High-throughput Analysis in Nanoscale Reactions
The BioMark HD and Juno enable high-throughput and sample-efficient analyses in low volume reactions. The instruments support several genomic-based applications, such as gene expression analysis, genotyping by allele-specific PCR, as well as NGS Library preparations for Illumina sequncers.
Fluidigm has developed chips with integrated fluidic circuits (IFCs), also referred to as Dynamic Arrays – pictured below. The IFC consists of two layers of microchannels. Pressurized liquids flowing in one layer is used to precisely control the microchannels below, creating hundreds of thousands of valves, channels, and chambers. The technology allows rapid and reproducible multistep sample preparation, and analysis of up to hundreds of genetic markers across thousands of samples is completed in just hours, rather than days or weeks.
The principle of the technology is based on assays being loaded on one side of the chip, and samples being loaded on the other. Once in the wells, the components are pressurized into the chip using an IFC controller. The components are then systematically combined, so that in each chamber of the chip one sample is mixed with one assay and the reaction takes place there. In this way thousands of reactions are run simultaneously.
At BioXpedia we combine the Fluidigm instruments BioMark HD and Juno, which enables maximal efficiency and workflow. We employ the Juno instrument as a controller, preparing the IFC chips, which are then run on the Biomark HD in either real-time or endpoint read mode.
The BioMark HD qPCR system supports several genomics-based applications, such as gene expression profiling (both mRNA and miRNA), SNP genotyping, digital PCR, mutant detection, and many more. In addition, we use the BioMark HD for quantification of protein in combination with Olink Protein screening.
The Juno also work as a stand-alone instrument, by providing library preparation for next-generation sequencing (NGS) on Illumina sequencers in a fully automated workflow.
Integrated Fluidic Circuit for Efficient and Reproducible Analysis
The Dynamic Array IFC chips come in different variants allowing flexibility regarding number of samples and assays.
Chip variants:
- 12.12 – 12 samples x 12 assays (144 reactions)
- 48.48 – 48 samples x 48 assays (2,304 reactions)
- 96.96 – 96 samples x 96 assays (9,216 reactions)
- 192.24 – 192 samples x 24 assays (4,608 reactions)
- 24.192 – 24 samples x 192 assays (4,608 reactions)
Loading the BioMark HD with a Dynamic Array, Integrated Fluidic Circuit (IFC)
SNP Genotyping
At BioXpedia we provide SNP genotyping service, using the Fluidigm systems allowing high-throughput and maximum reproducibility.
Genotyping can be key in understanding disease or to predict treatment response of a specific drug. Several technologies, like Next Generation Sequencing (NGS), microarrays or targeted SNP genotyping on the Fluidigm platform, can be used in genotyping studies, depending on the aim of the study.
Genotyping allows researchers to study genetic variations across the genome between different groups of patients. These variations can be both small differences, such as single nucleotide polymorphisms (SNPs) or larger structural DNA changes, e.g. loss of heterozygosity and copy number variation (CNV).
For animal or plant breeders, SNP genotyping is a widely used tool to identify specific traits in a population, and useful for breeding selection.
The BioMark HD system for SNP genotyping is a fast and reproducible method as multiple samples can be evaluated in a single run, and the workflow is almost entirely automated requiring minimal hands-on time.
Genotyping using BioMark HD requires only small amounts of DNA, making it ideal for analysis of precious samples, and the system is compatible with multiple types of assays, including:
- SNP Type assays,
- TaqMan genotyping assays – See available assays here
- IDT rhAmp® SNP assays
TaqMan assays include allele-specific probes, while SNP Type and IDT rhAmp SNP assays are based on universal probes, and all assay types allow customized assay design.
To learn more about the Fluidigm Technology click here
Gene Expression (qPCR) – mRNA and miRNA
BioXpedia offers Gene Expression analysis by qPCR using the Fluidigm system, which provides flexibility and high-throughput.
Transcription of genes is a dynamic process, allowing cells to adapt rapidly to external, environmental, or physiological changes. mRNA profiling is the first level of exploration for any regulation at the molecular and cellular level, and is essential in numerous research areas, such as oncology, immunology, or neurology. The approach can be both targeted at specific genes or more explorative in the sense of discovering multiple differences across the transcriptomic level – e.g. between different groups of patients.
In addition, miRNAs have been found to be important regulators of several processes in the human body, where they function by translational silencing or destabilization of mRNA. miRNAs are typically located and exert their mechanism within the cells, however circulating miRNAs are also found in the extracellular milieu (e.g. serum, plasma etc.). Here they may be important factors in cell-to-cell communication, and as potential biomarkers for some solid tumors.
By employing the Fluidigm systems, BioXpedia provides a rapid, flexible, and high throughput set up for multiplex gene expression assays. Assays can either be based on dual-probes (TaqMan) or DNA-binding dye (EvaGreen). The expression of selected genes, ranging from 12 to 192 genes, can be profiled in a single run. The system requires very little RNA as input and creates highly reproducible data.
To learn more about the Fluidigm Technology click here
Library Preparation and RNAseq
BioXpedia offers library preparation for NGS. It can be utilized as a stand-alone service or in combination with RNAseq on Illumina sequencers.
Transcriptome profiling by RNAseq enables differential gene expression, SNP identification, alternative splicing, or novel transcript identification. It can be used for comparison against a reference genome or de novo assembly.
The Juno system from Fluidigm provides a fully automated workflow from sample loading to harvest of barcoded libraries. On the IFC, a solid phase capture of poly(A)RNA selects full length mRNA and some lncRNA. After elution, the RNA is fragmented, and used for first-strand cDNA synthesis by reverse transcription using random primers. The second strand cDNA is generated from the first strand by utilizing a template switching primer. In this way the orientation of the original RNA is kept, generating a stranded library. Barcodes and adaptors are added via PCR, and the samples are harvested from the Juno. Finally, the samples are pooled, cleaned, and enriched. After QC the libraries are ready for NGS.
Each IFC can make 48 stranded libraries in a run and requires only 10-100 ng total RNA as input material per library. By the use of dual indexing barcoding, up to 96 libraries can be sequenced together on an Illumina Sequencer, making the system both cost- and sample efficient.
BioXpedia offers everything in the process of RNAseq starting from RNA extraction and library preparation to RNAseq and downstream demultiplexing and bioinformatics.
To learn more about the Fluidigm Technology click here