The DriverMap™ EXP Human Genome-Wide 19K (hGW19K) Profiling Kit for Dried Blood Microsamples (Version 3) measures the expression level of almost 19,000 human protein-coding genes by combining highly multiplexed RT-PCR amplification with the depth and precision of Next-Generation Sequencing (NGS) quantitation. The hGW19K Dried Blood Microsamples kit is designed to perform genome-wide expression profiling from 30 µl of dried human blood collected using Mitra® tips loaded with GentegraRNA-NEO stabilization reagent (Neoterix). Expression profiling of dried blood is a convenient strategy for longitudinal analysis of blood samples which can be collected directly by patients or study subjects. This method eliminates the need for blood donors to go to a blood collection facility and allows collection in remote locations that do not have the necessary infrastructure.
The development of the DriverMap™ EXP hGW19K Assay involved extensive optimization and experimental validation of tens of thousands of primer sets to identify a pool of primers that could be combined in a single multiplex RT-PCR reaction to amplify representative transcript sequences (i.e., conserved transcript regions common to different mRNA isoforms from the same gene) from each of ~19,000 protein-coding genes in the human genome. The amplified sequences are then quantified using NGS on the Illumina platform.
Main advantages of DriverMap™ EXP hGW19K Dried Blood Microsamples Assay:
- An easy-to-run, one-tube, single-day assay allows expression profiling directly from total RNA extracted from dried whole blood microsamples.
- The assay provides robust, quantitative, and reproducible measurements of the copy number of mRNAs over as many as 4-orders of magnitude differences in expression level.
- The assay provides increased sensitivity from a small amount of RNA (approx. 20 ng) isolated from whole blood dried microsamples using a set of highly experimentally validated primers.
- To achieve high specificity, the DriverMap protocol employs several mechanisms to minimize amplification of off-target and primer dimer background products.
- No mRNA enrichment, rRNA, mitochondrial, beta-globin depletion, or other processing is required.
- The primers are designed using Illumina’s dual UD Indexes labeling strategy to minimize NGS index-swapping background issues and amplified products are compatible with 100-150-cycle Paired-End NGS sequencing using Illumina’s NextSeq/NovaSeq instruments.
- NGS data can be analyzed using the Salmon software package available online. Cellecta also provides custom data analysis services on request.
Related Products and Services:
- DriverMap™ AIR TCR/BCR Profiling Kit (human/mouse RNA/DNA) is designed to generate comprehensive AIR CDR3 repertoire profiling of all seven TCR (TRA, TRB, TRD, and TRG) and/or BCR (IGH, IGK, and IGL) chains from RNA or DNA samples. AIR RNA assay provides higher sensitivity in detecting full-length (CDR1-CDR2-CDR3) or CDR3 clonotypes than AIR DNA assay, especially from small biological samples. Both AIR RNA and AIR DNA assay could be effectively used together to identify antigen-induced clonotypes. For more details, please refer to the AIR Technical Guide. Similar to the DriverMap EXP assay, AIR repertoire profiling can be also run using dried blood microsamples.
- DriverMap™ EXP Human Genome-wide 19K Profiling Kit allows profiling of almost 19,000 human protein-coding genes from total RNA isolated from cells, tissue, or blood, or directly from cellular extract using the DirectCell™ Protocol. It has a 104 – 105 fold dynamic range from 10-50 ng of total RNA and can be used effectively with as little as 1 ng of total RNA.
Cellecta also provides a DriverMap™ Expression Profiling Service. For more information, please visit our website:
DriverMap EXP Service Webpage
Please read the entire user manual before proceeding with your experiment.
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