International Journal of Legal Medicine

, Volume 130, Issue 3, pp 635–647 | Cite as

Evaluation of a 13-loci STR multiplex system for Cannabis sativa genetic identification

  • Rachel Houston
  • Matthew Birck
  • Sheree Hughes-Stamm
  • David GangitanoEmail author
Original Article


Marijuana (Cannabis sativa) is the most commonly used illicit substance in the USA. The development of a validated method using Cannabis short tandem repeats (STRs) could aid in the individualization of samples as well as serve as an intelligence tool to link multiple cases. For this purpose, a modified 13-loci STR multiplex method was optimized and evaluated according to ISFG and SWGDAM guidelines. A real-time PCR quantification method for C. sativa was developed and validated, and a sequenced allelic ladder was also designed to accurately genotype 199 C. sativa samples from 11 U.S. Customs and Border Protection seizures. Distinguishable DNA profiles were generated from 127 samples that yielded full STR profiles. Four duplicate genotypes within seizures were found. The combined power of discrimination of this multilocus system is 1 in 70 million. The sensitivity of the multiplex STR system is 0.25 ng of template DNA. None of the 13 STR markers cross-reacted with any of the studied species, except for Humulus lupulus (hops) which generated unspecific peaks. Phylogenetic analysis and case-to-case pairwise comparison of 11 cases using F st as genetic distance revealed the genetic association of four groups of cases. Moreover, due to their genetic similarity, a subset of samples (N = 97) was found to form a homogeneous population in Hardy-Weinberg and linkage equilibrium. The results of this research demonstrate the applicability of this 13-loci STR system in associating Cannabis cases for intelligence purposes.


Forensic DNA Forensic botany Cannabis sativa Short tandem repeats Reference population 



We would like to thank all staff and personnel at the U.S. Customs and Border Protection LSS Southwest Regional Science Center for their great assistance and help with this project.

Supplementary material

414_2015_1296_MOESM1_ESM.pptx (101 kb)
Suppl. Fig. 1 Electropherograms of homozygote Cannabis samples (at 60ºC, left) displaying the recovery of sister alleles when amplified at their specific annealing temperatures (53 or 55°°C, right). (PPTX 101 kb)
414_2015_1296_MOESM2_ESM.docx (13 kb)
ESM 2 (DOCX 12 kb)
414_2015_1296_MOESM3_ESM.docx (14 kb)
ESM 3 (DOCX 13 kb)


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Copyright information

© Springer-Verlag Berlin Heidelberg 2015

Authors and Affiliations

  1. 1.Department of Forensic Science, College of Criminal JusticeSam Houston State UniversityHuntsvilleUSA
  2. 2.New York Laboratory, U.S. Customs and Border Protection, U.S. Department of Homeland SecurityNewarkUSA

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