The invention discloses a method for massively and efficiently developing molecular markers on the basis of
Indel and SSR (simple
sequence repeat) site techniques. The method comprises the following steps: (1) selecting at least 3 samples to be developed, and respectively extracting DNAs (deoxyribonucleic acids) of the samples to be developed; (2) carrying out
enzyme digestion on the
DNA samples of the samples to be developed, establishing a sequencing
library, and carrying out sequencing; (3) mixing the genomes of all the samples to be developed, and assembling to obtain Contigs; (4) comparing the Contigs with the sequences of the sample individuals to be developed, and acquiring SSR sites with
Indel inside according to the
Indel and SSR site information as a candidate polymorphism SSR site; (5) designing primers according to the obtained candidate polymorphism SSR site, carrying out PCR (
polymerase chain reaction) amplification and sequencing, and selecting a ribbon-shaped stable clear strip as
molecular marker primers to be verified; and (6) carrying out PCR amplification on different samples to be developed by using the obtained
molecular marker primers, and selecting the molecular markers with diversity, thereby obtaining the molecular markers. The method enhances the
molecular marker development efficiency; and the developed SSR molecular markers can be efficiently applied to research in the aspects of
genetics, multiplication release evaluation and the like.