US 11,851,663 B2
Single-vector type I vectors
Virginia Martinez, Copenhagen Ø (DK); Ruben Vazquez-Uribe, Copenhagen Ø (DK); Adam Takos, Copenhagen Ø (DK); and Eric Van Der Helm, Copenhagen Ø (DK)
Assigned to SNIPR BIOME APS, Copenhagen Ø (DK)
Filed by SNIPR Biome ApS, Copenhagen Ø (DK)
Filed on Nov. 27, 2018, as Appl. No. 16/201,736.
Claims priority of application No. 1816700 (GB), filed on Oct. 14, 2018; and application No. 1817509 (GB), filed on Oct. 27, 2018.
Prior Publication US 2020/0115716 A1, Apr. 16, 2020
This patent is subject to a terminal disclaimer.
Int. Cl. C12N 15/70 (2006.01); C12N 9/22 (2006.01); C12N 15/11 (2006.01); A61K 38/46 (2006.01); A61K 31/7088 (2006.01); C07K 14/33 (2006.01); A61P 3/04 (2006.01); A61P 31/04 (2006.01)
CPC C12N 15/70 (2013.01) [A61K 31/7088 (2013.01); A61K 38/465 (2013.01); A61P 3/04 (2018.01); C07K 14/33 (2013.01); C12N 9/22 (2013.01); C12N 15/11 (2013.01); C12N 2310/20 (2017.05); C12N 2800/80 (2013.01); C12N 2820/002 (2013.01); C12N 2820/007 (2013.01); C12N 2820/55 (2013.01); C12N 2830/005 (2013.01)] 26 Claims
 
1. A production strain bacterial cell comprising a nucleic acid vector for introduction into a target bacterial host cell for expression of Type I Cas3 and Cascade proteins in the target bacterial host cell, the vector comprising a first nucleotide sequence encoding a Type I Cas3 and a second nucleotide sequence encoding one or more cognate Cascade proteins, wherein the first nucleotide sequence is under the control of a promoter for controlling the expression of Type I Cas3 in the target bacterial host cell, wherein the promoter has a strength that is weaker than the Anderson Score strength of promoter BBa_J23108,
wherein the target bacterial host cell is selected from the group consisting of Fusobacteria, Bacteroides, Staphylococcus, Clostridium, Lactobacillus, Bacillus, Escherichia, Streptococcus, Streptomyces, Pseudomonas, and Klebsiella,
wherein the nucleic acid vector further comprises: (i) a CRISPR array for producing crRNAs in the target bacterial host cell; or (ii) one or more nucleotide sequences encoding one or more guide RNAs (gRNA), wherein the crRNAs or gRNAs each comprise a spacer sequence complementary to a target sequence of the target bacterial host cell, and
wherein the production strain bacterial cell does not comprise a crRNA or gRNA operable with the Cas3 to target and cut a chromosomal sequence of the production strain cell.