Carrier RNA is a key factor afecting fully integrated short tandem repeats profling in challenging forensic samples models

Abstract

Background: Short tandem repeats (STRs) are used today to provide discriminatory power for DNA fngerprinting. The present results showed that diferent factors may afect STR profles in challenging samples including DNA quan‑ tity, DNA quality, PCR inhibitors and storage time. In the present study, blood stain samples were applied on two types of fabrics (black cotton and denim) to compare the efciency of two diferent DNA-extraction methods (automated magnetic based beads method (EZ1), and manual organic method), with and without adding carrier RNA molecules, and to assess the quality and quantity of the extracted DNA and their capabilities for producing reportable STR-pro‑ fles in the presence of PCR inhibitors at two diferent storage times. Results: Carrier RNA caused a dramatic increase in DNA recovery from black cotton or denim using EZ1 in contrast to organic method. EZ1 was found to be preferred than organic, especially when a time passed over, while organic method was preferred when samples are available in small quantities. In addition, using carrier RNA within the organic method steps showed no improvement in STR profling. EZ1 with carrier RNA was preferred for bloodstained samples on fabrics with textile dyes (black dye or denim indigo), especially when stored for a long time. Conclusions: Denim was found to be more problematic than black cotton due to presence of challenging inhibitors (indigo dye). DNA concentration, storage time and types of fabrics are key factors for choosing the appropriate extrac‑ tion method for reportable STR profle. Using EZ1 with carrier RNA gives less dropout profle than not using it, or when using organic method even in presence or absence of carrier RNA. Anyway, innovation of more sensitive, more robust analytical protocols could result in a better understanding of these inhibitory samples.