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Nuclear, chloroplast, and mitochondrial data of a US cannabis DNA database

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A Correction to this article was published on 12 March 2018

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Abstract

As Cannabis sativa (marijuana) is a controlled substance in many parts of the world, the ability to track biogeographical origin of cannabis could provide law enforcement with investigative leads regarding its trade and distribution. Population substructure and inbreeding may cause cannabis plants to become more genetically related. This genetic relatedness can be helpful for intelligence purposes. Analysis of autosomal, chloroplast, and mitochondrial DNA allows for not only prediction of biogeographical origin of a plant but also discrimination between individual plants. A previously validated, 13-autosomal STR multiplex was used to genotype 510 samples. Samples were analyzed from four different sites: 21 seizures at the US–Mexico border, Northeastern Brazil, hemp seeds purchased in the US, and the Araucania area of Chile. In addition, a previously reported multi-loci system was modified and optimized to genotype five chloroplast and two mitochondrial markers. For this purpose, two methods were designed: a homopolymeric STR pentaplex and a SNP triplex with one chloroplast (Cscp001) marker shared by both methods for quality control. For successful mitochondrial and chloroplast typing, a novel real-time PCR quantitation method was developed and validated to accurately estimate the quantity of the chloroplast DNA (cpDNA) using a synthetic DNA standard. Moreover, a sequenced allelic ladder was also designed for accurate genotyping of the homopolymeric STR pentaplex. For autosomal typing, 356 unique profiles were generated from the 425 samples that yielded full STR profiles and 25 identical genotypes within seizures were observed. Phylogenetic analysis and case-to-case pairwise comparisons of 21 seizures at the US–Mexico border, using the Fixation Index (FST) as genetic distance, revealed the genetic association of nine seizures that formed a reference population. For mitochondrial and chloroplast typing, subsampling was performed, and 134 samples were genotyped. Complete haplotypes (STRs and SNPs) were observed for 127 samples. As expected, extensive haplotype sharing was observed; five distinguishable haplotypes were detected. In the reference population, the same haplotype was observed 39 times and two unique haplotypes were also detected. Haplotype sharing was observed between the US border seizures, Brazil, and Chile, while the hemp samples generated a distinct haplotype. Phylogenetic analysis of the four populations was performed, and results revealed that both autosomal and lineage markers could discern population substructure.

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Change history

  • 12 March 2018

    The original version of this article contained a mistake. In page 10 of the original article, the significant level (p > 0.01) is incorrect. The correct significant level is (p < 0.01). The original article has been corrected.

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Acknowledgements

The authors would like to thank all staff and personnel at the U.S. Customs and Border Protection LSSD Southwest Regional Science Center for their great assistance and help with this project. The authors would also like to thank Roberta Marriot and Alejandra Figueroa for their kind donation of marijuana DNA extracts. Lastly, the authors greatly appreciate Haleigh Agot for her assistance with the chloroplast quantitation method.

Funding

This study was partially funded by a Graduate Research Fellowship Award #2015-R2-CX-0030 (National Institute of Justice, Office of Justice Programs, U.S. Department of Justice). The opinions, findings, conclusions, or recommendations expressed in this presentation are those of the authors and do not necessarily reflect those of the National Institute of Justice.

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Authors and Affiliations

  1. Department of Forensic Science, College of Criminal Justice, Sam Houston State University, 1003 Bowers Blvd., Huntsville, TX, 77340-2525, USA

    Rachel Houston, Bobby LaRue, Sheree Hughes-Stamm & David Gangitano

  2. New York Laboratory, U.S. Customs and Border Protection, U.S. Department of Homeland Security, 1100 Raymond Blvd., Newark, NJ, 07102, USA

    Matthew Birck

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  1. Rachel Houston
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Correspondence to Rachel Houston.

Ethics declarations

This study does not involve living human subjects as defined under Title 45, Part 46.102(f), and is therefore exempt from Institutional Review Board (IRB) review.

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The authors declare that they have no competing interest.

Additional information

The original version of this article was revised: The original version of this article contained a mistake. In page 10 of the original article, the significant level (p > 0.01) is incorrect. The correct significant level is (p < 0.01).

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Houston, R., Birck, M., LaRue, B. et al. Nuclear, chloroplast, and mitochondrial data of a US cannabis DNA database. Int J Legal Med 132, 713–725 (2018). https://doi.org/10.1007/s00414-018-1798-4

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  • DOI: https://doi.org/10.1007/s00414-018-1798-4

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