The invention relates to a long-chain dibasic acid with low content of fatty acid impurity and a method of producing it, in particular to the preparation of a long-chain dibasic acid producing strain by using directed evolution and homologous recombination, and to the fermentation production of a long-chain dibasic acid with low content of fatty acid impurity by using said strain. The invention relates to an isolated mutated CPR-b gene, homologous gene or variant thereof, relative to the GenBank Accession Number AY823228, taking the first base upstream of the start codon ATG as −1, comprising a base mutation −322G>A, and taking the first base downstream of the stop codon TAG as 1, comprising mutations 3TUTR.19C>T and 3′UTR.76_77insT. The invention also relates to a strain containing said mutated CPR-b gene, homologous gene or variant thereof, wherein, when the strain is used to produce a long-chain dibasic acid by fermentation, the content of fatty acid impurity in the fermentation product is significantly decreased.