CD74-ROS1 rearrangement DNA (deoxyribonucleic acid) standard substance for molecular diagnosis, RNA (ribonucleic acid) standard substance for molecular diagnosis and application of CD74-ROS1 rearrangement DNA standard substance and RNA standard substance

Abstract

The invention belongs to the technical field of gene rearrangement standard substances, and provides a CD74-ROS1 rearrangement DNA (deoxyribonucleic acid) standard substance for molecular diagnosis, an RNA (ribonucleic acid) standard substance for molecular diagnosis and application of the CD74-ROS1 rearrangement DNA standard substance and the RNA standard substance. The scheme provided by the invention comprises the following steps: constructing sgRNA of a No.6 intron aiming at a CD74 gene onto a vector pX330 to obtain a plasmid C4, constructing sgRNA of a No.33 intron aiming at an ROS1 gene onto the vector pX330 to obtain a plasmid R3, jointly transfecting a host cell with the plasmid C4 and the plasmid R3 to obtain a recombinant cell, and carrying out monoclonal treatment on the recombinant cell to obtain a cell for preparing a DNA standard substance or an RNA standard substance so as to obtain the DNA standard substance and the RNA standard substance. According to the CD74-ROS1 rearrangement standard substance sequence and cell provided by the invention, a sample can be stably provided for a long time, and the CD74-ROS1 rearrangement standard substance sequence and cell are very suitable for performance evaluation and long-term quality control requirements of LDT or IVD development.

Description

technical field [0001] The invention relates to the technical field of gene rearrangement standard products, in particular to a CD74-ROS1 rearrangement DNA standard product, RNA standard product and application thereof for molecular diagnosis. Background technique [0002] ROS proto-oncogene 1 (ROS1), encoded by the ROS1 gene located on chromosome 6q22.1, belongs to the insulin receptor subfamily of tyrosine kinases and was first discovered in 1986 in a study involving chicken tumor RNA UR2 tumor virus. The physiological role of ROS1 remains controversial; its expression is mainly observed in lung tissue, followed by cervix and colon. Conceivably, the tyrosine kinase insulin receptor plays a key role in embryonic development. The ROS1 protein shows substantial homology to ALK (both belonging to the insulin receptor superfamily), especially in the ATP-binding site (84% homology) and the kinase domain (64% homology). Fusion mutations in ROS1 are well known and often result i...

AI Technical Summary

Problems solved by technology

However, in the actual detection process, clinical samples are often hard to come by, and such patients need to be met just in time, and the samples are not reproducible, and the amount is small, so they are not suitable for long-term use as standards and quality controls for IVD kit development.

[0009] How to solve this contradiction, there are two methods, the first is to prepare the clinical samples containing the target CD74-ROS1 rearrangement into immortalized cell lines, but this preparation process often takes half a year to a year, and the success rate is not high. High, one patient corresponds to one type of rearrangement, and there is no diversity, so this method is basically ruled out and not preferred; the second method is to use gene recombination and gene editing on immortalized cell lines. The CD74-ROS1 rearrangement is introduced into the cell line to form a stable genome structure. In this way, the genome (DNA and RNA) containing the CD74-ROS1 rearrangement can be continuously and stably supplied, while also meeting the needs of diversity and solving The demand for standard and quality control products in the whole process of performance evaluation

However, there is little research on the development of CD74-ROS1 rearrangement standards

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