Heat-Seq Ultra Oncology HotSpot Panel

Discontinued HEAT-Seq Portfolio

The HEAT-Seq Ultra Oncology HotSpot Panel has been discontinued. Our new KAPA Target Enrichment Portfolio is now available and contains our new KAPA HyperExome, as well as our new custom probes, KAPA HyperChoice and KAPA HyperExplore. For additional questions, please contact your regional sales representative.

Overview

The HEAT-Seq® Ultra Oncology HotSpot Panel is based on amplification-based enrichment employing an advanced version of Molecular Inversion Probe (MIP) technology and combines a fast and easy workflow with powerful SNV detection. The panel contains 635 probes, and has a capture target of approximately 30.5 Kb across previously described mutation hotspots in 53 commonly targeted oncogenes and is ideal for detecting and characterizing low-frequency mutations. The panel helps with the detection of variants at 1% minor allele frequency (MAF) and below.

Benefits of Heat-Seq Oncology HotSpot Panel

Smaller overall design size with simplified analysis and high throughput
Detection and removal of PCR duplicates using molecular barcoding and the ability to confidently discern a biological replicate and a process artifact
Maximized variant detection over a wide range of allelic frequencies using molecular barcoding with UIDs without the need for deep sequencing
Simple workflow with sequencing-ready DNA in a single tube with just two hours of hands-on time
Minimized sequencing errors due to the stringent removal of false positives
Utilize HSQutils software, which is a separately sourced sequencing read mapper and variant caller for use in the analysis workflow

Heat-Seq Workflow

The final product of the workflow is a sequencing-ready amplicon that contains the target DNA sequence, a molecular barcode, and sample indices.

The entire workflow is completed in 8 hours with < 2 hours of hands-on time using a single tube. The protocol is easy to adopt and is also automation-friendly.

Product Highlights

Target the mutation hotspots in oncogenes with one panel

HEAT-Seq Ultra Oncology Panel genes
UABL1	EZH2	KDR	PDGFRA
AKT1	FBXW7	KIT	PIK3CA
ALK	FGFR1	KRAS	PTEN
APC	FGFR2	MAP2K1	PTPN11
ATM	FGFR3	MAP2k4	RB1
BRAF	FLT3	MET	RET
BRCA1	GNA11	MLH1	RUNX1
BRCA2	GNAQ	MPL	SMAD4
CDH1	GNAS	MSH2	SMARCB1
CDKN2A	HNF1A	MSH6	SMO
CSF1R	HRAS	NF1	SRC
CTNNB1	IDH1	NF2	STK11
EGFR	IDH2	NOTCH1	TERT
ERBB2	JAK2	NPM1	TP53
ERBB4	JAK3	NRAS	VHL

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Uniform and complete coverage of the capture target

HEAT-Seq Ultra Oncology HotSpot Panel performance

The hotspot panel was tested using 100 ng of FFPE DNA. 24 samples were multiplexed on a single MiSeq lane using v3 chemistry. Data from three replicate experiments is shown. High on-target coverage, uniformity, and coverage rates were observed for each replicate.

View performance data of the HEAT-Seq Ultra Oncology Panel showing uniform and complete coverage of capture targets.

Accurate and sensitive detection of allele frequency

The HEAT-Seq Ultra Oncology HotSpot Panel was tested using an FFPE control sample with known mutations at frequencies between 0.9% and 25%. Observed allele frequencies strongly matched the known frequencies, showing the sensitivity and accuracy of this panel..

View performance data showing accurate and sensitive detection of allele frequency using the HEAT-Seq Ultra Oncology Panel.

High coverage depth over regions of interest

Coverage depth over regions of interest.

The HEAT-Seq Ultra Oncology HotSpot panel was tested using the Formalin Compromised Quantitative Multiplex Reference Standard (HD-C749, Horizon Diagnostics). Using all available reads (3-5 M per sample), the panel provided coverage of between 250x and 1250x of known mutations.

View data showing high coverage depth (between 250x and 1250x) of known mutants using the HEAT-Seq Ultra Oncology Panel.

Sensitive detection of false positives

False-positive rate in detection of variants using various methods.

The performance of the HEAT-Seq Ultra Oncology HotSpot Panel is compared to similar kits to assess allele frequency detection accuracy and data quality. The HEAT-Seq panel presents a similar or improved variant detection ability, but also presents an extremely low false-positive rate. The proprietary process of removing duplicates using molecular barcoding is the core of the HEAT-Seq system that enables not only accurate variant calls but also the confident removal of process induced errors that appear as false positives.

	HEAT-Seq Ultra Oncology HotSpot Panel	Illumina TruSeq Amplicon Cancer Panel	Thermo Fisher Ion AmpliSeq Cancer Panel v2
Average False-Positive Rate	0.014%	0.213%	3.071%
Range	0.0% - 0.2%	0.0% - 0.4%	0.0 - 11.1%
Missed Calls	0	2	0

Design Files

The design and annotation files provide information about genomic regions covered by the capture probes and the genes included in these regions.

These files were designed for use with the following Roche products:

HEAT-Seq Ultra Oncology Panel, 24 Reactions (Catalog No. 07757069001)
HEAT-Seq Ultra Oncology Panel, 96 Reactions (Catalog No. 07768613001)

Download Design File

Demo Data

Demo data is available for evaluation. Please contact Roche representative for your country.

Research Use Only. Not for use in diagnostic procedures.
HEAT-SEQ is a trademark of Roche.