Reagents for reversibly terminating primer extension

Details Reagents for reversibly terminating primer extension Reagents for reversibly terminating primer extension

2009-03-27

2011-10-11

Olson, Eric S (Department: 1623)

Organic compounds -- part of the class 532-570 series

Organic compounds

Carbohydrates or derivatives

C536S025300, C536S025310, C536S025600, C536S026100, C536S026200, C536S026260

Reexamination Certificate

active

08034923

ABSTRACT:
Processes are disclosed that use 3′-reversibly terminated nucleoside triphosphates to analyze DNA for purposes other than sequencing using cyclic reversible termination. These processes are based on the unexpected ability of terminal transferase to accept these triphosphates as substrates, the unexpected ability of polymerases to add reversibly and irreversibly terminated triphosphates in competition with each other, the development of cleavage conditions to remove the terminating group rapidly, in high yield, and without substantial damage to the terminated oligonucleotide product, and the ability of reversibly terminated primer extension products to capture groups. The presently preferred embodiments of the disclosed processes use a triphosphate having its 3′-OH group blocked as a 3′-ONH2group, which can be removed in buffered NaNO2and use variants of Taq DNA polymerase, including one that has a replacement (L616A).

REFERENCES:
patent: 6214551 (2001-04-01), Sanghvi et al.
patent: 6576752 (2003-06-01), Manoharan et al.
patent: 7173125 (2007-02-01), Schwartz et al.
Trevisiol et al., “Synthesis of Nucleoside Triphosphates that Contain an Aminooxy Function for ‘Post-Amplification Labelling’” European Journal of Organic Chemistry (2000) pp. 211-217.

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