Fixative optimisation study for BRITROC project

The British Translational Research Ovarian Cancer Collaborative (BriTROC), spanning more than 10 centres across UK, is a prospective observational project focused on relapsed high-grade serous ovarian cancer (HGSOC). The first study, as a part of the project, is the Fixative optimisation study, which analyses the utility for next-generation sequencing of UMFIX samples compared to NBF-fixed and fresh frozen samples. Various DNA and sequencing quality analyses were performed to compare fixation methods in bulk tumour and biopsy samples.

Type: Other
Archiver: European Genome-Phenome Archive (EGA)

2 Datasets 1 Publication

Click on a Dataset ID in the table below to learn more, and to find out who to contact about access to these data

Dataset ID	Description	Technology	Samples
EGAD00001001937	Targeted amplicon sequencing of samples as part of the study "Methanol-based fixation is superior to buffered formalin for next-generation sequencing of DNA from clinical cancer samples. The amplicon panel consists of 48 amplicons in TP53, PTEN, EGFR, PIK3CA, KRAS and BRAF genes as described previously [Forshew, STM 2012]. All libraries were pooled and quantify using DNA 1000 kit on Agilent 2100 Bioanalyzer and KAPA SYBR FAST ABI Prism qPCR Kit (KAPA Biosystems) on 7900HT Fast Real-Time PCR System (Applied Biosystems) according to the supplier's recommendations. Reads were aligned using bwa-mem v0.7.12-r1039 to the 1000 genomes version of human genome build GRCh37, retaining duplicate reads.	Illumina MiSeq	66
EGAD00001001938	Shallow whole-genome sequencing of samples from the study "Methanol-based fixation is superior to buffered formalin for next-generation sequencing of DNA from clinical cancer samples". DNA from each sample (100ng) was sheared on Covaris S220 (Covaris): duty cycle - 10%, intensity -5.0, bursts per sec - 200, duration - 300 sec, mode - frequency sweeping, power - 23V, temperature -5:5 C to 6 C, water level - 13. Libraries were prepared with the TruSeq Nano DNA LT Sample Prep Kit (Illumina) using a modi?ed protocol - Sample Puri?cation Beads were replaced by Agencourt AMPure XP beads (Beckman Coultier) and size selection after the End Repair was done to remove only the short fragments. Quality and quantity for contructed libraries were assessed with DNA 7500 kit on Agilent 2100 Bioanalyzer and with Kapa Quanti?cation kit (KAPA Biosystems) on 7900HT Fast Real-Time PCR System (Applied Biosystems) according to the supplier's recommendations, respectively. Libraries from 18 barcoded samples were pooled together in equimolar amounts and each pool was loaded on a single lane of a HiSeq Single End Flowcell (Illumina), followed by cluster generation on a cBot (Illumina) and sequencing on a HiSeq 2500 (Illumina) in a single-read 50bp mode. Reads were aligned using bwa-mem v0.7.12-r1039 to the 1000 genomes version of human genome build GRCh37. Picard (http://picard.sourceforge.net) was used to remove duplicate reads.	Illumina HiSeq 2500	60

Publications	Citations
Methanol-based fixation is superior to buffered formalin for next-generation sequencing of DNA from clinical cancer samples. Piskorz AM, Ennis D, Macintyre G, Goranova TE, Eldridge M, Segui-Gracia N, Valganon M, Hoyle A, Orange C, Moore L, Jimenez-Linan M, Millan D, McNeish IA, Brenton JD. Ann Oncol 27: 2016 532-539	21