Cloning method for series parallel expression of a plurality of sgRNA based on grading assembling and application

A CRISPR/Cas9 system has an ultrahigh parallel capacity. In order to meet the requirement for expressing a plurality of sgRNA in some cases, the invention provides a quick assembling method for a plurality of parallel expressed sgRNA. The invention utilizes grading Golden Gate reaction to develop a multi-turn amplifying method based on polymerase chain reaction and independent of a carrier, so as to realize the quick connecting assembling for 20 sgRNA within one week. The method for serially assembling a plurality of sgRNA developed by the invention has the advantages of time-saving and labor-saving effect, flexibility, high efficiency and multifunction. The method can be used for quickly assembling 2-20 sgRNA in different quantity onto one carrier. A plurality of parallel expressed sgRNA are utilized to target to a section of DNA, so that the functions of marking and tracking unrepeated sequence chromatin locus in living cells, cooperatively activating or restraining a single gene, simultaneously editing a p