Methods for preparing a biological sample obtained from an individual for use in a genetic testing assay

A method for preparing a biological sample obtained from a single individual for use in a genetic testing assay, the method comprising:

• identifying 50 to 10000 polymorphic loci for targeting in the biological sample;
• isolating cell-free DNA fragments from the biologic sample, wherein the isolated DNA fragments are between 100 bp and 220 bp and, wherein at least some of the isolated DNA fragments are target DNA molecules, and wherein the target DNA molecules are DNA fragments comprising one or more of the 50 to 10000 targeted polymorphic loci;
• ligating at least some of the isolated DNA fragments with a plurality of adaptors comprising a known tail sequence and a molecular barcode to produce adaptor-ligated DNA fragments such that most or all of the target DNA molecules in the sample have a unique molecular barcode, wherein the first known tail sequence comprises a first universal priming sequence;
• amplifying at least some of the adaptor-ligated DNA fragments using a universal primer that binds to the first universal priming sequence and 50 to 10000 different target-specific primers in one PCR reaction volume, followed by a second, nested PCR amplification, to obtain amplified adaptor-ligated DNA fragments comprising the known tail sequence, a target sequence, and a molecular barcode;
• and
• sequencing at least some of the amplified adaptor-ligated DNA fragments by clonal sequencing to generate allelic data for at least some of the target DNA molecules.