Protected dye-labeled reagents

Patent metadata

Field	Value
Publication number	US-2017145495-A1
Application number	US-201615357965-A
Country	US
Kind code	A1
Filing date	Nov 21, 2016
Priority date	Nov 20, 2015
Publication date	May 25, 2017
Grant date	—

Abstract

Labeled nucleotide analogs comprising at least one avidin protein, at least one dye-labeled compound, and at least one nucleotide compound are provided. The analogs are useful in various fluorescence-based analytical methods, including the analysis of highly multiplexed optical reactions in large numbers at high densities, such as single molecule real time nucleic acid sequencing reactions. The analogs are detectable with high sensitivity at desirable wavelengths. They contain structural components that modulate the interactions of the analogs with DNA polymerase, thus decreasing photodamage and improving the kinetic and other properties of the analogs in sequencing reactions. Also provided are nucleotide and dye-labeled compounds of the subject analogs, as well as intermediates useful in the preparation of the compounds and analogs. Compositions comprising the compounds, methods of synthesis of the intermediates, compounds, and analogs, and mutant DNA polymerases are also provided.

First claim

1 - 79 . (canceled)

80 . A dye-labeled compound of structural formula (IIIG):

wherein
each L′ is independently a dye compound linker element;
each S is independently a shield element;
each Dye is independently either an acceptor dye or a donor dye;
each B″ is independently a terminal coupling element;
each p is independently 0 or 1; and
each r″ is independently an integer from 0 to 8;
s is an integer from 1 to 6; and
t is 0 or 1;

wherein the compound comprises at least one shield element, at least one acceptor dye, and at least one donor dye.

81 . The dye-labeled compound of claim 80 , wherein each r″ is independently an integer from 0 to 4.

82 . The dye-labeled compound of claim 80 , wherein s is an integer from 1 to 4.

83 . The dye-labeled compound of claim 80 comprising at least two donor dyes.

84 . The dye-labeled compound of claim 83 , wherein each donor dye is directly coupled to a donor shield element.

85 . The dye-labeled compound of claim 80 comprising at least two acceptor dyes.

86 . The dye-labeled compound of claim 85 , wherein each acceptor dye is directly coupled to an acceptor shield element.

87 . The dye-labeled compound of claim 80 comprising at least four donor dyes and at least two acceptor dyes.

88 . The dye-labeled compound of claim 80 , comprising at least two shield elements.

89 . The dye-labeled compound of claim 80 , wherein at least one dye compound linker element comprises a shield element or a side chain element.

90 . The dye-labeled compound of claim 80 , wherein the at least one shield element comprises a plurality of side chains.

91 . The dye-labeled compound of claim 90 , wherein at least one side chain has a molecular weight of at least 300.

92 . The dye-labeled compound of claim 90 , wherein all of the side chains have a molecular weight of at least 300.

93 . The dye-labeled compound of claim 90 , wherein at least one side chain comprises a polyethylene glycol.

94 . The dye-labeled compound of claim 90 , wherein at least one side chain comprises a negatively-charged component.

95 . The dye-labeled compound of claim 94 , wherein the negatively-charged component comprises a sulfonic acid.

96 . The dye-labeled compound of claim 90 , wherein at least one side chain comprises a substituted phenyl group.

97 . The dye-labeled compound of claim 96 , wherein the at least one side chain comprises the structure:

wherein each x is independently an integer from 1 to 6.

98 . The dye-labeled compound of claim 97 , wherein each x is independently an integer from 1 to 4.

99 . The dye-labeled compound of claim 90 , wherein at least one side chain comprises a triazole.

100 . The dye-labeled compound of claim 90 , wherein at least one side chain comprises the structure:

101 . The dye-labeled compound of claim 80 , wherein the at least one shield element comprises the structure:

wherein each y is independently an integer from 1 to 6.

102 . The dye-labeled compound of claim 80 , wherein the at least one shield element comprises the structure:

103 . The dye-labeled compound of claim 80 , wherein t is 1.

104 . The dye-labeled compound of claim 80 , wherein at least one B″ comprises a biotin moiety.

105 . The dye-labeled compound of claim 104 , wherein the at least one B″ comprises a bis-biotin moiety.

106 . The dye-labeled compound of claim 80 , wherein the at least one acceptor dye or the at least one donor dye is a cyanine dye.

107 . The dye-labeled compound of claim 80 , wherein the compound does not contain a nucleoside.

108 . A labeled reagent composition comprising an avidin protein and the dye-labeled compound of claim 80 .

Assignees

• Pacific Biosciences California Inc

Inventors

• Sebo Lubomir
• Osuna Honey
• Yue Stephen
• Lapin Yuri

Classifications

• G01N2021/6439

  with indicators, stains, dyes, tags, labels, marks · CPC title

• G01N21/6428

  Measuring fluorescence of fluorescent products of reactions or of fluorochrome labelled reactive substances, e.g. measuring quenching effects, using measuring "optrodes" (in vivo A61B5/00; immunoassay G01N33/53) · CPC title

• C07D495/04

  Ortho-condensed systems · CPC title

• C12Q1/6869 Primary

  Methods for sequencing · CPC title

• C09B69/105 Primary

  containing a methine or polymethine dye · CPC title