Provided herein are methods of selective screening. In addition, various targeting proteins and sequences, as well as methods of their use, are also provided.

Among other things, a method for normalizing a sample is provided. In some embodiments, the method comprises: (a) reacting a sample with a limiting amount of a single-turnover sequence-specific endonuclease that recognizes a target sequence, thereby cleaving a portion of the nucleic acid molecules that comprise the target sequence and producing a normalized amount of a first cleavage product; and (b) isolating, transcribing or selectively amplifying the normalized amount of the first cleavage product. In this method, because a limiting amount of the endonuclease is used, the normalized amount of the first cleavage product is determined by the limiting amount of the first single-turnover sequence-specific endonuclease used in step (a).

Technology provided herein relates in part to methods, processes, machines and apparatuses for determining sequences of nucleotides for nucleic acid templates in a nucleic acid sample. The technology provide herein also relates in part to methods, processes, machines and apparatuses for counting nucleic acid templates. Nucleic acid templates of a sample are tagged with nonrandom oligonucleotide adapters that include predetermined non-randomly generated sequences. The use of these nonrandom oligonucleotide adapters provides an efficient method to reduce sequencing errors, and increase the sensitivity of detection of low-frequency single nucleotide alterations.

A method for making an enriched library comprising specific nucleic acid sequence information allowing to identifying at least one binding entity that binds to at least one target wherein the specific binding entity has been present in an in vitro display library.

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.

Described herein are methods, compositions, and systems derived from uncultivated microorganisms useful for gene editing.

Provided herein are methods of selective screening. In addition, various targeting proteins and sequences, as well as methods of their use, are also provided.

Methods and systems for sample preparation techniques that allow amplification (e.g., whole genome amplification) and sequencing of chromatin accessible regions of single cells are provided. The methods and systems generally operate by forming or providing partitions (e.g., droplets) including a single biological particle and a single bead comprising a barcoded oligonucleotide. The preparation of barcoded next-generation sequencing libraries prepared from a single cell is facilitated by the transposon-mediated transposition and fragmentation of a target nucleic acid sequence. The methods and systems may be configured to allow the implementation of single-operation or multi-operation chemical and/or biochemical processing within the partitions.

Methods for the automated template-free synthesis of user-defined sequence controlled biopolymers using microfluidic devices are described. The methods facilitate simultaneous synthesis of up to thousands of uniquely addressed biopolymers from the controlled movement and combination of regents as fluid droplets using microfluidic and EWOD-based systems. In some forms, biopolymers including nucleic acids, peptides, carbohydrates, and lipids are synthesized from step-wise assembly of building blocks based on a user-defined sequence of droplet movements. In some forms, the methods synthesize uniquely addressed nucleic acids of up to 1,000 nucleotides in length. Methods for adding, removing and changing barcodes on biopolymers are also provided. Biopolymers synthesized according to the methods, and libraries and databases thereof are also described. Modified biopolymers, including chemically modified nucleotides and biopolymers conjugated to other molecules are described.

The present inventors used aminoacylated tRNAs prepared by ligating a pCpA-amino acid to a tRNA lacking the 3-terminal CA sequence using a ligase, and thereby succeeded in discrimination between the NNA and NNG codons in specific codon boxes, which had been difficult to achieve due to the presence of wobble base pairing. Furthermore, the inventors assigned another amino acid to the NNU or NNC codon in the same codon box, and performed translation of a sequence containing the NNU, NNA, and NNG codons, or the NNC, NNA, and NNG codons to evaluate the codon discrimination ability. As a result, the codons of interest were specifically translated into only the corresponding amino acids, achieving accurate discrimination successfully. Furthermore, the inventors confirmed that similar effects can also be obtained even when the nucleotide sequence of the tRNA body is altered.