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Ampli1™ workflow: Single Cell DNA analysis

Successful analysis of single cell genome starts with an appropriate Whole Genome Amplification step and ends with an optimized sequencing process which matches the characteristics of the adopted WGA.

Ampli1™ provides a complete range of products covering the entire workflow and optimized for different kinds of genetic analysis on the amplified DNA. The complete validated workflows - which inludes Ampli1™ WGA, Ampli1™ QC Kit, Ampli1™ ReAmp/ds Kit and Ampli1™ LowPass Kit - will drive you through the processing of single cell DNA for Copy Number Aberration analysis by next-generation sequencing (NGS).

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Ampli1™ WGA Kit

Balanced, robust whole genome amplification from single cells

image-5The Ampli1™ Whole Genome Amplification (WGA) kit has been developed and optimized for balanced and complete amplification of the total DNA content of a single cell. The output of the Ampli1WGA protocol is a well-balanced library of approximately 19 million fragments representing the entire genome.  This DNA library is ideally suited for downstream analysis of both SNPs and CNV in single cells, due to its low rate of preferential amplification.

 

Single tube protocol ideal for all sample types

The Ampli1™ WGA Kit is designed to provide optimal whole genome amplification—not only from samples containing DNA from a few cells, but from DNA obtained from one single cell. Input DNA may be derived from fresh, fixed, and/or stained cells.

Following cell lysis, DNA is digested with a restriction enzyme and adaptors are ligated onto the DNA fragments. Amplification, performed in the same tube, is mediated by a single highly specific PCR primer for all 19 million fragments.


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Reliable, Reproducible, Balanced Single Cell Amplification

Key features of the Ampli1™ WGA Kit:
  • Single-tube, no-precipitation protocol minimizes template loss
  • Single, highly specific primer-mediated PCR ensures balanced amplification of both alleles
  • Robust protocol generally produces about 2 micrograms dsDNA and 5 micrograms of ssDNA.
Amplified DNA may be used in a variety of downstream applications, including:
  • Sanger Sequencing
  • Next-generation sequencing (NGS)
  • Microsatellite analysis
  • Copy number variation (CNV) analysis by RT-PCR or aCGH

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Ampli1 QC Kit Scatola_Ampli1 QC Kit_newlogo.jpg

The Ampli1™ QC Kit provides a useful quality control check of the WGA procedure by utilizing a PCR-based assay to establish DNA integrity.

By testing for the presence of one short and three long DNA fragments in the WGA library, the quality of the DNA as starting material for downstream analyses can be assured.

Cells with good quality DNA typically produce 3 or 4 PCR bands, while fixation of cells or cells with degraded DNA will show fewer QC bands. It has been proved that the number of the bands revealed is useful to define a “Genome Integrity Index” (GII) for predicting success of various downstream analytical methods and for assessing the quality of the starting DNA.

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 Ampli1™ LowPass Kit

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Ampli1™ LowPass Kit is designed to generate multiplexed, sequencing-ready libraries in a streamlined single-day protocol, to detect chromosomal aneuploidies and copy number aberrations (CNAs) by low-pass whole genome sequencing.

Ampli1™ LowPass Kit is compatible with Ion Torrent systems (Ion PGM™ and Ion S5™) and NOW also compatible with Illumina® systems (MiSeq®, HiSeq® 1000/1500, 2000/2500, 3000/4000 and NovaSeq™).

Learn more by visiting the dedicated page Ampli1™ LowPass Kit.

 

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 Ampli1™ ReAmp/ds Kit 

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Reamplify the Ampli1™ WGA product obtained from single cells, or from a batch of cells, in a reliable and reproducible way.

Reamplification will provide additional WGA-amplified DNA from the same cell. For downstream genetic analysis, you may use the Ampli1™ Reamplification product in the same manner as Ampli1™ WGA product.

 

Ampli1™ ReAmp/ds Kit can also be used to convert ssDNA molecules into dsDNA molecules both in Ampli1™ WGA products and in Ampli1™ reamplification products in order to increase the total dsDNA content. This procedure is recommended for accurate DNA quantitation, high-throughput high dimensional downstream analysis, and NGS applications to ensure high sequencing uniformity.

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Menarini Silicon Biosystems has developed Sanger sequencing assays allowing analysis of commonly-examined target genes starting from Ampli1™ WGA products.

  • PIK3CA kit— includes reagents for evaluation of the most common mutations in exons 9 and 20 of the PIK3CA gene.
  • EGFR kit—includes reagents for evaluation of the most common mutations in exons 18, 19, 20, and 21 of the EGFR gene.
  • KRAS kit—includes reagents for evaluation of somatic mutations in codons 12 and 13 of exon 2 in the KRAS gene.
  • BRAF kit—includes reagents for evaluation of V600 mutations of the BRAF gene.

If your research includes analysis of other genes, please contact us for information on obtaining custom primers compatible with the Ampli1™ WGA generated DNA libraries.

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To download Material Safety Data Sheets (MSDS), please click here.

The Ampli1™ WGA Kit is for Research Use Only. Not for use in diagnostic procedures. Ampli1™ WGA Kits are protected by U.S. patent 6,673,541 and corresponding patent claims outside the United States.

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