crRNA (Clustered Regularly Interspaced Short Palindromic Repeat-Derived Ribonucleic Acid) for specifically detecting exon mutation of human KRAS genes #2 and #3 based on CRISPR technique

Abstract

The invention discloses crRNA (Clustered Regularly Interspaced Short Palindromic Repeat-Derived Ribonucleic Acid) for specifically detecting exon mutation of human KRAS genes #2 and #3 based on a CRISPR technique. The crRNA is characterized by comprising a CRISPR-Cas13a system as well as crRNA, wherein the crRNA can be combined with the CRISPR-Cas13a system; and the crRNA has a sequence format of5'-Cas13a protein anchoring sequence-crRNA guide sequence-3'. The crRNA has the advantages that according to detection results, whether the human KRAS gene have mutation or not can be judged intuitively through fluorescent reading, high-flux sequencing can be avoided, and the crRNA has the advantages of being low in cost, possible in multi-time detection, high in detection speed, possible in direct result analysis, and the like, and is applicable to large-scale clinical sample detection; the nucleic acid testing technique disclosed by the invention is different from a conventional detection technique which is based on PCR (Polymerase Chain Reaction) techniques, and no complex temperature control instrument or system is needed in a whole reaction process; and three steps of reactions are carried out in a same reaction system, so that operation procedures can be further simplified, and nucleic acid fragments with specific sequences can be detected within two hours.

Description

technical field [0001] The invention relates to the detection of biological materials, in particular to a method suitable for rapid detection of specific nucleic acid fragments in biological samples based on CRISPR-Cas13a technology. Background technique [0002] Researching and diagnosing diseases at the genetic level is the key to realizing the medical concept of individualized precision medicine. Kras is a murine sarcoma virus oncogene. There are three genes in the ras gene family related to human tumors—H-ras , K-ras and N-ras are located on chromosome 11, 12 and 1, respectively. K-ras is also known as the p21 gene because it encodes a 21kD ras protein. Among the ras genes, K-Ras has the greatest impact on human cancer. It is like a molecular switch: when normal, it can control the pathways that regulate cell growth; when abnormal, it causes cells to continue to grow and prevents cells from self-destruction. It participates in intracellular signal transmission. When th...

AI Technical Summary

Problems solved by technology

[0003] At present, conventional gene detection is realized by PCR technology, but the existing PCR technology has some defects such as low sensitivity, low specificity, DNA is often used for the target fragment, etc., resulting in high cost, low repeatability, and long detection time And other issues

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