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

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

Öz

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.

Anahtar Kelimeler

• Andrena,DNA,moleküler,müze örnekleri,kum arıları

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