sequencing, and respectively mapping and encoding the DNA short sequence by a first hash algorithm and a second hash algorithm, so as to respectively obtain a first index and a second index; according to a preset index query library, the first index and the second index, contrasting the DNA short sequence and a reference gene group, wherein the index query library consists of an unit structure array, and each unit structure comprises value and index 2; storing the array index offset of each unit structure as the corresponding index 1, namely the index value corresponding to the structure array, wherein K is the length of segment sequence; according to the contrast result, when the contrast result is correct, obtaining the value of the K-mer segment contrasted with the corresponding DNA short sequence, and determining the chromosome number of the corresponding DNA short sequence and the site on the chromosome.