CFTR Gene Mutation Detection via Segmented PCR and Hybridization
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Solution Overview
Problem
Current methods for diagnosing cystic fibrosis are limited by the inability to effectively identify novel mutations in the CFTR gene, which are essential for accurate diagnosis and genetic counseling.
Innovation Solution
The discovery and detection of new CFTR gene mutations, including specific nucleotide changes and deletions, using methods such as PCR, sequencing, and oligonucleotide ligation, allowing for the identification of individuals predisposed to cystic fibrosis or associated conditions.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If conventional diagnostic methods are used for CFTR gene mutations, then existing known mutations can be detected, but novel mutations cannot be effectively identified
Solution Approach 1:
The patent applies universality by creating a comprehensive screening method that can detect both known and novel CFTR mutations through a combination of PCR amplification, restriction enzyme digestion, and sequencing. The method is designed to be universally applicable across different mutation types including deletions, insertions, and point mutations, rather than being limited to specific pre-known mutations.
Solution Approach 2:
The patent segments the CFTR gene into multiple regions for targeted analysis. By dividing the gene into specific exons and introns that are prone to mutations, the method enables systematic detection of novel mutations in each segment while maintaining overall gene coverage. This segmentation allows for more thorough screening compared to analyzing the entire gene at once.
2Measurement precision
If comprehensive CFTR gene sequencing is performed to identify all mutations, then diagnostic accuracy improves, but detection complexity and time increase
Solution Approach 1:
The patent reduces complexity by segmenting the CFTR gene into specific regions of interest (exons 2-10, introns 2-10, and specific splice sites) rather than sequencing the entire 250,000 base pair gene. This targeted approach maintains high diagnostic accuracy for clinically relevant mutations while significantly reducing the complexity and time required compared to whole-gene sequencing.
Solution Approach 2:
The patent applies partial action by focusing sequencing efforts on specific high-risk regions of the CFTR gene where mutations are most commonly found and have the greatest clinical impact. Rather than attempting to detect all possible mutations throughout the entire gene, the method concentrates resources on the most diagnostically important segments, achieving sufficient accuracy for clinical diagnosis without excessive complexity.
3Loss of information
If traditional mutation screening methods are used, then known mutations can be detected, but the ability to provide comprehensive genetic counseling information is limited
Solution Approach 1:
The patent applies preliminary action by performing PCR amplification and restriction enzyme digestion as preparatory steps before sequencing. This preliminary processing enriches the sample for mutation detection and removes non-informative sequences, thereby preserving more relevant genetic information while maintaining efficient workflow. The method prepares the genetic material in advance to maximize information yield from subsequent sequencing.
Solution Approach 2:
The patent uses restriction enzymes as intermediaries to facilitate mutation detection. These enzymes selectively cut DNA at specific recognition sequences, creating patterns that reveal the presence of mutations. This intermediary step translates complex genetic information into detectable physical patterns, preserving genetic information in a form that can be efficiently analyzed and interpreted for genetic counseling.
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
Enables precise determination of CFTR gene mutations, improving diagnostic accuracy and providing valuable information for genetic counseling and risk assessment of cystic fibrosis and related conditions.
Implementation Method 1
contacting the sample with a substantially purified nucleic acid (an oligonucleotide probe) that hybridizes to a mutant CFTR sequence
Implementation Method 2
amplifying said CFTR gene segments by a method such as, but not limited to, the polymerase chain reaction
Data Source
AI summary
The present invention provides novel mutations of the CFTR gene related to cystic fibrosis or to conditions associated with cystic fibrosis. Also provided are probes for detecting the mutant sequences. Methods of identifying if an individual has a genotype containing one or more mutations in the CFTR gene are further provided.