Efficiency of different protocols of apple tree DNA extraction

Authors

  • Thyana Lays Brancher Universidade Federal de Lavras, Lavras, MG, Brasil
  • Maraisa Crestani Hawerroth Empresa de Pesquisa Agropecuária e Extensão Rural de Santa Catarina, Caçador, SC, Brasil
  • Marcus Vinícius Kvitschal Empresa de Pesquisa Agropecuária e Extensão Rural de Santa Catarina (Epagri)-Estação Experimental de Caçador/Pesquisador
  • Danielle Caroline Manenti Universidade Estadual de Maringá, Maringá, PR, Brasil.

DOI:

https://doi.org/10.5965/223811711732018361

Keywords:

Malus domestica, molecular biology, leaf tissue, polymerase chain reaction

Abstract

Several protocols to extract plant DNA exist, which can provide varying concentrations and quality of DNA due to the interaction between the protocol components and plant tissue compounds that affect the extraction, such as polyphenols and proteins. The objective of this study was to test the efficiency of different protocols for extracting DNA from apple tree leaves. Three DNA extraction protocols were tested: FastDNA® SPIN Kit - MP Biomedicals; adaptation from Qiagen DNeasy® Plant Mini Kit; adapted CTAB. Fresh and frozen young leaves of the genotypes M-10/09, SCS426 Venice and SCS427 Elenise were used. The purity and concentration of the DNA were analyzed using a spectrophotometer. The DNA extracted was tested using PCR with one primer related to regions of the genome that access transcription factors in apples (SAND). The highest concentrations of DNA were provided by the adapted CTAB protocol. Regarding purity, the adaptation from Qiagen DNeasy® Plant Mini Kit and the adapted CTAB protocols were considered satisfactory. Regarding DNA quality verified by PCR, all protocols were efficient for amplification of the target fragments. Therefore, all three protocols can be used for DNA extraction from apple leaves; however, the use of the adapted CTAB protocol showed high efficiency, due to the higher quality and concentration of DNA, with the lowest relative cost in the process of DNA extraction.

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Author Biographies

Thyana Lays Brancher, Universidade Federal de Lavras, Lavras, MG, Brasil

Biotecnologista Industrial. M.Sc. Produção Vegetal.

Aluna de Pós-Graduação em Biotecnologia Vegetal
(Doutorado) na Universidade Federal de Lavras - UFLA. Av. Doutor Sylvio Menicucci, 1001 - Kennedy, Lavras - MG, 37200-000

 

Maraisa Crestani Hawerroth, Empresa de Pesquisa Agropecuária e Extensão Rural de Santa Catarina, Caçador, SC, Brasil

Engenheira-agrônoma, D.Sc. Melhoramento Genético de
Plantas. Pesquisadora da Estação Experimental de
Caçador. Rua Abílio Franco, 1500. Bairro Bom Sucesso. Caçador - SC. Brasil. Cep 89501-032.

Marcus Vinícius Kvitschal, Empresa de Pesquisa Agropecuária e Extensão Rural de Santa Catarina (Epagri)-Estação Experimental de Caçador/Pesquisador

Engenheiro-agrônomo, D.Sc. Genética e Melhoramento.
Pesquisador da Estação Experimental de Caçador. Rua Abílio Franco, 1500. Bairro Bom Sucesso. Caçador - SC. Brasil. Cep 89501-032.

Danielle Caroline Manenti, Universidade Estadual de Maringá, Maringá, PR, Brasil.

Engenheira-agrônoma. M.Sc. Genética e Melhoramento.

Aluna de Pós-Graduação em Genética e Melhoramento (Doutorado) na Universidade Estadual de Maringá - UEM. Av. Colombo, 5790 - Zona 7, Maringá - PR, 87020-900.

 

References

ANTANAVICIUTE L et al. 2015. An inexpensive and rapid genomic DNA extraction protocol for rosaceous species. Journal of Horticultural Science and Biotechnology 90: 427-432.

AUBAKIROVA K et al. 2014. Evaluation of five protocols for DNA extraction from leaves of Malus sieversii, Vitis vinifera, and Armeniaca vulgaris. Genetics and Molecular Research 13: 1278-1287.

BAUMGARTNER IO et al. 2016. Development of SNP-based assays for disease resistance and fruit quality traits in apple (Malus × domestica Borkh.) and validation in breeding pilot studies. Tree Genetics & Genomes 12:35.

BIANCO L et al. 2014. Development and validation of a 20K single nucleotide polymorphism (SNP) whole genome genotyping array for apple (Malus x domestica Borkh). Plos One 9: 1-9.

CHENG FS et al. 1997. A DNA extraction protocol from various tissues in woody species. HortScience 32: 921-922.

CUSIN R et al. 2017. New biotechnological tools to accelerate scab-resistance trait transfer to apple. Genetics and Molecular Biology 40: 305-311.

DESJARDINS P & CONKLIN D. 2010. NanoDrop microvolume quantitation of nucleic acids. Journal of Visualized Experiments 45: 2565.

EDGE-GARZA DA et al. 2014. A high-throughput and cost-efficient DNA extraction protocol for the tree fruit crops of apple, sweet cherry, and peach relying on silica beads during tissue sampling. Molecular Breeding 34: 2225-2228.

FAOSTAT. 2014. Food and Agriculture Organization of The United Nations Statistics Database. Prodution/Crops. Disponível em: http://www.fao.org/faostat/en/#data/QC. Acesso em: 24 jul. 2017.

GUAN Y et al. 2015. QTLs detected for individual sugars and soluble solids content in apple. Molecular Breeding 35: 1-13.

LEFORT F & DOUGLAS GC. 1999. An efficient micro-method of DNA isolation from mature leaves of four hardwood tree species Acer, Fraxinus, Prunus and Quercus. Annals of Forest Science 56: 259-263.

MPBIO. 2016. Application manual FastDNA® SPIN Kit. Disponível em: https://www.mpbio.com/includes/protocol/FastDNA%20SPIN%20kit.pdf. Acesso em: 24 jul. 2017.

PERINI P et al. 2014. Reference genes for transcriptional analysis of flowering and fruit ripening stages in apple (Malus × domestica Borkh.). Molecular Breeding 34: 829-842.

REVERS LF et al. 2006. Uso prático de marcadores moleculares para seleção assistida no melhoramento de uvas de mesa apirênicas. Revista Brasileira de Fruticultura 28: 104-108.

ZHANG XJ et al. 2014. A and MdMYB1 allele-specific markers controlling apple (Malus x domestica Borkh.) skin color and suitability for marker-assisted selection. Genetics and Molecular Research 13: 9103-9114.

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Published

2018-09-26

How to Cite

BRANCHER, Thyana Lays; HAWERROTH, Maraisa Crestani; KVITSCHAL, Marcus Vinícius; MANENTI, Danielle Caroline. Efficiency of different protocols of apple tree DNA extraction. Revista de Ciências Agroveterinárias, Lages, v. 17, n. 3, p. 361–367, 2018. DOI: 10.5965/223811711732018361. Disponível em: https://periodicos.udesc.br/index.php/agroveterinaria/article/view/10456. Acesso em: 21 nov. 2024.

Issue

Vol. 17 No. 3 (2018)

Section

Research Article - Science of Plants and Derived Products

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Copyright (c) 2018 Revista de Ciências Agroveterinárias (Journal of Agroveterinary Sciences)

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