US 11,718,639 B2
Fluorescent polymerase enzyme substrates having protein shields
Keith Bjornson, Fremont, CA (US); Jeremiah Hanes, Woodside, CA (US); Erik Miller, Berkeley, CA (US); Satwik Kamtekar, Mountain View, CA (US); Lubomir Sebo, Hayward, CA (US); and Louis Brogley, Santa Cruz, CA (US)
Assigned to Pacific Biosciences of California, Inc., Menlo Park, CA (US)
Filed by Pacific Biosciences of California, Inc., Menlo Park, CA (US)
Filed on Apr. 14, 2021, as Appl. No. 17/230,007.
Application 17/230,007 is a continuation of application No. 16/012,428, filed on Jun. 19, 2018, granted, now 11,014,958.
Application 16/012,428 is a continuation of application No. 14/727,553, filed on Jun. 1, 2015, granted, now 10,023,605.
Application 14/727,553 is a continuation of application No. 13/767,619, filed on Feb. 14, 2013, granted, now 9,062,091.
Claims priority of provisional application 61/599,149, filed on Feb. 15, 2012.
Prior Publication US 2021/0347808 A1, Nov. 11, 2021
This patent is subject to a terminal disclaimer.
Int. Cl. C07H 21/04 (2006.01); C07H 19/06 (2006.01); C07H 19/16 (2006.01); G01N 33/58 (2006.01); C07K 14/36 (2006.01); C12Q 1/6874 (2018.01); C12N 9/10 (2006.01); C09B 23/08 (2006.01); C09B 69/10 (2006.01); C12Q 1/6869 (2018.01)
CPC C07H 21/04 (2013.01) [C07H 19/06 (2013.01); C07H 19/16 (2013.01); C07K 14/36 (2013.01); C09B 23/083 (2013.01); C09B 69/105 (2013.01); C12N 9/10 (2013.01); C12Q 1/6869 (2013.01); C12Q 1/6874 (2013.01); G01N 33/582 (2013.01); G01N 33/583 (2013.01); C12Q 2523/313 (2013.01); C12Q 2535/113 (2013.01); C12Q 2563/107 (2013.01); C12Q 2563/131 (2013.01)] 13 Claims
 
1. A method of making a labeled nucleotide analog comprising:
(a) providing an avidin protein having four subunits, each subunit comprising one biotin binding site;
(b) adding a nucleotide component, the nucleotide component comprising one or more phospholinked nucleotide moieties covalently connected to a bis-biotin moiety, whereby the bis-biotin moiety binds to two of the four subunits on the avidin protein, and
(c) adding a dye component comprising one or more dye moieties covalently connected to a bis biotin moiety, whereby this bis-biotin moiety binds to the other two subunits on the avidin protein thereby forming a nucleotide analog.