A method including (a) providing an amplification reagent including an array of sites, and a solution having different target nucleic acids; and (b) reacting the amplification reagent to produce amplification sites each having a clonal population of amplicons from a target nucleic acid from the solution. The reacting can include simultaneously transporting the nucleic acids to the sites at an average transport rate, and amplifying the nucleic acids that transport to the sites at an average amplification rate, wherein the average amplification rate exceeds the average transport rate. The reacting can include producing a first amplicon from a nucleic acid that transports to each of the sites, and producing subsequent amplicons from the nucleic acid or from the first amplicon, wherein the average rate at which the subsequent amplicons are generated exceeds the average rate at which the first amplicon is generated.

A method including (a) providing an amplification reagent including an array of sites, and a solution having different target nucleic acids; and (b) reacting the amplification reagent to produce amplification sites each having a clonal population of amplicons from a target nucleic acid from the solution. The reacting can include simultaneously transporting the nucleic acids to the sites at an average transport rate, and amplifying the nucleic acids that transport to the sites at an average amplification rate, wherein the average amplification rate exceeds the average transport rate. The reacting can include producing a first amplicon from a nucleic acid that transports to each of the sites, and producing subsequent amplicons from the nucleic acid or from the first amplicon, wherein the average rate at which the subsequent amplicons are generated exceeds the average rate at which the first amplicon is generated.

A method of manufacturing and using a nanofluidic NAND transistor sensor array scheme including a plurality of nanopore channel pillars, a plurality of respective fluidic channels, a plurality of gate electrodes, a top chamber, and a bottom chamber includes placing a sensor substrate in an electrolyte solution comprising biomolecules and DNA. The method also includes placing first and second electrodes in the electrolyte solution (Vpp and Vss of the nanofluidic NAND transistor); forming the nanopore channel pillars; placing the gate electrodes and gate insulators in respective walls of the nanopore channel pillars; applying an electrophoretic bias in the first and second electrodes; applying a bias in the gate electrodes; detecting a change in an electrode current in the electrolyte solution caused by a change in a gate voltage; and detecting a change in a surface charge in nanopore channel electrodes in the respective fluidic channels.

A method of manufacturing and using a nanofluidic NAND transistor sensor array scheme including a plurality of nanopore channel pillars, a plurality of respective fluidic channels, a plurality of gate electrodes, a top chamber, and a bottom chamber includes placing a sensor substrate in an electrolyte solution comprising biomolecules and DNA. The method also includes placing first and second electrodes in the electrolyte solution (Vpp and Vss of the nanofluidic NAND transistor); forming the nanopore channel pillars; placing the gate electrodes and gate insulators in respective walls of the nanopore channel pillars; applying an electrophoretic bias in the first and second electrodes; applying a bias in the gate electrodes; detecting a change in an electrode current in the electrolyte solution caused by a change in a gate voltage; and detecting a change in a surface charge in nanopore channel electrodes in the respective fluidic channels.

Provided herein are methods for quantifying foreign cell-free DNA (cfDNA) via SNP profiling of low-volume blood sample. The methods allow for monitoring the status of organ transplant rejection through analysis of small volumes of patient capillary blood samples collected non-invasively with fingersticks or other devices. The methods also allow for guiding the dosage of immunosuppressant and for preparing for a new organ transplant in case of imminent organ failure.

The invention relates to a method for enriching a target polynucleotide sequence containing a genetic variation said method comprising: (a) providing two primers targeted to said target polynucleotide sequence; (b) providing a target specific xenonucleic acid clamp oligomer specific for a wildtype polynucleotide sequence; (c) generating multiple amplicons using PCR under specific temperature cycling conditions; and (d) detecting said amplicons. We introduce a novel molecule, Xenonucleic Acid (XNA) for the NGS library preparation. XNA is able to selectively suppress amplification of DNA with wild type alleles and amplify DNA containing mutant alleles. Mutants with low allelic frequency will be easily detectable without deep sequencing after enrichment by adding XNA in multiplex PCR. The 17 actionable mutants related to lung or colorectal cancer diseases at different variant allelic frequency (VAF) % were investigated. Clinical sensitivity is significantly improved with XNA in various types of samples.

This disclosure provides, among other things, methods for generating and applying therapeutic interventions. The methods involve, for example, (a) sequencing polynucleotides from cancer cells from a subject; (b) identifying and quantifying somatic mutations in the polynucleotides; (c) developing a profile of tumor heterogeneity in the subject indicating the presence and relative quantity of a plurality of the somatic mutations in the polynucleotides, wherein different relative quantities indicates tumor heterogeneity; and (d) determining a therapeutic intervention for a cancer exhibiting the tumor heterogeneity, wherein the therapeutic intervention is effective against a cancer having the profile of tumor heterogeneity determined.

This disclosure provides, among other things, methods for generating and applying therapeutic interventions. The methods involve, for example, (a) sequencing polynucleotides from cancer cells from a subject; (b) identifying and quantifying somatic mutations in the polynucleotides; (c) developing a profile of tumor heterogeneity in the subject indicating the presence and relative quantity of a plurality of the somatic mutations in the polynucleotides, wherein different relative quantities indicates tumor heterogeneity; and (d) determining a therapeutic intervention for a cancer exhibiting the tumor heterogeneity, wherein the therapeutic intervention is effective against a cancer having the profile of tumor heterogeneity determined.

A method for sensing an analyte utilizing a sensor having a working electrode, the method includes providing the working electrode with an analyte-specific enzyme and a redox mediator, providing the working electrode to the analyte, accumulating charge derived from the analyte reacting with the analyte-specific enzyme and the redox mediator for a set period of time, connecting the working electrode to circuit after the set period of time, and measuring the signal from the accumulated charge.

Provided are a method for constructing a library based on an RNA sample and uses thereof. The method includes: step 1 of subjecting the RNA sample to a reverse transcription reaction to obtain DNA-RNA hybrid strands; step 2 of performing reaction of the DNA-RNA hybrid strands with an endoribonuclease, a first DNA polymerase, a second DNA polymerase, and dATPs to obtain a double-stranded DNA added with dA-tail, where the first DNA polymerase has a 5′-3′ exonuclease activity and a 3′-5′ exonuclease activity, and the second DNA polymerase has no 3′-5′ exonuclease activity; step 3 of ligating the double-stranded DNA added with dA-tail and a sequencing adaptor to obtain a ligated product; and step 4 of subjecting the ligated product to PCR amplification to obtain a sequencing library.