Compositions and methods for highly efficient genetic screening using barcoded guide rna constructs
AssigneesPeking University, Edigene Biotechnology Inc.
InventorsWensheng WEI, Shiyou ZHU, Zhongzheng CAO, Zhiheng LIU, Yuan HE, Pengfei YUAN
Life of this patent
Estimating the term…
Abstract
Compositions, kits and methods are provided for genetic screening using one or more sets of guide RNA constructs having internal barcodes (“iBAR”). Each set has three or more guide RNA constructs targeting the same genomic locus, but embedded with different iBAR sequences.
Claims40
A set of sgRNAiBAR constructs comprising three or more sgRNAiBAR constructs each comprising or encoding an sgRNAiBAR, wherein each sgRNAiBAR has an sgRNAiBAR sequence comprising a guide sequence and an internal barcode (iBAR) sequence, wherein each guide sequence is complementary to a target genomic locus, wherein the guide sequences for the three or more sgRNAiBAR constructs are the same, wherein the iBAR sequence for each of the three or more sgRNAiBAR constructs is different from each other, and wherein each sgRNAiBAR is operable with a Cas protein to modify the target genomic locus.
A method of preparing an sgRNAiBAR library comprising a plurality of sets of sgRNAiBAR constructs, wherein each set corresponds to one of a plurality of guide sequences complementary to different target genomic loci, wherein the method comprises:
- a) designing three or more sgRNAiBAR constructs for each guide sequence, wherein each sgRNAiBAR construct comprises or encodes an sgRNAiBAR having an sgRNAiBAR sequence comprising the corresponding guide sequence and an iBAR sequence, wherein the iBAR sequence corresponding to each of the three or more sgRNAiBAR constructs is different from each other, and wherein each sgRNAiBAR is operable with a Cas protein to modify the corresponding target genomic locus;
- and
- b) synthesizing each sgRNAiBAR construct, thereby producing the sgRNAiBAR library.
Dependencies are inferred from the claim text.