The present disclosure relates to methods of assessing a sample of guide RNAs (gRNAs).

The disclosure provides a synthetic standard which includes polynucleotides (e.g., DNA or RNA) containing multiple clinically important germline and somatic variants. These materials are utilized to calibrate, evaluate, and/or validate the performance of polynucleotide-based genetic analysis assays, such as NGS assays. In one aspect the disclosure provides a method for validating assay performance including generating synthetic variant DNA fragments comprising variants with known allele frequencies, wherein the fragments comprise a molecular tag; combining the synthetic variant DNA with wild-type DNA to create test samples; preparing one or more dilutions of the test samples; performing an assay of interest on the one or more dilutions of test samples; and comparing the outcome of the assay with the test samples with known allele frequencies of interest, thereby validating the performance of the assay.

The disclosure provides a synthetic standard which includes polynucleotides (e.g., DNA or RNA) containing multiple clinically important germline and somatic variants. These materials are utilized to calibrate, evaluate, and/or validate the performance of polynucleotide-based genetic analysis assays, such as NGS assays. In one aspect the disclosure provides a method for validating assay performance including generating synthetic variant DNA fragments comprising variants with known allele frequencies, wherein the fragments comprise a molecular tag; combining the synthetic variant DNA with wild-type DNA to create test samples; preparing one or more dilutions of the test samples; performing an assay of interest on the one or more dilutions of test samples; and comparing the outcome of the assay with the test samples with known allele frequencies of interest, thereby validating the performance of the assay.

A replacement fluid suitable to replace a PCR Mastermix, wherein the replacement fluid is different from the PCR Mastermix. The replacement fluid has pipetting characteristics substantially equivalent to the PCR Mastermix, wherein for a desired pipetted volume of the target fluid delivered by a pipette, a pipetted volume of the replacement fluid delivered by the pipette is substantially equivalent to the desired pipetted volume of the PCR Mastermix. The replacement fluid is a primary equivalent fluid having substantially similar pipetting characteristics. One or more additives may be added to the primary equivalent fluid. The one or more additives may include pipettability modifying additives and non pipettability modifying additives.

A replacement fluid suitable to replace a PCR Mastermix, wherein the replacement fluid is different from the PCR Mastermix. The replacement fluid has pipetting characteristics substantially equivalent to the PCR Mastermix, wherein for a desired pipetted volume of the target fluid delivered by a pipette, a pipetted volume of the replacement fluid delivered by the pipette is substantially equivalent to the desired pipetted volume of the PCR Mastermix. The replacement fluid is a primary equivalent fluid having substantially similar pipetting characteristics. One or more additives may be added to the primary equivalent fluid. The one or more additives may include pipettability modifying additives and non pipettability modifying additives.

Nucleic acid reagents and corresponding methods of using the same for monitoring and evaluating nucleic acid extraction and amplification reactions.

Nucleic acid reagents and corresponding methods of using the same for monitoring and evaluating nucleic acid extraction and amplification reactions.

Aspects of the present disclosure include methods of detecting the presence or absence of one or more diseases using quantitative approaches. Aspects of the present disclosure include methods for determining the abundance of endogenous targets. Aspects of the present disclosure also include determining the presence or absence of an aneuploidy.

Aspects of the present disclosure include methods of detecting the presence or absence of one or more diseases using quantitative approaches. Aspects of the present disclosure include methods for determining the abundance of endogenous targets. Aspects of the present disclosure also include determining the presence or absence of an aneuploidy.

The present invention provides a method of determining integrity and/or quantity of cell free DNA (cfDNA) in a biological sample comprising amplifying target sequences with at least a first primer/probe set and at least a second primer probe/set, amplifying the target sequences of differing lengths, and monitoring for detection of the labels of the oligonucleotide probes, and determining the integrity and/or quantity of the cfDNA based on the level of detection of the label of the oligonucleotide probe from the first primer/probe set compared to the level detection of the label of the oligonucleotide probe from the second primer/probe set. The present invention also provides methods for generating a library with the cfDNA for sequencing and analysis.