The invention relates to the field of
biotechnology, and provides improved identification for target
RNA sequences bound to
RNA-
binding protein in cells. Through UV crosslinking-immunopricipitation, target
RNA of RNA-
binding protein is obtained, micrococcal
nuclease is adopted for carrying out incomplete enzymolysis on the target RNA after
ultraviolet crosslinking, after enzymolysis, a chelating agent capable of removing Ca<2+> is adopted for inactivating the
enzyme, the 3' terminal of the target RNA obtained through the method is phosphorylated, the phosphorylated 3' terminal is ligated witha 3' RNA
linker, then, the
ligation product is phosphorylated, the phosphorylated
ligation product is separated, the separated phosphorylated
ligation product is recycled, then the 3' terminal is ligated with a 5' RNA
linker, RT-PCR amplification is carried out, so that a
cDNA library corresponding to the target RNA is obtained. With the technical scheme provided by the invention, the experiment flow is simplified, the step of
isotope labelling is eliminated, due to the adding of an IgG
antibody negative control sample, the influences of non-specific binding and background on the experimentalresult are eliminated, and the data result with the quality equal to that of Clip-seq is obtained.