Fluorescent quantitative PCR detection method and detection kit of KIR2DL2 mRNA

Abstract

The invention belongs to the field of biological detection, and particularly relates to a fluorescent quantitative PCR detection method and detection kit of KIR2DL2 mRNA. A method for designing a primer and a probe comprises the following steps: (1) performing KIR genetic typing on a DNA sample by applying a PCR-SSOP method, screening out the positive DNA of KIR2DL2; (2) performing PCR amplification and electrophoresis on the positive DNA sample (3) extracting a specific DNA band of the KIR2DL2, performing tapping and purifying, linking with a T vector, transforming, selecting flora, and extracting plasmids for sequencing; (4) comparing the sequencing result with KIR / IPD database to prove the sequence is a characteristic gene sequence; and (5) designing the primer and probe by applying primer design software according to the characteristic KIR2DL2 gene sequence. The designed primer and probe are used for gene detection. The method can be used for improving the specificity and accuracyof PCR detection.

Description

technical field [0001] The invention belongs to the technical field of biological detection, and in particular relates to a fluorescent quantitative PCR detection method of KIR2DL2 mRNA and a detection kit thereof. Background technique [0002] Killer cell immunoglobulin-like receptor (KIR) is a transmembrane glycoprotein mainly expressed on the surface of NK cell membrane, which interacts with ligand HLA molecules to generate allogeneic NK cells, exerts GVL effect, and plays a role in hematopoietic stem cells Play an important role in the prognosis of transplantation. KIR is located on human chromosome 19q13.4, contains 2 (KIR2D) or 3 (KIR3D) extracellular Ig-like domains, and is divided into two types: inhibitory KIR (inhibitory KIR, iKIR) and activating KIR (activating KIR, aKIR) , iKIR genes include KIR2DL1, KIR 2DL2, KIR3DL1, etc. with heterologous reactive functions, and aKIR genes include KIR2DS1, KIR2DS2, KIR2DL2, KIR2DS4, KIR2DS5, KIR3DS1, etc. The DNA sequences o...

AI Technical Summary

Problems solved by technology

The DNA sequences of KIR genes are 80-90% identical. At present, for primer design, most of the target gene sequences are found on NCBI, or the gene is sequenced, and then the primers are designed according to the gene sequence through primer design software. Perform blast; these methods have certain limitations when designing primers for KIR genes, mainly manifested in low efficiency, poor specificity, and low accuracy

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