Compositions, methods, and kits for synthesizing, detecting, and/or quantifying nucleic acids are provided herein. Embodiments comprise a nucleic acid amplification composition comprising a thermostable DNA polymerase and agents which improve nucleic acid synthesis, amplification, detection, and/or quantification of nucleic acid targets in a crude extract or crude lysate sample.

Compositions, methods, and kits for synthesizing, detecting, and/or quantifying nucleic acids are provided herein. Embodiments comprise a nucleic acid amplification composition comprising a thermostable DNA polymerase and agents which improve nucleic acid synthesis, amplification, detection, and/or quantification of nucleic acid targets in a crude extract or crude lysate sample.

A chemically-enhanced primer is provided comprising a negatively charged moiety (NCM), an oligonucleotide sequence having a) non-nuclease resistant inter-nucleotide linkages or b) at least one nuclease resistance inter-nucleotide linkage. The chemically-enhanced primer can be used for sequencing and fragment analysis. Methods for synthesizing the chemically-enhanced primer as well as a method of preparing DNA for sequencing, a method of sequencing DNA, and kits containing the chemically-enhanced primer are also provided. The method of sequencing DNA can comprise contacting amplification reaction products with the composition wherein excess amplification primer is degraded by the nuclease and the chemically-enhanced primer is essentially non-degraded.

A chemically-enhanced primer is provided comprising a negatively charged moiety (NCM), an oligonucleotide sequence having a) non-nuclease resistant inter-nucleotide linkages or b) at least one nuclease resistance inter-nucleotide linkage. The chemically-enhanced primer can be used for sequencing and fragment analysis. Methods for synthesizing the chemically-enhanced primer as well as a method of preparing DNA for sequencing, a method of sequencing DNA, and kits containing the chemically-enhanced primer are also provided. The method of sequencing DNA can comprise contacting amplification reaction products with the composition wherein excess amplification primer is degraded by the nuclease and the chemically-enhanced primer is essentially non-degraded.

Methods, compositions and kits are described for resequencing, confirming or verifying Next Generation Sequencing (NGS) results with Sanger Sequencing. These methods are particularly useful for samples having very limited quantities such as formalin-fixed, paraffin-embedded (FFPE), Laser Capture Microdissection (LCM), fine needle biopsies or aspirates.

Methods, compositions and kits are described for resequencing, confirming or verifying Next Generation Sequencing (NGS) results with Sanger Sequencing. These methods are particularly useful for samples having very limited quantities such as formalin-fixed, paraffin-embedded (FFPE), Laser Capture Microdissection (LCM), fine needle biopsies or aspirates.

A nucleic acid isothermal amplification method is based on a polymerase spiral reaction using only one pair of primers. The method employs a self-spiraling amplification method, and has a high amplification efficiency.

A nucleic acid isothermal amplification method is based on a polymerase spiral reaction using only one pair of primers. The method employs a self-spiraling amplification method, and has a high amplification efficiency.

The present disclosure provides compositions and methods that employ the compositions for conducting pairwise sequencing and for generating concatemer template molecules for pairwise sequencing. The concatemers can be generated using a rolling circle amplification reaction which is conducted either on-support, or conducted in-solution and then distributed onto a support. The rolling circle amplification reaction generates concatemers containing tandem copies of a sequence of interest and at least one universal adaptor sequence. An increase in the number of tandem copies in a given concatemer increases the number of sites along the concatemer for hybridizing to multiple sequencing primers which serve as multiple initiation sites for polymerase-catalyzed sequencing reactions. When the sequencing reaction employs detectably labeled nucleotides and/or detectably labeled multivalent molecules (e.g., having nucleotide units), the signals emitted by the nucleotides or nucleotide units that participate in the parallel sequencing reactions along the concatemer yields an increased signal intensity for each concatemer.

The present disclosure provides compositions and methods that employ the compositions for conducting pairwise sequencing and for generating concatemer template molecules for pairwise sequencing. The concatemers can be generated using a rolling circle amplification reaction which is conducted either on-support, or conducted in-solution and then distributed onto a support. The rolling circle amplification reaction generates concatemers containing tandem copies of a sequence of interest and at least one universal adaptor sequence. An increase in the number of tandem copies in a given concatemer increases the number of sites along the concatemer for hybridizing to multiple sequencing primers which serve as multiple initiation sites for polymerase-catalyzed sequencing reactions. When the sequencing reaction employs detectably labeled nucleotides and/or detectably labeled multivalent molecules (e.g., having nucleotide units), the signals emitted by the nucleotides or nucleotide units that participate in the parallel sequencing reactions along the concatemer yields an increased signal intensity for each concatemer.