US 12,110,498 B2
Compositions and methods for genomic integration of nucleic acids into exogenous landing pads
Kevin George, Oakland, CA (US); Andrew Main, Benicia, CA (US); and Chia-Hong Tsai, Martinez, CA (US)
Assigned to AMYRIS, INC., Emeryville, CA (US)
Filed by AMYRIS, INC., Emeryville, CA (US)
Filed on Mar. 9, 2022, as Appl. No. 17/690,429.
Application 17/690,429 is a division of application No. 16/302,079, granted, now 11,293,033, previously published as PCT/US2017/033369, filed on May 18, 2017.
Claims priority of provisional application 62/338,412, filed on May 18, 2016.
Prior Publication US 2022/0251606 A1, Aug. 11, 2022
Int. Cl. C12N 15/70 (2006.01); C12N 1/19 (2006.01); C12N 15/10 (2006.01); C12N 15/81 (2006.01); C12N 15/90 (2006.01)
CPC C12N 15/905 (2013.01) [C12N 15/102 (2013.01); C12N 15/1082 (2013.01); C12N 15/81 (2013.01)] 17 Claims
 
1. A method for integrating one or more exogenous donor nucleic acids into a yeast host cell's genome, the method comprising:
contacting a host cell, the host cell comprising one or more exogenous landing pads integrated in the host cell's genome, wherein each exogenous landing pad comprises a nuclease target sequence (NTS) positioned between an upstream landing pad homology sequence (ULP) and a downstream landing pad homology sequence (DLP), with:
(i) one or more exogenous donor nucleic acids (ES), comprising a nucleic acid of interest (D) positioned between an upstream library sequence (UL) and a downstream library sequence (DL), wherein each (UL) is capable of homologously recombining at any (ULP), and each (DL) is capable of homologously recombining at any (DLP) of any of the one or more exogenous landing pads
wherein any of the exogenous donor nucleic acids (ES) is integrated at any of the exogenous landing pads and is independent of genomic sequences surrounding each landing pad, and
wherein the one or more exogenous landing pads are selected from SEQ ID NOs: 8, 9, and 10.