Extraction of high quality DNA from biological materials and calcified tissues

Abstract

Calcified tissues, such as bone and tooth, and some other sample types, such as those containing adhesive, present a challenge to standard extraction protocols. We have developed a lysis reagent, BTA™ lysis buffer, which is designed for use with PrepFiler™ Kit reagents. The BTA™ lysis buffer disrupts calcified tissue matrices and achieves effective extraction of DNA from pulverized bone and tooth samples. In addition, the BTA™ lysis buffer mildly but efficiently extracts DNA from challenging substrates like tape, chewing gum, and cigarette butts and, as with bone and tooth, DNA from these lysates is purified using established PrepFiler™ reagent extraction protocols. We successfully extracted DNA from powdered human bone samples, chewed gum and smoked cigarettes using BTA™ lysis buffer. Extraction yields for bone, gum and cigarette samples tested were consistent and reproducible. This extraction method efficiently removed potential PCR inhibitors from all samples tested, and C T values for the internal PCR control of Quantifiler ® Human DNA Quantification Kit were consistent and within the normal range. The DNA extracted from these samples also provided conclusive profiles that were free of PCR artifacts when amplified using the AmpFℓSTR ® Identifiler ® PCR Amplification Kit. The protocol is easily adapted for automation.