Development And Application Of Laser Capture Microdissection In Cast-off Cells For Forensic Genetics

ObjectivesThe purpose of this project is to establish a set of suitable for cast-off cells detection LCM technology, and the method in cast-off cells of type specimens for the degradation of cast-off cells in particular, an exploratory study.Method1Cast-off cells in oral epithelium as a model, from the staining method, extraction method, and cycling parameters of other aspects were compared, the establishment of LCM method.2The mixture of acidic drinks and oral epithelial cells to study Chelex-100method, Chelex-100method with the Microcon100combined, DNA IQTM extraction method, a comparative with the LCM method.3To urinary cast-off cells for the study, the conventional method were compared with the LCM method.Results1Established by this study, LCM method captures10oral epithelial cells, you can get the complete DNA typing, reproducible, practical case for forensic biological evidence the DNA testing of biological samples provides a reference method.2Acidic mixture of drinks and oral epithelial cells using Chelex-100method only or combined with Microcon100, it is difficult to obtain high quality DNA, making it difficult to obtain satisfactory results; for the class of mixtures of citric acid, direct extraction method using DNA IQIM extraction, and can reduce the elution volume, increased DNA concentration, thereby improving the detection rate of DNA; lactic acid protein-rich mixture may consider the LCM method, the target cells isolated from the complex environment, more conducive to obtaining more good test results.3Fresh urine group:satisfactory results can be obtained in all loci in DNA extracted by LCM technology. And in DNA extracted by Chelex-100, locus allele loss, low peak, non-specific expansion and other issues happen; stored urine group:for urine samples stored at4℃for10days and at room temperature for4days, by LCM technology combined with DNA IQTM extraction method, more than12loci can be clearly interpreted; and the valid typing can not be obtained by Chelex-100. ConclusionLaser Capture Microdissection provides an effective method in detection of cast-off cells, particularly of cast-off cells with degradation, and also provides an idea for detection of other types of trace biological samples.