METHODS AND SYSTEMS FOR DETECTION OF A GENETIC MUTATION

摘要

Methods and systems for the detection of genetic mutations from a tissue sample (e.g., a preserved tissue sample) are provided. The method includes the steps of a) extracting a nucleic acid from a tissue or biological sample; b) preparing a targeted nucleic acid amplicon library from the extracted nucleic acid; c) sequencing the targeted nucleic acid amplicon library to produce tissue sample target nucleic acid sequence data; and d) analyzing the sample target nucleic acid sequence data to determine whether it contains a mutation (e.g., a mutation associated with a risk for a particular disease). The methods described herein advantageously can be performed in less than 36 hours.

主权项

1. A method for extracting nucleic acid from a preserved tissue sample, the method comprising the steps of:
(a) incubating the preserved tissue sample with a tissue digestion solution to form a tissue digestion mixture, wherein the tissue digestion solution is selected from the group consisting of:
(i) a tissue digestion solution comprising NaCl at a concentration of 10 mM to 140 mM, Na2HPO4 at a concentration of 0.5 mM to 10 mM, KH2PO4 at a concentration of 0.1 mM to 5 mM, and Tween 20;(ii) a tissue digestion solution comprising NaCl at a concentration of 10 mM to 140 mM, Na2HPO4 at a concentration of 0.5 mM to 10 mM, KH2PO4 at a concentration of 0.1 mM to 5 mM, and Triton-X100;(iii) a tissue digestion solution comprising NaCl at a concentration of 10 mM to 140 mM, Na2HPO4 at a concentration of 0.5 mM to 10 mM, and KH2PO4 at a concentration of 0.1 mM to 5 mM;(iv) a tissue digestion solution comprising TAPS sodium salt at a concentration of 0.5 mM to 25 mM, DTT at a concentration of 0.05 mM to 5 mM, and KCl at a concentration of 0.2 mM to 200 mM;(v) a tissue digestion solution comprising HEPES buffer at a concentration of 1 mM to 100 mM;(vi) a tissue digestion solution comprising HEPES buffer at a concentration of 1 mM to 100 mM and Triton-X100;(vii) a tissue digestion solution comprising HEPES buffer at a concentration of 1 mM to 100 mM and Tween 20;(viii) a tissue digestion solution comprising TAPS sodium salt at a concentration of 0.5 mM to 25 mM, DTT at a concentration of 0.05 mM to 5 mM, KCl at a concentration of 0.2 mM to 200 mM, and Triton-X100;(ix) a tissue digestion solution comprising a TAPS sodium salt at a concentration of 0.5 mM to 25 mM, DTT at a concentration of 0.05 mM to 5 mM, KCl at a concentration of 0.2 mM to 200 mM, and Tween 20; and(x) a tissue digestion solution comprising a TAPS sodium salt at a concentration of 0.5 mM to 25 mM, KCl at a concentration of 0.2 mM to 200 mM, β-Mercaptoethanol at a concentration of 0.1 mM to 1 mM, and Triton-X100, (b) heating the tissue digestion mixture at 80 to 110° C. for 1-30 minutes; (c) adding a protease solution comprising a proteinase to the tissue digestion mixture to form a protein degradation mixture and incubating the protein degradation mixture at 50 to 70° C. for 1-30 minutes; and (d) incubating the protein degradation mixture at 80 to 110° C. for 1-30 minutes; thereby extracting nucleic acid from the preserved tissue sample.