Project description:In this study, small RNAs were isolated from individual donations of eight forensically relevant biological fluids (blood, semen, vaginal fluid, menstrual blood, saliva, urine, feces, and perspiration) and subjected to next generation sequencing using the Illumina® Hi-Seq platform. Sequencing reads were aligned and annotated against miRbase release 21, resulting in a list of miRNAs and their relative expression levels for each sample analyzed. Body fluids with high bacterial loads (vaginal fluid, saliva, and feces) yielded relatively low annotated miRNA counts, likely due to oversaturation of small RNAs from the endogenous bacteria. Both body-fluid specific and potential normalization miRNAs were identified for further analysis as potential body fluid identification tools for each body fluid.
Project description:MicroRNAs (miRNAs) in body fluids are candidate diagnostics for a variety of conditions and diseases, including breast cancer. One premise for using extracellular miRNAs to diagnose disease is the notion that the abundance of the miRNAs in body fluids reflects their abundance in the abnormal cells causing the disease. As a result, the search for such diagnostics in body fluids has focused on miRNAs that are abundant in the cells of origin. Here we report that released miRNAs do not necessarily reflect the abundance of miRNA in the cell of origin. We find that release of miRNAs from cells into blood, milk and ductal fluids is selective and that the selection of released miRNAs may correlate with malignancy. In particular, the bulk of miR-451 and miR-1246 produced by malignant mammary epithelial cells was released, but the majority of these miRNAs produced by non-malignant mammary epithelial cells was retained. Our findings suggest the existence of a cellular selection mechanism for miRNA release and indicate that the extracellular and cellular miRNA profiles differ. This selective release of miRNAs is an important consideration for the identification of circulating miRNAs as biomarkers of disease.
Project description:The ability to predict tissue type and donor’s age from molecular profiles of crime scene samples has practical implications in forensics. In order to identify body fluid- and age-associated DNA methylation changes, genome-wide DNA methylation profiling was carried out for body fluids including blood, saliva, semen, menstrual blood, and vaginal fluid obtained from individuals aged 20 to 59. The Illumina Infinium 450k Human DNA methylation Beadchip was used to obtain DNA methylation profiles across approximately 450,000 CpGs in bisulfite converted DNA. Samples included 12 of each blood, saliva and semen samples from 18 male donors aged 20 to 59, and 3 of each vaginal fluid and menstrual blood samples from 4 female donors in their twenties. Genome-wide DNA methylation profiling of body fluids obtained from individuals aged 29 to 41. The Illumina Infinium 450K Human DNA methylation Beadchip was used to obtain DNA methylation profiles across approximately 450K CpGs from human body fluids including blood, saliva and semen. Bisulfite converted DNA from the 6 samples were hybridised to the Illumina Infinium 450k Human Methylation Beadchip
Project description:The ability to predict tissue type and donor’s age from molecular profiles of crime scene samples has practical implications in forensics. In order to identify body fluid- and age-associated DNA methylation changes, genome-wide DNA methylation profiling was carried out for body fluids including blood, saliva, semen, menstrual blood, and vaginal fluid obtained from individuals aged 20 to 59. The Illumina Infinium 450k Human DNA methylation Beadchip was used to obtain DNA methylation profiles across approximately 450,000 CpGs in bisulfite converted DNA. Samples included 12 of each blood, saliva and semen samples from 18 male donors aged 20 to 59, and 3 of each vaginal fluid and menstrual blood samples from 4 female donors in their twenties. Genome-wide DNA methylation profiling of body fluids obtained from individuals aged 37 to 48. The Illumina Infinium 450K Human DNA methylation Beadchip was used to obtain DNA methylation profiles across approximately 450K CpGs from human body fluids including blood, saliva and semen. Bisulfite converted DNA from the 12 samples were hybridised to the Illumina Infinium 450k Human Methylation Beadchip
Project description:The ability to predict tissue type and donor’s age from molecular profiles of crime scene samples has practical implications in forensics. In order to identify body fluid- and age-associated DNA methylation changes, genome-wide DNA methylation profiling was carried out for body fluids including blood, saliva, semen, menstrual blood, and vaginal fluid obtained from individuals aged 20 to 59. The Illumina Infinium 450k Human DNA methylation Beadchip was used to obtain DNA methylation profiles across approximately 450,000 CpGs in bisulfite converted DNA. Samples included 12 of each blood, saliva and semen samples from 18 male donors aged 20 to 59, and 3 of each vaginal fluid and menstrual blood samples from 4 female donors in their twenties. Genome-wide DNA methylation profiling of body fluids obtained from young and old individuals. The Illumina Infinium 450K Human DNA methylation Beadchip was used to obtain DNA methylation profiles across approximately 450K CpGs from human body fluids including blood, saliva, semen, vaginal fluid and menstrual blood. Bisulfite converted DNA from the 24 samples were hybridised to the Illumina Infinium 450k Human Methylation Beadchip
Project description:In this study, small RNAs were isolated from individual donations of eight forensically relevant biological fluids (blood, semen, vaginal fluid, menstrual blood, saliva, urine, feces, and perspiration) and subjected to next generation sequencing using the Illumina® Hi-Seq platform. Sequencing reads were aligned and annotated against miRbase release 21, resulting in a list of miRNAs and their relative expression levels for each sample analyzed. Body fluids with high bacterial loads (vaginal fluid, saliva, and feces) yielded relatively low annotated miRNA counts, likely due to oversaturation of small RNAs from the endogenous bacteria. Both body-fluid specific and potential normalization miRNAs were identified for further analysis as potential body fluid identification tools for each body fluid. 32 samples - 3-5 replicates of each human biological fluid: venous blood, urine, semen (normal and vasectomized), vaginal secretions, menstrual secretions, perspiration, feces, saliva
Project description:Preterm birth (PTB) is defined as childbirth occurring at less than 37 completed weeks or 259 days of gestation. Premature babies have higher rates of cerebral palsy, sensory deficits, learning disabilities and respiratory illnesses that extend into adulthood. This lifelong morbidity results in high economic and social costs to families and communities. PTB is a syndrome initiated by multiple mechanisms, including infection or inflammation, uteroplacental ischaemia or haemorrhage, uterine overdistension, stress, and other immunologically mediated processes. Identifying and monitoring molecular signals in easily accessible body fluids that can diagnose or predict the risk of preterm labor in pregnant women will reduce or prevent PTBs. A number of studies reported the identification of putative biomarkers for PTB including protein, miRNA and hormone from different body fluids such as serum/plasma, cervical vaginal fluid, saliva and amniotic fluids. These putative biomarkers identified can largely be grouped into three main functional categories: inflammatory related molecules, placenta or fetal derived molecules and stress related molecules. In the past few years next generation sequencing (NGS) has become the major platform for miRNA analysis especially with body fluids. However, studies have shown significant sequence bias among different small RNA library preparation protocols. We have modified the small RNA library construction protocol which greatly reduces the sequence bias and increase miRNA coverage in sample. We also adapted a newly developed size exclusion chromatography (SEC) based EV purification protocol which can provide cleaner EVs compared to other methods. We are using these improved approaches to gain more reliable profile of circulating RNA in body fluid as well as its associated EVs. With these new approaches, we explore the possibility of using specific circulating miRNAs, specifically those encapsulated in EVs, as a noninvasive biomarker for PTB by comparing the miRNA profiles in maternal plasma, EV and EV-depleted plasma between individuals who had a spontaneous preterm birth and uncomplicated pregnancies.
Project description:Genome-wide expression profiling of four kinds of body fluid samples (blood, saliva, semen and vaginal swab). The purpose of the present study was selection of specific mRNA markers for identification of the four body fluids. Results provide important information about gene expression level of each body fluid for forensic science.