Comparison of Different DNA Isolation Protocols in Old and New Insect Samples

Muhammed Tatar; Aslıhan Esra Bildirici; Fatih Ölmez; Zemran Mustafa; Göksel Tozlu

Makale detayları

Comparison of Different DNA Isolation Protocols in Old and New Insect Samples

Yazarlar:

Muhammed Tatar | Aslıhan Esra Bildirici | Fatih Ölmez | Zemran Mustafa | Göksel Tozlu

Yayıncı:

Igdir University

Ana Başlıklar:

Genetik (Diğer) | Hayvan Hücresi ve Moleküler Biyoloji

Dergi:

Iğdır Üniversitesi Fen Bilimleri Enstitüsü Dergisi Vol.14 Issue 2, pp.594-603

Belge Tipi:

Makale

Yayın Tarihi:

1/2024

Belge Dili:

İngilizce

Document ID (Org No.):

2001317794

pISSN:

2536-4618

Anahtar kelimeler:

Özet:

 Insects belong to the Arthropoda phylum and are the most numerous animal group in the world, with more than 1.000.000 species. Insects ensure the sustainability of life on earth by pollinating plants, improving soil texture by decomposing organic materials, and providing nutrients to humans. DNA isolation is a necessity for performing analyses such as Sanger sequencing, DNA fingerprinting, and gene-genome analysis. Isolated insect DNA samples can be used to carry out studies such as species identification, taxonomic relations, comparison of insect-plant and microfauna relations, revealing genomic, population, and genetic diversity, bio-geography determination, combating agricultural pests, etc. In this study, it is aimed to compare three different isolation methods in terms of time and quality for obtaining genetic material from museum samples old and new samples of insects. The samples used in this investigation belong to the same insect species of Cerambycidae family Coleoptera. Samples collected at most 9-12 months ago are considered as new specimen, whereas old specimens consist samples of 7-28 years old. Three DNA isolation protocols: Phenol-Chloroform, Salting Out and DArT seq, are compared in this study. Considering quality gel images and A260280 and A260230 values in newly collected samples, the best results in terms of quality were obtained with the phenol-chloroform method. When A260280 values were examined alone, DArT seq protocol appears to be the most appropriate protocol in terms of purity in both old and new insect samples. In terms of time, the Phenol-chloroform method was found to be the shortest protocol lasting for about 2 h. Accordingly, the phenol-chloroform method was deduced to be the better choice for both old and new insect samples. After concentration adjustments, the obtained DNAs can be used for the next experimental stages or stored as stock material at -20 ºC. Determining the appropriate protocol to obtain high-quality DNA in a short time is important for the continuation of the studies. Our findings are important in terms of accelerating the studies for identification and determining systematic status and especially in examining the taxonomic relationships of the insect species. Using molecular taxonomy in addition to classical taxonomy allows the identification of insects on a species basis. These types of studies will generate useful information for researchers who study insect molecular biology.

 Insects belong to the Arthropoda phylum and are the most numerous animal group in the world, with more than 1.000.000 species. Insects ensure the sustainability of life on earth by pollinating plants, improving soil texture by decomposing organic materials, and providing nutrients to humans. DNA isolation is a necessity for performing analyses such as Sanger sequencing, DNA fingerprinting, and gene-genome analysis. Isolated insect DNA samples can be used to carry out studies such as species identification, taxonomic relations, comparison of insect-plant and microfauna relations, revealing genomic, population, and genetic diversity, bio-geography determination, combating agricultural pests, etc. In this study, it is aimed to compare three different isolation methods in terms of time and quality for obtaining genetic material from museum samples old and new samples of insects. The samples used in this investigation belong to the same insect species of Cerambycidae family Coleoptera. Samples collected at most 9-12 months ago are considered as new specimen, whereas old specimens consist samples of 7-28 years old. Three DNA isolation protocols: Phenol-Chloroform, Salting Out and DArT seq, are compared in this study. Considering quality gel images and A260280 and A260230 values in newly collected samples, the best results in terms of quality were obtained with the phenol-chloroform method. When A260280 values were examined alone, DArT seq protocol appears to be the most appropriate protocol in terms of purity in both old and new insect samples. In terms of time, the Phenol-chloroform method was found to be the shortest protocol lasting for about 2 h. Accordingly, the phenol-chloroform method was deduced to be the better choice for both old and new insect samples. After concentration adjustments, the obtained DNAs can be used for the next experimental stages or stored as stock material at -20 ºC. Determining the appropriate protocol to obtain high-quality DNA in a short time is important for the continuation of the studies. Our findings are important in terms of accelerating the studies for identification and determining systematic status and especially in examining the taxonomic relationships of the insect species. Using molecular taxonomy in addition to classical taxonomy allows the identification of insects on a species basis. These types of studies will generate useful information for researchers who study insect molecular biology.

 Insects belong to the Arthropoda phylum and are the most numerous animal group in the world, with more than 1.000.000 species. Insects ensure the sustainability of life on earth by pollinating plants, improving soil texture by decomposing organic materials, and providing nutrients to humans. DNA isolation is a necessity for performing analyses such as Sanger sequencing, DNA fingerprinting, and gene-genome analysis. Isolated insect DNA samples can be used to carry out studies such as species identification, taxonomic relations, comparison of insect-plant and microfauna relations, revealing genomic, population, and genetic diversity, bio-geography determination, combating agricultural pests, etc. In this study, it is aimed to compare three different isolation methods in terms of time and quality for obtaining genetic material from museum samples old and new samples of insects. The samples used in this investigation belong to the same insect species of Cerambycidae family Coleoptera. Samples collected at most 9-12 months ago are considered as new specimen, whereas old specimens consist samples of 7-28 years old. Three DNA isolation protocols: Phenol-Chloroform, Salting Out and DArT seq, are compared in this study. Considering quality gel images and A260280 and A260230 values in newly collected samples, the best results in terms of quality were obtained with the phenol-chloroform method. When A260280 values were examined alone, DArT seq protocol appears to be the most appropriate protocol in terms of purity in both old and new insect samples. In terms of time, the Phenol-chloroform method was found to be the shortest protocol lasting for about 2 h. Accordingly, the phenol-chloroform method was deduced to be the better choice for both old and new insect samples. After concentration adjustments, the obtained DNAs can be used for the next experimental stages or stored as stock material at -20 ºC. Determining the appropriate protocol to obtain high-quality DNA in a short time is important for the continuation of the studies. Our findings are important in terms of accelerating the studies for identification and determining systematic status and especially in examining the taxonomic relationships of the insect species. Using molecular taxonomy in addition to classical taxonomy allows the identification of insects on a species basis. These types of studies will generate useful information for researchers who study insect molecular biology.

 Eklem bacaklılar şubesinde yer alan ve 1.000.000dan fazla tür sayısı ile dünyada en kalabalık hayvan grubu, böceklerdir. Böcekler, bitkileri tozlaştırarak, organik maddeleri ayrıştırarak, toprak dokusunu iyileştirerek ve insanlara besin sağlayarak yeryüzündeki yaşamın sürdürülebilirliğini sağlar. DNA izolasyonu, Sanger dizileme, DNA parmak izi, gen-genom analizi gibi analizlerin yapılabilmesi için bir zorunluluktur. İzole edilmiş böcek DNA örnekleri; tür tanımlama, taksonomik ilişkiler, böcek-bitki ve -mikrofauna ilişkilerinin karşılaştırılması, genomik, popülasyon ve genetik çeşitliliğin ortaya çıkarılması, biyocoğrafya tespiti, tarımsal zararlılarla mücadele vb. çalışmaların yürütülmesinde kullanılabilir. Bu çalışmada, müze eski ve yeni böcek örneklerinden genetik materyal elde etmek için üç farklı izolasyon yönteminin zaman ve kalite açısından karşılaştırılması amaçlanmıştır. Çalışmada kullanılan örnekler, Cerambycidae familyasından Coleoptera aynı böcek türüne aittir. En fazla 9-12 ay önce toplanan numuneler yeni örnek olarak kabul edilirken, eski örnekler 7-28 yıllık numunelerden oluşmaktadır. Karşılaştırma için, Fenol-Kloroform, Salting Out ve DArT Seq olmak üzere üç farklı DNA izolasyon protokolü takip edilmiştir. Yeni toplanan örneklerde, kalite jeli görüntüleri ve A260280 ve A260230 değerleri dikkate alındığında, kalite açısından en iyi sonuçlar fenol-kloroform yöntemi ile elde edilmiştir. A260280 değerleri tek başına incelendiğinde hem eski hem de yeni böcek örneklerinde saflık açısından DArT seq protokolü en uygun protokol olarak görünmektedir. Zaman açısından, fenol-kloroform yöntemi yaklaşık 2 saat süren en kısa protokoldür. Buna göre, fenol-kloroform yönteminin hem eski hem de yeni böcek örnekleri için daha iyi bir seçim olduğu sonucuna varılmıştır. Konsantrasyon ayarlanmasından sonra elde edilen DNAlar, sonraki deney aşamaları için kullanılabilir veya -20de stok materyal olarak saklanabilir. Bu bağlamda, kısa sürede yüksek-kalite DNA elde etmek için en uygun protokolün belirlenmesi, çalışmaların devamlılığı için önemlidir. Bulgularımız, teşhis ve sistematik durum belirleme çalışmalarına hız kazandırması ve özellikle böcek türlerinin taksonomik ilişkilerinin incelenmesi açısından önemlidir. Klasik taksonomiye ek olarak moleküler taksonominin kullanılması, böceklerin tür bazında tanımlanmasına olarak sağlar. Bu tür çalışmalar, böcek moleküler biyolojisi üzerine çalışan araştırmacılar için faydalı bilgiler üretecektir.

Makale detayları

Comparison of Different DNA Isolation Protocols in Old and New Insect Samples

Yazarlar:

Muhammed Tatar | Aslıhan Esra Bildirici | Fatih Ölmez | Zemran Mustafa | Göksel Tozlu

Yayıncı:

Igdir University

Ana Başlıklar:

Genetik (Diğer) | Hayvan Hücresi ve Moleküler Biyoloji

Dergi:

Iğdır Üniversitesi Fen Bilimleri Enstitüsü Dergisi Vol.14 Issue 2, pp.594-603

Belge Tipi:

Makale

Yayın Tarihi:

1/2024

Belge Dili:

İngilizce

Document ID (Org No.):

2001317794

pISSN:

2536-4618

Insect

Coleoptera

Cerambycidae

DNA isolation

Genetic materialBöcek

Coleoptera

Cerambycidae

DNA İzolasyonu

Genetik materyal

Sayı

Makale detayları

Yazarlar: Muhammed Tatar | Aslıhan Esra Bildirici | Fatih Ölmez | Zemran Mustafa | Göksel Tozlu

Yayıncı: Igdir University

Ana Başlıklar: Genetik (Diğer) | Hayvan Hücresi ve Moleküler Biyoloji

Dergi: Iğdır Üniversitesi Fen Bilimleri Enstitüsü Dergisi Vol.14 Issue 2, pp.594-603

Belge Tipi: Makale

Yayın Tarihi: 1/2024

Belge Dili: İngilizce

Document ID (Org No.): 2001317794

pISSN: 2536-4618

Sayı

Yazarlar:

Muhammed Tatar | Aslıhan Esra Bildirici | Fatih Ölmez | Zemran Mustafa | Göksel Tozlu

Yayıncı:

Igdir University

Ana Başlıklar:

Genetik (Diğer) | Hayvan Hücresi ve Moleküler Biyoloji

Dergi:

Iğdır Üniversitesi Fen Bilimleri Enstitüsü Dergisi Vol.14 Issue 2, pp.594-603

Belge Tipi:

Makale

Yayın Tarihi:

1/2024

Belge Dili:

İngilizce

Document ID (Org No.):

2001317794

pISSN:

2536-4618