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Genomic DNA isolation methods from honey bee (Apis mellifera L.) Spermatheca

2022Conference object Open access
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dc.contributor.authorAriel Yadró, Carlos
dc.contributor.authorLopes, Ana
dc.contributor.authorHenriques, Dora
dc.contributor.authorSoltani, Chiraz
dc.contributor.authorMarques, Manoela
dc.contributor.authorWegener, Jakob
dc.contributor.authorMusin, Eduard
dc.contributor.authorPinto, M. Alice
dc.date.accessioned2022-10-17T13:19:09Z
dc.date.available2022-10-17T13:19:09Z
dc.date.issued2022
dc.description.abstractThe honey bee queen (Apis mellifera L.) has a polyandrous mating system, meaning that the queen mates on average with 17 drones from the surroundings in a congregation area. After the mating event, the spermatozoa of the drones are stored in an organ called spermatheca. Genetic analysis of the spermathecal content can provide an estimate of the genetic diversity and purity of the surrounding honey bee populations. This can be particularly useful for conservation and mating centers that need to monitor their populations’ genetic backgrounds. However, isolating enough DNA for genomic applications from such a small and complex matrix can be a challenge. Here, we compared the quantity and quality of DNA isolated using five methods: (i) phenol-chloroform-isopropanol, (ii) QIAamp DNA Minikit, (iii) QIAamp DNA Microkit, (iv) Macherey- Nagel Nucleospin Tissue, and (v) NEB Monarch Genomic DNA Purification Tissue. For each kit, when appropriate, variations including different isolation protocols, lysis incubation times, and the addition of RNA carrier were assayed. The quantity and quality of DNA extracted was assessed by spectrophotometric (SpectroStar®Nano LVis Plate) and fluorometric methods (Quantus ™ Fluorometer). Spectrophotometric quantification indicated nucleic acid concentrations ranging from 2.00 to 55.58 ng/μL, and in 91.43% of the cases, the A260/280 ratios were over 2.00, indicating an elevated presence of RNA. The fluorometric quantification, specific for double-stranded DNA, provided values ranging from 0.02 to 2.30 ng/μL. From the five methods, two alternative protocols of the commercial kit QIAamp DNA Microkit produced a sufficient DNA quantity (≥1.7 ng/μL measured by Quantus) for applications involving SNP genotyping, namely: the Tissue protocol with 6 hours of lysis incubation and the Tissue protocol with 3 hours of incubation, both with addition of RNA carrier. In contrast, overnight lysis decreased the DNA yield. The other methods generally produced low and/or inconsistent DNA recovery. According to our results, QIAamp DNA Microkit with the use of RNA carrier and lysis incubation times between 3 to 6 hours produce the required DNA quantities for SNP genotyping.pt_PT
dc.description.sponsorshipContributions of JW and EM were financed through the financial support of the German Federal Ministry for Food and Agriculture, through the intermediary of the Federal Office for Agriculture and Food (project no. 2818BM040).pt_PT
dc.description.versioninfo:eu-repo/semantics/publishedVersionpt_PT
dc.identifier.citationYadró, Carlos; Lopes, Ana Rita; Henriques, Dora; Soltani, Chiraz; Marques, Manoela; Wegener, Jakob; Musin, Eduard; Pinto, M. Alice (2022). Genomic DNA isolation methods from honey bee (Apis mellifera L.) Spermatheca. In ICBEES-International Congress on Bee Sciences. Onlinept_PT
dc.identifier.urihttp://hdl.handle.net/10198/26016
dc.language.isoengpt_PT
dc.peerreviewedyespt_PT
dc.rights.urihttp://creativecommons.org/licenses/by/4.0/pt_PT
dc.subjectDNA isolationpt_PT
dc.subjectApis melliferapt_PT
dc.subjectSpermathecapt_PT
dc.subjectConservationpt_PT
dc.titleGenomic DNA isolation methods from honey bee (Apis mellifera L.) Spermathecapt_PT
dc.typeconference object
dspace.entity.typePublication
oaire.citation.conferencePlaceOnlinept_PT
oaire.citation.endPage88pt_PT
oaire.citation.startPage88pt_PT
oaire.citation.titleICBEES-International Congress on Bee Sciencept_PT
person.familyNameLopes
person.familyNameHenriques
person.familyNamePinto
person.givenNameAna
person.givenNameDora
person.givenNameM. Alice
person.identifier.ciencia-idF014-6835-054D
person.identifier.ciencia-id291F-986F-07DA
person.identifier.ciencia-idF814-A1D0-8318
person.identifier.orcid0000-0002-1888-2262
person.identifier.orcid0000-0001-7530-682X
person.identifier.orcid0000-0001-9663-8399
person.identifier.scopus-author-id55761737300
person.identifier.scopus-author-id8085507800
rcaap.rightsopenAccesspt_PT
rcaap.typeconferenceObjectpt_PT
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relation.isAuthorOfPublicationd2abd09f-a90c-4cfb-9a60-7fc32f56184d
relation.isAuthorOfPublication0667fe04-7078-483d-9198-56d167b19bc5
relation.isAuthorOfPublication.latestForDiscoveryd2abd09f-a90c-4cfb-9a60-7fc32f56184d

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