Compositions and techniques for nucleic acid primer extension

Abstract

A method of characterizing a nucleic acid that includes steps of (a) contacting a primer-template nucleic acid hybrid with a polymerase and a mixture of nucleotides to produce an extended primer hybrid and to form a stabilized ternary complex including the extended primer hybrid, the polymerase and a nucleotide cognate of the next base in the template, wherein the mixture contains nucleotide cognates of four different base types, wherein the nucleotide cognate of the first base type has a reversible terminator, and wherein nucleotide cognates of the second, third and fourth base types are extendable; (b) detecting the stabilized ternary complex to distinguish the next base from other base types in the template; and (c) determining the presence of a base multiplet in the template nucleic acid, the base multiplet including the first base type followed by the next base.

Description

CROSS-REFERENCES TO RELATED APPLICATIONS[0001]This application claims the benefit of U.S. Provisional Application No. 62 / 626,836, filed Feb. 6, 2018, which is incorporated herein by reference in its entirety and for all purposes.REFERENCE TO A “SEQUENCE LISTING,” A TABLE, OR A COMPUTER PROGRAM LISTING APPENDIX SUBMITTED AS AN ASCII FILE[0002]The Sequence Listing written in file 053195-505001US_Sequence Listing_ST25.txt, created Feb. 1, 2019, 1,865 bytes, machine format IBM-PC, MS Windows operating system, is hereby incorporated by reference.BACKGROUND[0003]The present disclosure relates generally to molecular analysis and diagnostics and has specific applicability to nucleic acid sequencing.[0004]The time required to sequence a human genome has dropped precipitously in the last decade. The procedure, which used to take several years and millions of dollars to perform, can now be completed in mere hours, for a few thousand dollars. Although the rate of improvement has been impressive...

AI Technical Summary

Benefits of technology

The present patent provides a method for characterizing a nucleic acid by using a polymerase and a mixture of nucleotides to produce an extended primer hybrid. The method can detect the presence of a base multiplet in the template nucleic acid, which is a reversible termination of the polymerase. The method can also determine the presence of a series of base multiplets in the template nucleic acid. The technical effect of this patent is to provide a reliable and accurate method for characterizing nucleic acids.

Problems solved by technology

Although the rate of improvement has been impressive, the currently available commercial methods are still unsatisfactory for many clinical applications.

Even a few days of delay in receiving such confirmation can cause a significant adverse toll on the emotional and psychological state of the patient.

Nevertheless, too many newborns die every year for lack of a timely diagnosis.

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