Whole Exome Sequencing Identifying Frequent Alternations In Genes In Ovarian Endometriosis

Endometriosis is a complex gynaecology disease, which is affected by multiplefactors including genetic, environmental and immune factors. Studies to date have notaddressed the actual causal genetic variants for this disease and finding candidate genesis a hot spot in recent years. The whole exome sequencing, as a newly high-throughputsequencing technology, is economic and effective. So far, the identifications ofcandidate genes in complex diseaseshave already made a breakthrough.However, therearestill no reports using whole-exome sequencing to search endometriosisat home andabroad.Thus, we conducted whole-exome sequencing to comprehensively search forsomatic mutations in both eutopic and ectopic endometriumand hope to find out therelated gene changes. This study can be divided into two parts.Part1Extracting and amplificatingwhole genomic DNA from endometrioticcells obtained by laser capture microdissectionObjective：To detect a standard protocol for extracting low amount DNA andWGA from the endometriotic cells obtained by laser capture microdissection.Methods:Using LCM to collect endometriotic cells from10paraffin specimens and10frozenspecimens which were used to extract DNA. Fiveother samples are prepared to dowhole genome amplification. The concentrations and OD values of final productionswere validated by spectrophotometer. And PCR was performed with housekeepinggenes in different chromosomes. Results: Both the frozen samples and formalin-fixedparaffin embedded tissue can extract micro DNA products. However, paraffinspecimens cannot remain intact DNA informationsafter doing whole genomeamplification. Conclusions: The simple and stable method of extracting andamplificating whole genomic DNA from endometriotic cells obtained by LCMcan beused as a routine approach for the future genome research of endometriosis.Part2Whole exome sequencing identifying frequent alternations in genes inoverian endometriosisObjective: Usewhole exome sequencing to search for somatic mutations in botheutopic and ectopic endometrium in endometriosis. Methods: Extracting DNA fromofectopic and eutopicglandular cells and blood from7endometriosis patients.Choose qualified specimen Illumina Hiseq2000to sequence the DNA samples. After gotprimitive data, we used database to call and filter useful informations.Results: Weidentifiedseven genes including MUC16, MGA, AHNAK, CENPE, HERC1, MAGEC1and PCLO which may be involved in the pathogenesis of endometriosis.Conclusion:1.Overall, the rates of alternations that we observed inendometriosis, whether in theeutopic or ectopic endometrium samples, were significantly higher than hadbeenpreviously determined rates for other tumors, which might be an natural instinct ofendometrium.2. The preexistingsomatic mutations in ovarian endometriosiswererandomly distributed and do not exsist “driver” mutations like tumors.3. Geneticalterations inadhesion and invasionsustained the theory of eutopicendometriumdeterminism.