Kit for detecting gene expression of cytokeratin 18

Abstract

The invention relates to a cytokeratin 18 gene expression detection kit and a detection method thereof. The cytokeratin 18 gene expression detection kit comprises a capture probe aiming at CK18 gene mRNA and a signal amplification system, the capture probe is a stem-ring-shaped gamma PNA-PNA probe, which sequentially comprises a stem structure sequence, a P1 sequence, a Linker sequence and a P2 sequence from the 5' end to the 3' end; and the signal amplification system comprises a marking probe SP1 and a marking probe SP2, wherein each labeled probe SP1 sequentially comprises a P3 sequence, a spacer sequence and a P4 sequence from the 5' end to the 3' end; the 5' end of the P3 sequence is modified with a fluorophore, each labeled probe SP2 sequentially comprises a P5 sequence, a spacer sequence and a P6 sequence from the 5' end to the 3' end, and the 3' end of the P6 sequence is modified with a fluorophore. The kit has the advantages of high sensitivity, strong specificity, high accuracy, short detection time and the like.

Description

technical field [0001] The invention relates to the field of molecular biology, in particular to a cytokeratin 18 gene expression detection kit. Background technique [0002] Cytokeratin 18 (CK18) is a type I acidic intermediate filament protein belonging to the cytoskeleton protein family. Its gene is located on human chromosome 12q13.13 and contains 8 exons. CK18 is mainly expressed in monolayer or "simple" epithelial tissues, such as liver, lung, kidney, pancreas, gastrointestinal tract, breast, etc., and is also expressed in cancers arising from these tissues (WengYR et al., MolCancer Res, 2012, 10:485–493). The known biological function of CK18 is to provide a flexible intracellular scaffold to build the cytoplasm, resist external pressure on the cell, and maintain normal mitochondrial structure (WengYR et al., Mol Cancer Res, 10:485–493) . CK18 also plays an important role in processes such as apoptosis, mitosis, cell cycle progression and cell signal transduction. ...

AI Technical Summary

Problems solved by technology

Immunohistochemistry detects the expression of CK18 at the protein level; however, the limitations of this method are: the tissue samples required for the detection are limited, and the accuracy of the detection results is limited by the type of antibody, antigen retrieval method, staining method and conditions, etc. Influenced by factors, result interpretation is highly subjective

Real-time quantitative PCR is mainly used to detect the expression level of CK18 gene in tissue / cell samples, which has the advantages of high sensitivity and strong sequence specificity; however, this method has high requirements on experimental conditions, and contamination must be strictly avoided, otherwise it is difficult to guarantee the detection results accuracy and reliability

In view of the above problems, Chinese patent CN201410228511.9 provides an RNA in situ hybridization method for detecting gene expression. The detection probe of the method can realize the fluorescence signal amplification of RNA in situ detection and improve the sensitivity of detection, but further research It was found that the hybridization selectivity of the capture probe and the signal amplification system in the above RNA in situ hybridization detection method need to be further optimized to achieve better detection results

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