Success of DNA extraction and PCR amplification from dry pinned sand bees (Andrena spp. Fabricius, 1775) using newly-designed primers

Year 2019, , 79 - 96, 01.03.2019

Canan Hazır , Clive H. Bock

https://doi.org/10.16970/entoted.460948

Abstract

The suitability of dry pinned museum specimens for DNA extraction of sand bees (Andrenaspp. Fabricius, 1775) (Hymenoptera: Andrenidae) and the effectiveness of existing and new primers used in DNA analysis of specimens for future studies were evaluated. A total 256 specimens were analyzed, including 222 dry pinned bee specimens representing 37 subgenera and 101 species and34 ethanol-preserved specimens belonging to 21 species. Several different protocols were tested for DNA extraction, and DNA was extracted from almost all of the specimens. The samples preserved in ethanol had the highest quality DNA.Of 31 primersets tested for amplification of the DNA, 14 of them were newly designed or redesigned. The amplified sequence length ranged from 130 to 1571 bp. DNA from 32 specimens belonging to 25 species was successfully amplified at three to four loci. This study demonstrates the importance ofstorage conditions for specimens possibly destined for later DNA extraction, and for selecting suitable primers when dealingwith older bee specimens. Some primers can be diagnostically informative provided appropriate gene regions are used.

Keywords

Andrena, DNA, molecular, museum specimens, sand bees

İğnelenerek kurutulmuş kum arılarından (Andrena spp. Fabricius, 1775) DNA eldesi ve yeni tasarlanmış primerler kullanıldığında PCR amplifikasyonu başarısı

Abstract

İğnelenerek kurutulmuş kum arısı müze örneklerinin (Andrenaspp. Fabricius, 1775) (Hymenoptera: Andrenidae) DNA ekstraksiyonu için uygunluğu incelenmiş ve gelecekteki çalışmalarda DNA analizi amacıyla yeni primerlerin etkinliği değerlendirilmiştir. Otuz yedi altcins ve 101 türü temsil eden 222 iğnelenmiş kuru arı örneği ve etanol içerisinde saklanmış 21 türe ait 34 arı örneği olmak üzere toplamda 256 örnek analiz edilmiştir. DNA ekstraksiyonu için birkaç farklı protokol denenmiş ve örneklerin tamamından DNA izole edilmiştir. En yüksek kaliteli DNA etanol içerisinde saklanan örneklerden elde edilmiştir. DNA amplifikasyonunda test edilen 31 primerden 14 tanesi ya yeni ya da yeniden tasarlanmıştır. Primerlerin sekans uzunluğu 130 ile 1571 bp arasında değişkenlik göstermiştir. Yirmi beş türe ait 32 örneğin üç-dört DNA lokusu başarılı bir şekilde çoğaltılmıştır. Bu çalışma, gelecekte DNA elde edilme ihtimali olan arı örneklerinin saklama koşullarının ve uygun primer seçiminin önemini ortaya koymuştur. Bazı primerler uygun gen bölgelerinin kullanılması şartıyla tür teşhislerini yapmaya yarayacak bilgileri sağlayabilirler.

Keywords

Andrena, DNA, moleküler, müze örnekleri, kum arıları

References

• Almeida, E. A. B & B. N. Danforth, 2009. Phylogeny of colletid bees (Hymenoptera: Colletidae) inferred from four nuclear genes. Molecular Phylogenetics and Evolution, 50: 290-309.
• Andersen, J. C & N. Mills, 2012. DNA extraction from museum specimens of parasitic Hymenoptera. PLoS ONE, 7 (10): e45549.
• Andersen, S. O., 2009. “Cuticle, 245-246”. In: Encyclopedia of Insects (Second Edition) (Eds. V. H. Resh & R. T. Cardé). Academic Press, United States, 1168 pp.
• Cameron, S. A & P. H. Williams, 2003. Phylogeny of bumble bees in the new world subgenus Fervidobombus (Hymenoptera: Apidae): congruence of molecular and morphological data. Molecular Phylogenetics and Evolution, 28: 552-563.
• Crane, S., 2011. DNA extraction from archival museum insect specimens modified from Qiagen® DNeasy Blood & Tissue Kit. (Web page: CC-BY http://dx.doi.org/10.6084/m9.figshare.741214) (Date accessed: December 2018).
• Danforth, B. N., H. Sauquet & L. Packer, 1999. Phylogeny of the bee genus Halictus (Hymenoptera: Halictidae) based on parsimony and likelihood analyses of nuclear EF-1'alpha' sequence data. Molecular Phylogenetics and Evolution, 13: 605-618.
• Danforth, B. N., 2013. Protocol for DNA extraction from small Hymenoptera. (Web page: http://www.danforthlab.entomology.cornell.edu/wp-content/uploads/dna_sequencing_protocols_v6.pdf) (Date accessed: December 2018).
• Danforth, B. N., J. Fang & S. Sipes, 2006. Analysis of family-level relationships in bees (Hymenoptera: Apiformes) using 28S and two previously unexplored nuclear genes: CAD and RNA polymerase II. Molecular Phylogenetics and Evolution, 39: 358-343.
• Danforth, B. N., S. Cardinal, C. Praz, E. A. B. Almeida & D. Michez, 2013. The impact of molecular data on our understanding of bee phylogeny and evolution. Annual Review of Entomology, 58: 57-78.
• Dillon, N., A. D. Austin & E. Bartowsky, 1996. Comparison of preservation techniques for DNA extraction from hymenopterous insects. Insect Molecular Biology, 1: 21-24.
• Dubitzky, A., 2005. Studies in Phylogeny and Biosystematics of Bees: The Bee Genus Andrena (Andrenidae) and the Tribe Anthophorini (Apidae) (Insecta: Hymenoptera: Apoidea). Fakultät für Biologie der Ludwig-Maximilians-Universität, (Published) PhD Thesis, München, Germany, 244 pp.
• Dubitzky, A., J. Plant & K. Schönitzer, 2010. Phylogeny of the bee genus Andrena Fabricius based on morphology (Hymenoptera: Andrenidae). Mitteilungen der Münchner Entomologischen Gesellschaft, 100: 137-202.
• Francoso, E. & M. C. Arias, 2013. Cytochrome c oxidase I primers for corbiculate bees: DNA barcode and mini-barcode. Molecular Ecology Resources, 13: 844-850.
• Gilbert, M. T. P., W. Moore, L. Melchior & M. Worobey, 2007. DNA extraction from dry museum beetles without conferring external morphological damage. PLoS ONE, 2 (3): e272.
• Hebert, P. D. N., E. H. Penton, J. M. Burns, D. H. Janzen & W. Hallwachs, 2004. Ten species in one: DNA barcoding reveals cryptic species in the neotropical skipper butterfly Astraptes fulgerator. Proceedings of the National Academy of Sciences of the United States of America, 101: 14812-14817.
• Hedtke, S. M., S. Patiny & B. N. Danforth, 2013. The bee tree of life: a supermatrix approach to apoid phylogeny and biogeography. BMC Evolutionary Biology, 13: 138.
• Hernandez-Triana, L. M., S. W. Prosser, M. A. Rodriguez-Perez, L. G. Chaverri, P. D. N. Hebert & T. R. Gregory, 2014. Recovery of DNA barcodes from blackfly museum specimens (Diptera: Simuliidae) using primer sets that target a variety of sequence length. Molecular Ecology Resources, 14: 508-518.
• Hines, H. M., S. A. Cameron & P. H. Williams, 2006. Molecular phylogeny of the bumble bee subgenus Pyrobombus (Hymenoptera: Apidae: Bombus) with insights into gene utility for lower-level analysis. Invertebrate Systematics, 20: 289-303.
• Klein, A-M., B. E. Vaissiere, J. H. Cane, I. Steffan-Dewenter, S. A. Cunningham, C. Kremen & T. Tscharntke, 2007. Importance of pollinators in changing landscapes for world crops. Proceedings of the Royal Society of London (B), 274: 303-313.
• Koulianos, S. & P. Schmid-Hempel, 2000. Phylogenetic relationships among bumble bees (Bombus Latreille) inferred from mitochondrial cytochrome b and cytochrome oxidase ı sequences. Molecular Phylogenetics and Evolution, 14 (3): 335-341.
• Larkin, L. L., J. L. Neff & B. B. Simpson, 2006. Phylogeny of the Callandrena subgenus of Andrena (Hymenoptera: Andrenidae) based on mitochondrial and nuclear DNA data: polyphyly and convergent evolution. Molecular Phylogenetics and Evolution, 38 (2): 330-343.
• Meusnier, I., G. A. C. Singer, J. F. Landry, D. A. Hickey, P. D. N. Hebert & M. Hajibabaei, 2008. A universal DNA mini-barcode for biodiversity analysis. BMC Genomics, 9: 214.
• Michez, D., S. Patiny & B. N. Danforth, 2009. Phylogeny of the bee family Melittidae (Hymenoptera: Anthophila) based on combined molecular and morphological data. Systematic Entomology, 34: 574-597.
• Nishiguchi, M. K., P. Doukakis, M. Egan, D. Kizirian, A. Phillips, L. Prendini, H. C. Rosenbaum, E. Torres, Y. Wyner, R. DeSalle & G. Giribet, 2002. “Methods and Tools in Biosciences and Medicine Techniques, 249-287”. In: Molecular Systematics and Evolution (Eds: R. Desalle, G. Giribet & W. Wheeler). Birkhäuser Verlag Basel, Switzerland, 309 pp.
• Peeters, C., M. Molet, C. C. Lin & J. Billen, 2017. Evolution of cheaper workers in ants: comparative study of exoskeleton thickness. Biological Journal of the Linnean Society, XX: 1-8.
• Praz, C. J., A. Müller, B. N. Danforth, T. L. Griswold, A. Widmer & S. Dorn, 2008. Phylogeny and biogeography of bees of the tribe Osmiini (Hymenoptera: Megachilidae). Molecular Phylogenetics and Evolution, 49: 185-197.
• Ramirez, S. R., J. C. Nieh, T. B. Quental, D. W. Roubik, V. L. Imperatriz-Fonseca & N. E. Pierce, 2010. A molecular phylogeny of the stingless bee genus Melipona (Hymenoptera: Apidae). Molecular Phylogenetics and Evolution, 56: 519-525.
• Reemer, M., D. S. J. Groenenberg, C. Van Achterberg & T. M. J. Peeters, 2008. Taxonomie assessment of Andrena rosae and A stragulata by DNA-sequencing (Hymenoptera: Apoidea: Andrenidae). Entomologia Generalis, 31 (1): 21-32.
• Rehan, S. M., T. W. Chapman, A. I. Craigie, M. H. Richards, S. J. B. Cooper & M. P. Schwarz, 2010. Molecular phylogeny of the small carpenter bees (Hymenoptera: Apidae: Ceratinini) indicates early and rapid global dispersal. Molecular Phylogenetics and Evolution, 55: 1042-1054.
• Schander, C. & K. M. Halanychi, 2003. DNA, PCR and formalinized animal tissue – a short review and protocols. Organisms Diversity and Evolution, 3: 195-205.
• Schmidt, S., C. Schmid-Egger, J. Moriniere, G. Haszprunar & P. D. N. Hebert, 2015. DNA barcoding largely supports 250 years of classical taxonomy: identifications for Central European bees (Hymenoptera, Apoidea partim). Molecular Ecology Resources, 15: 985-1000.
• Schwarz, M. P., S. M. Tierney, S. J. B. Cooper & N. J. Bull, 2004. Molecular phylogenetics of the Allodapine bee genus Braunsapis: A-T bias and heterogeneous substitution parameters. Molecular Phylogenetics and Evolution, 32: 110-122.
• Strange, J. P., J. Knoblett & T. Griswold, 2009. DNA amplification from pin-mounted bumble bees (Bombus) in a museum collection: effects of fragment size and specimen age on successful PCR. Apidologie, 40: 134-139.
• Ward, P. S., 2009. DNA extraction of dried (point-mounted) ant specimens modified v2.0. (personal communication).
• Willows-Munro, S. & M. C. Schoeman, 2015. Influence of killing method on Lepidoptera DNA barcode recovery. Molecular Ecology Resources, 15: 613-618.
• Woodard, S. H., J. D. Lozier, D. Goulson, P. H. Williams, J. P. Strange & S. Jha, 2015. Molecular tools and bumble bees: revealing hidden details of ecology and evolution in a model system. Molecular Ecology, 24: 2916-2936.
• Ye, J., G. Coulouris, I. Zaretskaya, I. Cutcutache, S. Rozen & T. L. Madden, 2012. Primer-BLAST: a tool to design target-specific primers for polymerase chain reaction. BMC Bioinformatics, 13: 134.
• Zimmermann, J., M. Hajibabaei M, D. C. Blackburn, J. Hanken, E. Cantin, J. Posfai & T. C. Evans Jr, 2008. DNA damage in preserved specimens and tissue samples: a molecular assessment. Frontiers in Zoology, 5:18.