Next Generation Sequencing

Sirona Dx offers Next Generation Sequencing (NGS), using the Ion Proton  and Ion PGM systems. Our scientists leverage the powerful Ion Torrent TM  sequencing technology to generate large amounts of high quality  sequencing data including from challenging clinical samples such as  FFPE.

Ion Torrent Next Generation Sequencing (NGS) utilizes a  semiconductor-based, sequencing-by-synthesis technology and emulsion PCR  (emPCR) similar to other platforms. It measures the H+ ions released  during base incorporation.

Bases of small fragments of DNA are sequentially identified from  signals emitted as each fragment is re-synthesized from a DNA template  strand. NGS extends this process across millions of reactions in a  massively parallel fashion, rather than being limited to a single or a  few DNA fragments. This advance enables rapid sequencing, or large  stretches of DNA base pairs spanning entire genomes, with the latest  instruments capable of producing many gigabases of data in a single  sequencing run.

For example, consider a single genomic DNA (gDNA) sample. The gDNA is  first fragmented into a library of small segments that can be uniformly  and accurately sequenced during millions of parallel reactions. The  newly identified strings of bases, called reads, are then reassembled  using a known reference genome as a scaffold (resequencing) or in the  absence of a reference genome (de novo sequencing).


Applications for NGS include:

  • Targeted sequencing for DNA and RNA
  • Exome sequencing, enabling identification of single nucleotide variants (SNVs) and small insertions and deletions (indels)
  • Transcriptome sequencing (RNA sequencing can quantitate changes in  gene expression, as well as detect fusion transcripts from chromosomal  rearrangements)
  • Genome sequencing, including de novo sequencing, bacterial typing, and viral typing
  • Aneuploidy detection (Large chromosomal duplications or deletions and copy number variations (CNVs))
  • Epigenetic Applications (Chromatin Immunoprecipitation sequencing).
  • Microbial Population Analysis, including metagenomics and viral, bacterial, and yeast genome sequencing


Targeted Sequencing

Sirona Dx is an Ion Torrent Certified Service Provider,  offering an extensive suite of Next Generation Sequencing capabilities  with the Ion PGM and Proton systems.Targeted sequencing enables the interrogation of specific genomic regions in a high throughput and cost-effective manner.There are a number of applications for targeted sequencing, including  the detection of various genetic aberrations such as single-nucleotide  polymorphisms (SNP's), insertions and deletions (indels), copy number  variants (CNV's), and genetic rearrangements. Some applications include  cancer genomics, cancer mutation analysis, and genetic disease/disorder  research - including analysis of FFPE (formalin-fixed paraffin-embedded)  tissue samples.Ion AmpliSeq ready-to-use targeted primer panels, or custom-designed  panels, contain primer pairs that are specifically designed to flank  regions of interest for polymerase chain reaction (PCR) amplification  with as low as 10 ng of input DNA.

Targeted Sequencing Panels Offered by Sirona Dx Genomics Team

  • The most comprehensive cancer panel available, the Ion AmpliSeq  Comprehensive Cancer Panel provides complete coverage of over 400 of the  genes most frequently implicated in cancer including KRAS, EGFR, FLT3,  PTEN, TP53 and many others.
  • For a more focused and cost effective approach to identify cancer  variants the Ion Ampliseq Cancer Hotspot Panel v2 targets approximately  2,800 COSMIC mutations from 50 oncogenes and tumor suppressor genes.
  • The Ion Ampliseq Inherited Disease Panel provides coverage of the  coding regions of 300 genes that are implicated in over 700 inherited  Mendelian diseases.
  • Custom Ampliseq Panels can be designed to target specific genomic regions of interest.


Exome Sequencing

Sirona Dx CLIA Laboratory is a Life Technologies certified service  provider for Ion Ampliseq Exome Sequencing. This certification indicates  that the sequencing data generated by our laboratory meets a high  quality standard. Using the Ion Proton platform and its straightforward  technical approach also allows for a rapid turnaround time.
Recent advances in next generation sequencing have enabled scientists  to explore massive quanities of genomic data. The exome represents only  one percent of the total human genome, however the majority of  disease-causing mutations are found within this region.

The Sirona Dx services for the Ion Ampliseq Exome include:

  • Coverage of 58Mb including 33Mb of exonic DNA representing >97% of known coding regions;
  • Suitable for samples with low DNA concentrations, requiring only 50ng input DNA;
  • Basic data analysis includes production of aligned reads and a list of annotated variants;
  • Custom bioinformatics analysis available to address additional needs.

Potential applications for exome sequencing:

Exome sequencing has enhanced our understanding of the role  of genetics in disease leading to a more personalized approach to  medicine.Key areas where exome sequencing is transforming clinical research include:

  • Enhancing knowledge of the genomic mutations involved in the progression of various types of cancers;
  • Understanding why individuals have different responses to  pharmaceuticals and predicting personalized treatment regimens that will  most effectively meet the needs of patients;
  • Investigating the genetic basis of complex diseases including  Alzheimer's, autism, multiple sclerosis, diabetes, and other polygenic  disorders;
  • Diagnosis of rare genetic disorders


SNP Genotyping

Our Laboratory offers SNP Genotyping (single-nucleotide polymorphism)  services using the Quant Studio Real-Time PCR instrumentation. This  platform not only detects the amount of fluorescence through each cycle  of polymerase chain reactions, it confidently distinguishes between more  ambiguous genotypes. The instrument provides high sensitivity with  lower DNA input, accurate analysis of the results, and faster turnaround  time.
Our  team uses Taqman assays to perform SNP genotyping. These asays  are mixtures of allele-specific primers and minor groove binding probes  that identify polymorphisms by generating a fluorscent signal to  indicate the presence of either or both targeted alleles.SNP genotyping applications include;

  • Providing insight into the molecular basis of disease,
  • Pharmacogenomics,
  • The ability to link genetic variations with phenotypic changes.

Performance specifications include:

  • Both 96-well and 384-well plate options
  • Simple workflow
  • Low reaction volumes require little input DNA
  • Shorter run times allow for higher throughput of samples