Abstract: A retrotransposable element based multiplexed qPCR assay to robustly quantitate and distinguish cell free DNA integrity and concentration in blood plasma and serum is described. The multiplexed system for characterizing cancer in humans includes a sample of serum, plasma, urine, or other biological fluid, the sample comprising cell free DNA, the cell free DNA comprising long and short retrotransposable element targets and an added internal positive control, the long and short targets being independent of each other, a distinctly labeled TaqMan probe corresponding to each target, a forward primer and a reverse primer corresponding to each target, a DNA standard for generating standard curves, a qPCR system for amplifying the targets and a qPCR data analysis system.

Abstract: By utilizing a Mini-Primer strategy targeting the target site duplication (TSD) sequence of retrotransposons, INNUL markers, which include SINEs, LINEs, and SVAs, can be effectively used as markers for human identification and bio-ancestry studies regardless of the size of the inserted element. The size of the amplicons for INNULs and the difference between allelic states can be reduced substantially such that these markers have utility for analyzing high and low quality human DNA samples. A 15 RE marker and Amelogenin (for sex determination) multiplex for a single tube amplification of DNA, in four color detection was successfully designed. The multiplex provided power of discrimination suitable for forensic and paternity analysis. In addition, sensitivity of detection can enable human identity and bio-ancestry studies on forensic and anthropological samples. Depending on the distribution of the alleles in global populations, INNULs can be selected for human identity testing or for bio-ancestry studies.

Abstract: A process of quantifying the extent of degradation present in a human DNA sample is described. The process makes use of a real time PCR system to separately quantitate within a sample a first retrotransposon interspersed element and a relatively longer second retrotransposon interspersed element, where the longer element is expected to be disrupted at a faster pace than is the shorter element as the sample degrades. In one embodiment, the process makes use of the appearance of the relatively young (on an evolutionary scale) Alu Yb-lineage subfamily sequences appearing in every human genome and their virtual absence in non-human samples. In a preferred embodiment, the process quantifies longer 290 bp sequences of “SVA” elements and shorter 80 bp sequences of Alu Yb8-lineage. Newly designed primers and TaqMan probes that are useful in the process are presented. A related process additionally quantifies male specific human DNA.

Abstract: A family of PCR assays is disclosed for determining, both qualitatively and quantitatively, presence of material from a predetermined species source and for quantifying the amount of such material. The assays are based respectively on SINEs uniquely characteristic of pig species, cow species, chicken species, and ruminant sub-order, and having a high copy number. The assays disclosed permit rapid, inexpensive evaluation of meat samples to facilitate elimination from their diet of pork or beef by persons desiring to avoid such food sources; as well as the assay of cattle feed to determine presence therein of ruminant-source proteins, which are a potential source of bovine spongiform encephalopathy (BSE), commonly referred to as “mad cow disease.” The assays amplify the predetermined unique SINEs and the resulting amplified mixture is then evaluated qualitatively by electrophoresis on gel containing ethidium bromide or quantitatively by SYBR Green-based detection or TaqMan chemistry.

Abstract: The way to design a “filled” site (which contains an interspersed element) primer set to target a particular locus is to design one of the two primers such that it encompasses that unique information (e.g., interspersed element+flanking genomic sequence+direct repeat). The way to design an “empty” site primer is to design one of the two primers such that the entire direct repeat sequence in addition to flanking genomic sequence is included on both sides. To improve efficiency, the “empty” site primer designed around the direct repeat should not be too long. This primer design of the present invention allows for the ability to test any type of interspersed genetic element containing characteristic direct repeat sequences (direct repeats). This gives the option of many new polymorphic marker sites because Alu elements are not the only interspersed genetic elements having direct repeats flanking their core sequence.

Abstract: A family of PCR assays is disclosed for determining, both qualitatively and quantitatively, presence of material from a predetermined species source and for quantifying the amount of such material. The assays are based respectively on SINEs uniquely characteristic of pig species, cow species, chicken species, and ruminant sub-order, and having a high copy number. The assays disclosed permit rapid, inexpensive evaluation of meat samples to facilitate elimination from their diet of pork or beef by persons desiring to avoid such food sources; as well as the assay of cattle feed to determine presence therein of ruminant-source proteins, which are a potential source of bovine spongiform encephalopathy (BSE), commonly referred to as “mad cow disease.” The assays amplify the predetermined unique SINEs and the resulting amplified mixture is then evaluated qualitatively by electrophoresis on gel containing ethidium bromide or quantitatively by SYBR Green-based detection or TaqMan chemistry.

Abstract: An assay for determining presence of human DNA in a sample in which non-human DNA may also be present and for quantitating such human DNA. One embodiment uses intra-Alu based PCR and another uses inter-Alu based PCR. The assays are performed without unique, expensive equipment. The assays are based on detection of multiple-copy Alu elements recently integrated into the human genome that are largely absent from non-human primates and other mammals.