Described herein is a system and process for long read sequencing using PCR primers with incorporated Unique Molecular Identifiers (UMIs) for generating a single molecule consensus for each starting molecule in the sample population. This method reduces the sequencing error rate by generating a consensus from the individual reads in each UMI group, averaging out sequencing errors to give better confidence in the actual sequence, to allow for increased accuracy of quantifying the precise knock-in event, and reporting perfect HDR integration.

Methods for detection of a target nucleic acid in a sample are provided according to aspects of the present disclosure which include: providing a reaction mixture comprising a LAMP assay primer set specific for the target nucleic acid, magnesium, dNTPs, a reaction buffer, a DNA polymerase, and a sample to be tested for presence of the target nucleic acid, wherein the LAMP assay primer set comprises a forward inner primer (FIP), a backward inner primer (BIP), a forward outer primer (F3) and a backward outer primer (B3), wherein the FIP and BIP are present in a non-equal ratio such that molar concentration of BIP is highly skewed relative to molar concentration of FIP. The reaction mixture is incubated under amplification reaction conditions to produce a reaction product comprising amplified target nucleic acids. The amplified target nucleic acids are then detected.

Methods for detection of a target nucleic acid in a sample are provided according to aspects of the present disclosure which include: providing a reaction mixture comprising a LAMP assay primer set specific for the target nucleic acid, magnesium, dNTPs, a reaction buffer, a DNA polymerase, and a sample to be tested for presence of the target nucleic acid, wherein the LAMP assay primer set comprises a forward inner primer (FIP), a backward inner primer (BIP), a forward outer primer (F3) and a backward outer primer (B3), wherein the FIP and BIP are present in a non-equal ratio such that molar concentration of BIP is highly skewed relative to molar concentration of FIP. The reaction mixture is incubated under amplification reaction conditions to produce a reaction product comprising amplified target nucleic acids. The amplified target nucleic acids are then detected.

The present disclosure provides methods for determining the ploidy status of a chromosome in a gestating fetus from genotypic data measured from a mixed sample of DNA comprising DNA from both the mother of the fetus and from the fetus, and optionally from genotypic data from the mother and father. The ploidy state is determined by using a joint distribution model to create a plurality of expected allele distributions for different possible fetal ploidy states given the parental genotypic data, and comparing the expected allelic distributions to the pattern of measured allelic distributions measured in the mixed sample, and choosing the ploidy state whose expected allelic distribution pattern most closely matches the observed allelic distribution pattern. The mixed sample of DNA may be preferentially enriched at a plurality of polymorphic loci in a way that minimizes the allelic bias, for example using massively multiplexed targeted PCR.

The present disclosure provides methods for determining the ploidy status of a chromosome in a gestating fetus from genotypic data measured from a mixed sample of DNA comprising DNA from both the mother of the fetus and from the fetus, and optionally from genotypic data from the mother and father. The ploidy state is determined by using a joint distribution model to create a plurality of expected allele distributions for different possible fetal ploidy states given the parental genotypic data, and comparing the expected allelic distributions to the pattern of measured allelic distributions measured in the mixed sample, and choosing the ploidy state whose expected allelic distribution pattern most closely matches the observed allelic distribution pattern. The mixed sample of DNA may be preferentially enriched at a plurality of polymorphic loci in a way that minimizes the allelic bias, for example using massively multiplexed targeted PCR.

The invention concerns a method for detecting isothermal loop-mediated (LAMP) amplification of a target nucleic acid sequence which is based on the fluorescence resonance energy transfer (FRET) mechanism. The invention also concerns a set of oligonucleotides and a kit adapted for carrying out the LAMP-FRET method of the invention.

The invention concerns a method for detecting isothermal loop-mediated (LAMP) amplification of a target nucleic acid sequence which is based on the fluorescence resonance energy transfer (FRET) mechanism. The invention also concerns a set of oligonucleotides and a kit adapted for carrying out the LAMP-FRET method of the invention.

The present disclosure provides methods for determining the ploidy status of a chromosome in a gestating fetus from genotypic data measured from a mixed sample of DNA comprising DNA from both the mother of the fetus and from the fetus, and optionally from genotypic data from the mother and father. The ploidy state is determined by using a joint distribution model to create a plurality of expected allele distributions for different possible fetal ploidy states given the parental genotypic data, and comparing the expected allelic distributions to the pattern of measured allelic distributions measured in the mixed sample, and choosing the ploidy state whose expected allelic distribution pattern most closely matches the observed allelic distribution pattern. The mixed sample of DNA may be preferentially enriched at a plurality of polymorphic loci in a way that minimizes the allelic bias, for example using massively multiplexed targeted PCR.

The present disclosure provides methods for determining the ploidy status of a chromosome in a gestating fetus from genotypic data measured from a mixed sample of DNA comprising DNA from both the mother of the fetus and from the fetus, and optionally from genotypic data from the mother and father. The ploidy state is determined by using a joint distribution model to create a plurality of expected allele distributions for different possible fetal ploidy states given the parental genotypic data, and comparing the expected allelic distributions to the pattern of measured allelic distributions measured in the mixed sample, and choosing the ploidy state whose expected allelic distribution pattern most closely matches the observed allelic distribution pattern. The mixed sample of DNA may be preferentially enriched at a plurality of polymorphic loci in a way that minimizes the allelic bias, for example using massively multiplexed targeted PCR.

The present disclosure provides methods, devices and systems that enable simultaneous multiplexing amplification reaction and real-time detection in a single reaction chamber.