Methods and compositions are provided for amplifying a pool of oligonucleotides, such as dual guide oligonucleotide constructs comprising sequences encoding a first guide RNA segment and a sequence encoding a second guide RNA segment. An amplification mixture is formed comprising the pool of oligonucleotides, an amplification enzyme, deoxyribonucleotide triphosphates, and primers. The amplification mixture is thermocycled a sufficient number of times and under conditions to produce a library of oligonucleotide constructs. The present methods and compositions provide dual guide libraries, including libraries that are essentially free of scrambled library members.

The present disclosure methods for identifying binding partners using cell surface display libraries, where the cells of the library display engineered peptides on their cell surfaces for identification of peptides that bind to targets of interest. The engineered peptides are preferably expressed in the cells under conditions that provide both secretion and display of the engineered peptides on the cell surfaces, thus providing access of the engineered peptides to identify potential binding pairs. The cell libraries cab be engineered using an automated editing system that provides for one or more targeted edits per cell.

Disclosed herein is a novel method for multisite saturation mutagenesis. Provided in the instant disclosure is a means of generating genetic diversity at each desired codon position within the coding sequence of a gene using a multi-oligonucleotide primer pool and Gibson assembly approach where a mutant DNA library can be cloned into a target vector.

This application provides a bead with a covalently attached chemical compound and a covalently attached DNA barcode and methods for using such beads. The bead has many substantially identical copies of the chemical compound and many substantially identical copies of the DNA barcode. The compound consists of one or more chemical monomers, where the DNA barcode takes the form of barcode modules, where each module corresponds to and allows identification of a corresponding chemical monomer. The nucleic acid barcode can have a concatenated structure or an orthogonal structure. Provided are method for sequencing the bead-bound nucleic acid barcode, for cleaving the compound from the bead, and for assessing biological activity of the released compound.

This application provides a bead with a covalently attached chemical compound and a covalently attached DNA barcode and methods for using such beads. The bead has many substantially identical copies of the chemical compound and many substantially identical copies of the DNA barcode. The compound consists of one or more chemical monomers, where the DNA barcode takes the form of barcode modules, where each module corresponds to and allows identification of a corresponding chemical monomer. The nucleic acid barcode can have a concatenated structure or an orthogonal structure. Provided are method for sequencing the bead-bound nucleic acid barcode, for cleaving the compound from the bead, and for assessing biological activity of the released compound.

Disclosed herein is a novel method for multisite saturation mutagenesis. Provided in the instant disclosure is a means of generating genetic diversity at each desired codon position within the coding sequence of a gene using a multi-oligonucleotide primer pool and Gibson assembly approach where a mutant DNA library can be cloned into a target vector.

The present disclosure methods for identifying binding partners using cell surface display libraries, where the cells of the library display engineered peptides on their cell surfaces for identification of peptides that bind to targets of interest. The engineered peptides are preferably expressed in the cells under conditions that provide both secretion and display of the engineered peptides on the cell surfaces, thus providing access of the engineered peptides to identify potential binding pairs. The cell libraries cab be engineered using an automated editing system that provides for one or more targeted edits per cell.

Disclosed herein include systems, methods, compositions, and kits for whole transcriptome analysis (WTA) with random priming and extension (RPE). The RPE-based WTA method can comprise hybridizing random primers with a plurality of first strand barcoded polynucleotides associated with a solid support and extending the random primers to generate a plurality of extension products. The method can comprise amplifying the plurality of extension products to generate a sequencing library.

Disclosed herein include systems, methods, compositions, and kits for whole transcriptome analysis (WTA) with random priming and extension (RPE). The RPE-based WTA method can comprise hybridizing random primers with a plurality of first strand barcoded polynucleotides associated with a solid support and extending the random primers to generate a plurality of extension products. The method can comprise amplifying the plurality of extension products to generate a sequencing library.

The present disclosure provides instrumentation and automated methods for creating cell surface display libraries, where the cells of the library display engineered peptides on their cell surfaces for identification of binding pairs. The engineered peptides may be displayed using various cell surface display technologies.