The invention belongs to the technical field of
molecular pathology and
immunology, and relates to construction of pig
peripheral bloodmononuclear lymphocytePD-L1 (
programmed death-ligand1) recombinantplasmids, a real-time
gene abundance detection method and an application of the method. TotalRNA of pig
peripheral bloodmononuclear lymphocytes is collected and reverselytranscribed into cDNA (
complementary DNA); PD-L1
target gene fragments are amplified through a general PCR (
polymerase chain reaction), detected throughagarose
gel electrophoresis, recovered and purified; the PD-L1 targetgene fragments are connected with pMD18-T carriers and converted into competent cells DH5alpha, and the recombinantplasmids are extracted; sequencing analysis is performed after
cloning and screening, positive plasmids with the same sequence as the
target gene fragments are selected to serve as standard plasmids, and a
standard curve is drawn according to the copy concentration; thePD-L1
gene abundance is measured according to
fluorescence signal changes and the
standard curve. The real-time pig PD-L1
gene abundancedetection method has the advantages of high detection
throughput, high susceptibility, high specificity, convenience inoperation, low cost, accuracy inquantification and the like.