Compositions and methods for chemical cleavage and deprotection of surface-bound oligonucleotides

By using palladium or nickel complex for chemical cleavage in DNA sequencing, the problems of high stability and cost in enzymatic methods are solved, efficient linearization and deprotection are achieved, and background strength is reduced.

CN119684383APending Publication Date: 2025-03-25ILLUMINA INC +1
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Patent Information

Application Number
CN202411878056.7
Authority / Receiving Office
CN · China
Patent Type
Applications(China)
Current Assignee / Owner
Priority Date
2019-01-03
Filing Date
2019-05-14
Publication Date
2025-03-25

AI Technical Summary

Technical Problem

In the prior art, enzymatic methods have high stability, cost, storage and processing requirements in DNA sequencing applications, as well as high background intensity in sequencing readouts, and need to develop alternative linearization and deprotection methods.

Method used

Linearize and deprotect polynucleotides by providing a solid support containing a double-stranded polynucleotide, chemical cleavage with a palladium or nickel complex. The method includes immobilizing the double-stranded polynucleotide on a solid support, performing chemical cleavage in the presence of a transition metal complex, removing the cleaved first nucleic acid, and sequencing and resynthetic of the second strand, if necessary.

Benefits of technology

Effective linearization and deprotection in DNA sequencing are achieved, reducing enzyme dependence, improving the stability and efficiency of the reaction, and reducing background strength.

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Abstract

Embodiments of the present disclosure relate to methods of making templates for polynucleotide sequencing. In particular, the present disclosure relates to linearizing clustered polynucleotides prepared for sequencing by cleaving one or more first strands of a double-stranded polynucleotide immobilized on a solid support with a transition metal complex (e.g., a palladium complex or a nickel complex). Further disclosure relates to linearization of clustered polynucleotides by cleaving one or more second strands of a double-stranded polynucleotide comprising an azobenzene linker immobilized on a solid support using Na2S2O4. Also disclosed are nucleotides and oligonucleotides comprising 3 'phosphate capping groups, as well as methods of removing them using fluoride reagents.
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