Multiplicom's MASTR Technology - A powerful multiplex PCR based approach enabling MPS based diagnostics


Abstract: While standard PCR technology is limited to amplifying only one or a few target DNA sequences in a single reaction, Multiplicom’s approach allows the amplification of up to 200 target sequences per reaction using standard thermal cyclers and laboratory equipment. Multiplicom’s assay design platform allows robust, rapid, and cost effective development of novel MASTR assays for diagnostic massively parallel sequencing. The resulting MASTR assays substantially reduce processing cost and front-end workload to the end-user as proven by multiple European clinical diagnostic labs. Using BRCA MASTR assays, those labs were able to reduce their turnaround time from ~ 6 months to less than 6 weeks without any sacrifice of data quality. This is due to the fact that (i) only 5 robust PCR reactions are required to amplify all of the coding exons of BRCA1/2 compared to about 100 reactions in a simplex PCR approach and (ii) multiple DNA samples can be processed in parallel (16 DNA samples can be processed per 96-well plate). Furthermore, total processing cost is reduced two to five fold compared to Sanger based sequencing as a result of (i) highly specific amplification (> 96% of resulting MPS reads map back to the target sequences) and (ii) more than 99% of the amplicons have read counts in excess of 0.2 times the mean. This means that at an average read count of 100, more than 99% of the amplicons will have read counts in excess of 20. The high specificity and the narrow amplification range enable efficient use of MPS resources and provide a significant cost reduction over other MPS based resequencing technologies. Indeed, competing technologies (i.e. TruSeq, AmpliSeq and Haloplex) typically show 75% of MPS reads that fall within a 0.2 times the mean number of read counts and only about 80% of reads can be mapped back to their corresponding target sequences. Multiplicom currently has six MASTR assays on the market: predisposition for breast & ovarian cancer (BRCA1 and BCRA2; IVD-CE label pending); colon cancer: FAP (APC and MUTYH) and HNPCC (MLH1, MSH2, MSH6, PMS2, 3’ UTR of EPCAM); Cystic Fibrosis (CFTR); Marfan Syndrome (FBN1) and Alport syndrome (Col4A3, Col4A4 and Col4A5). Several other assays are under development including MASTR assays for DMD, cardiovascular diseases, renal diseases, targeted cancer therapy (on FFPE derived DNA) and non-invasive trisomy testing. Multiplicom also offers a rapid, cost-effective custom assay development program.

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