A method for quantitating a plurality of nucleic acid molecules is provided. The method includes contacting a plurality of detectably-labeled probes and a plurality of extension primers with the plurality of nucleic acid molecules, each detectably-labeled probe including a first labeled nucleic acid domain having a first label; producing an extension product of each of the plurality of target nucleic acid fragments by extending a respective one of the plurality of extension primers with a polymerase; hydrolyzing each of the plurality of detectably-labeled probes hybridized to the respective one of the target nucleic acid fragments during extending the respective one of the plurality of extension primers with the polymerase; detecting a first signal produced as a result of hydrolyzing the plurality of detectably-labeled probes; and calculating a number of the plurality of target nucleic acid fragments based on signals detected upon a single cycle of extension reactions.

The present disclosure provides a transposon system comprising: i) a nucleotide sequence encoding polypeptides that form a CRISPR-associated transposase complex; ii) a nucleotide sequence encoding a guide RNA; and iii) a transposon, or an insertion site for a transposon, flanked by CAST complex recognition sites. The present disclosure provides a prokaryotic cell comprising a subject transposon system. The transposon system is useful for editing the genome of a target prokaryotic cell. The present disclosure provides methods for editing the genome of a target prokaryotic cell. The present disclosure further provides systems and methods for identifying, within a heterogeneous population of prokaryotic cells, prokaryotic species that are susceptible to genetic modification and gene editing.

The present invention provides methods to barcode nucleic acid for detection and sequencing. It applies a barcode template in a compartment with various targets, including nucleic acid fragments, nuclei and/or cells. After clonal amplification within the compartment, barcode sequence will integrate into its targets before the compartment is broken so that it will effectively barcode nucleic acid fragments originated from a nucleic acid fragment, a nucleus or a cell clonally. The barcode information can be used for tracking the origin of the fragment, nucleus or cell and be used for haplotype phasing and a variety of single cell-based applications N including whole genome sequencing, targeted sequencing, RNA sequencing and immune repertoire sequencing.

The present invention provides methods to barcode nucleic acid for detection and sequencing. It applies a barcode template in a compartment with various targets, including nucleic acid fragments, nuclei and/or cells. After clonal amplification within the compartment, barcode sequence will integrate into its targets before the compartment is broken so that it will effectively barcode nucleic acid fragments originated from a nucleic acid fragment, a nucleus or a cell clonally. The barcode information can be used for tracking the origin of the fragment, nucleus or cell and be used for haplotype phasing and a variety of single cell-based applications N including whole genome sequencing, targeted sequencing, RNA sequencing and immune repertoire sequencing.

Methods and compositions for digital profiling of nucleic acid sequences present in a sample are provided.

The present disclosure provides compositions, methods, systems, and devices for polynucleotide processing. Such polynucleotide processing may be useful for a variety of applications, including polynucleotide sequencing.

A population of nucleic acid adaptors is provided. In some embodiments, the population contains at least 50,000 different molecular barcode sequences, where the barcode sequences are double-stranded and at least 90% of the barcode sequences have an edit distance of at least 2. In certain cases, the adaptor may have an end in which the top and bottom strands are not complementary (i.e., may be in the form of a Y-adaptor). In some embodiments and depending on the how the adaptor is going to be employed, the other end of the adaptor may have a ligatable end or may be a transposon end sequence.

A population of nucleic acid adaptors is provided. In some embodiments, the population contains at least 50,000 different molecular barcode sequences, where the barcode sequences are double-stranded and at least 90% of the barcode sequences have an edit distance of at least 2. In certain cases, the adaptor may have an end in which the top and bottom strands are not complementary (i.e., may be in the form of a Y-adaptor). In some embodiments and depending on the how the adaptor is going to be employed, the other end of the adaptor may have a ligatable end or may be a transposon end sequence.

A method for analysing insertion sites of an exogenous nucleotide sequence in a subject's genome, wherein the method comprises: (a) providing a sample from the subject comprising cell-free double-stranded DNA polynucleotides; (b) blunting the ends of the polynucleotides; (c) ligating an oligonucleotide to both ends of the polynucleotides; (d) amplifying polynucleotides comprising an insertion site; and (e) sequencing the product of step (d).

Systems and methods for enhancing RNA translatability and stability are disclosed. Some embodiments describe RNA molecules exhibiting increased translatability and/or stability. Additional embodiments describe methods for screening RNA molecules for increased translatability and/or stability. Various embodiments utilize screening methods, including degenerative sequences to identify sequences or regions that increase the translatability and/or stability of RNA molecules.