MLH1 V384D Mutation Detection Kit Using Specific Primers
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Solution Overview
Problem
Current cancer detection methods lack specificity and sensitivity, particularly in identifying the V384D mutation in the hMLH1 gene, which is a promising biomarker for various cancers and related to drug resistance in EGFR-TKI treatment for lung cancer.
Innovation Solution
A kit comprising specific primer sets and probes, such as those with sequences listed in SEQ ID NOs 06, 07, 08, 09, and 10, designed to target the V384 alteration of the MLH1 gene, utilizing DNA polymerase and DNA binding dyes like SYBR Green, for accurate identification of the V384D mutation through PCR and HRM assays.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If conventional primer sets and probes are used for cancer detection, then the detection process is simple, but the specificity and sensitivity for identifying V384D mutation are insufficient
Solution Approach 1:
The primer sets and probes are designed with specific local sequences that target the V384D mutation site in the hMLH1 gene. The primers contain sequences complementary to the mutated region, allowing selective binding and amplification only of samples containing the V384D mutation, thereby achieving high detection specificity and sensitivity without requiring complex overall system design
Solution Approach 2:
The detection system is divided into multiple functional components: forward primers, reverse primers, and probes, each with specific sequences targeting different aspects of the V384D mutation. This segmentation allows each component to be optimized independently for maximum specificity and sensitivity while maintaining overall system manageability
2Reliability
If early cancer detection is performed to improve survival rate, then the survival rate increases, but the detection methods must be highly accurate to avoid false results
Solution Approach 1:
The detection system incorporates control mechanisms where the specific primer-probe interactions provide feedback on the presence or absence of the V384D mutation. The fluorescent probes emit signals only when bound to the target sequence, providing real-time feedback that enhances detection reliability and allows for quality control assessment
Solution Approach 2:
The primer sets and probes are pre-designed and pre-tested to specifically recognize the V384D mutation sequence before clinical use. This preliminary optimization ensures that the detection system is ready to provide highly accurate results immediately upon application, without requiring complex runtime adjustments or calibration procedures
Applied Scientific Principles
This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.
Function Achieved in This Case
The kit enables more specific and sensitive detection of the V384D mutation, facilitating early cancer diagnosis and evaluation of EGFR-TKI treatment efficacy and progression-free survival, thereby informing follow-up treatment strategies.
Implementation Method 1
a first primer comprising a sequence of SEQ ID NO 06; and a second primer comprising a sequence of SEQ ID NO 07; and/or a first primer comprising a sequence of SEQ ID NO 08; and a second primer comprising a sequence of SEQ ID NO 09
Implementation Method 2
said kit further comprises a double strands DNA binding dye. Preferably, said double strands DNA binding dye is SYBR® Green I, SYBR® Green II, SYBR® Gold, Oxazole Yellow, Thiazole Orange, PicoGreen, EvaGreen®
Data Source
AI summary
The present invention provides a primer set and a kit containing same for analyzing a sequence of an MLH1 DNA, an MLH1 mRNA, and/or a cDNA sequence derived from said MLH1 mRNA of a subject to identify a V384 alteration of an MLH1 gene encoded protein.