Lynch Syndrome / Hereditary Non-Polyposis Colorectal Cancer (HNPCC)


Related Gene(s): MLH1, MSH2, MSH6, PMS2, EPCAM

Lynch syndrome, previously known as hereditary non-polyposis colorectal cancer (HNPCC), is caused by mutations in the mismatch repair (MMR) genes MLH1, MSH2, MSH6, PMS2, and EPCAM. Lynch syndrome is the most common hereditary form of colorectal cancer. It affects about 1 in 440 individuals in the United States.

Testing of Lynch Syndrome genes can be ordered sequentially or concurrently. Analysis for the MSH2 inversion of exons 1-7 can be ordered as a stand-alone test, but this inversion is automatically included in all tests with MSH2 sequencing.

MLH1 coding exons 1-19, MSH2 coding exons 1-16, MSH6 coding exons 1-10, and PMS2 coding exons 1-15, and well into the 5’ and 3’ ends of all the introns and untranslated regions are analyzed by sequencing. Gross deletion/duplication analysis determines gene copy number for all coding exons of MLH1, MSH2, MSH6, PMS2, and coding exon 9 of EPCAM. The inversion of coding exons 1-7 of the MSH2 gene is detected by NGS and confirmed by PCR and agarose gel electrophoresis. Clinically significant intronic findings beyond 5 base pairs are always reported. Intronic variants of unknown or unlikely clinical significance are not reported beyond 5 base pairs from the splice junction. Genomic deoxyribonucleic acid (gDNA) is isolated from the patient’s specimen using standardized methodology and quantified. Sequence enrichment of the targeted coding exons and adjacent intronic nucleotides is carried out by a bait-capture methodology, using long biotinylated oligonucleotide probes followed by polymerase chain reaction (PCR) and next generation sequencing (NGS). Sanger sequencing is performed for any regions missing or with insufficient read depth coverage for reliable heterozygous variant detection. Reportable small insertions and deletions, potentially homozygous variants, variants in regions complicated by pseudogene interference, and single nucleotide variant calls not satisfying 100x depth of coverage and 40% het ratio thresholds are verified by Sanger sequencing.

Gross deletion/duplication analysis of MLH1, MSH2, MSH6, and PMS2 using read-depth from NGS data and EPCAM using multiplex ligation-dependent probe amplification (MLPA) is also performed. Any copy number changes detected by NGS are confirmed by targeted chromosomal microarray and/or MLPA. If a deletion is detected in exons 13, 14, or 15 of PMS2, double stranded sequencing of the appropriate exon(s) of the pseudogene, PMS2CL, will be performed to determine if the deletion is located in the PMS2 gene or pseudogene. If a deletion is detected in exon 9 of EPCAM, deletion/duplication analysis of coding exons 3 and 8 of EPCAM will be performed. For EPCAM, only gross deletions encompassing the 3’ end of the gene are reported.

Single gene testing and genetic testing for a known familial mutation can also be ordered. If there is a previously-identified mutation in a family member, please include a copy of the test report with the requisition form.

Specimen Requirements

  • Whole blood: Two 4.5 mL EDTA tubes (lavender top)
  • For transfusion patients, please wait at least 2 weeks after a packed cell/platelet transfusion, and at least 4 weeks after a whole blood transfusion prior to blood draw for testing
  • For chemotherapy patients, the DNA quality may be affected if patient has received chemotherapy within the last 120 days. Sema4 may request an additional specimen if DNA quality is insufficient


  • Saliva: 2 mL of freshly-collected saliva in an Oragene container per kit’s specific instructions
  • Fill up to black line with 2 mL of saliva and close the lid. Once the lid is closed, it automatically adds 2 mL of buffer, for a total volume of 4 mL
  • Please note that 2 containers are required for all pediatric saliva testing kits


Saliva and blood are the most common specimen types we receive. For questions regarding other specimen types and requirements for patients with a significant medical history (including allogenic transplant and hematological diseases), please call us at 203-483-3459 to discuss before sample submission.

Ordering Information


  • Ship at room temperature
  • Ship to:
    1 Commercial Street, Branford, CT 06405
    Phone: 203-483-3400 / Fax: 203-483-3443



Turnaround Time

  • 14-21 days from receipt of specimen



Hereditary Cancer Requisition
Hereditary Cancer Genetic Testing Consent