Gene Expression of Flavanone 3-Hydroxylase (F3H), Anthocyanidin Synthase (ANS), and p-Coumaroyl Ester 3-Hydroxilase (C3H) in Tzimbalo Fruit

Juan M. Morales, Raúl H. Blas, Viviana P. Chiluisa-Utreras, Joel N. Flores, Geovanny D. Ortega

Abstract


The current investigation emphasises the expression of candidate genes for the future improvement of fruit quality; the objectives were to describe morphological variation on Solanum caripense Dunal (tzimbalo) ecotypes; identify gene expression of F3H and ANS; and analyse gene expression of C3H. Ecuadorian (BIO) and Peruvian (IBT) samples were utilized. Morphological descriptors for Solanum muricatum Aiton were used; RNA was isolated for identification of F3H and ANS transcripts in BIO-Ltg1 and BIO-Cyb1 through reverse transcription followed by semiquantitative PCR (RT-PCR); C3H relative expression was analysed in IBT-Lib1 for zero, five and fourteen days under influence of controlled conditions (10 ± 2 °C; 16 h day/8 h night) through reverse transcription followed by quantitative PCR (RT-qPCR). The cophenetic correlation (0.88) of conglomerate analysis (CA) pointed out good similarity for Ecuadorian ecotypes and two subgroups for Peruvian ecotypes. The first three principal components (PC) explained qualitatively 71.39% and quantitatively 81.34% of total variation; Fr-Flavour, Se-Diameter, Fl-CorollaColour, Fr-stripes, Fr-Length, Fr-PlacentLength and Fr-PlacentBreadth were characters that contributed more to the variability. The expression of F3H was identified in BIO-Ltg1. The expression of ANS was similar (BIO-Ltg1→48.20 ng/µL; BIO-Cyb1→36.19 ng/µL). The value of the mean fold change in C3H expression was 3.32, 4.52 and 6.24 for zero, five and fourteen days; C3H transcripts level was significantly different and increased 2.92 units after fourteen days. These results demonstrate the expression of F3H and ANS in BIO-Ltg1 and BIO-Cyb1, differential expression of C3H in IBT-Lib1, and focus the nutritional value of tzimbalo fruit.

Keywords


Reverse transcription; wild relatives; fruit quality; improvement; commercial potential

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References


E. Zuriaga, “Análisis de la variabilidad en poblaciones naturales de Solanum, secciones Lycopersicon y Basarthrum,†Doctoral thesis, Univ. Politécnica de Valencia, Spain, 2009. https://riunet.upv.es/handle/10251/6362

X. Palomeque, A. Verdugo, S. Criollo, and D. Peña, “Diversidad de frutales nativos comestibles Caricaceae - Solanaceae, fenología, usos y recolección de germoplasma en el Sur del Ecuador,†INIAP (Instituto Nacional de Investigaciones Agropecuarias), Estación Experimental Chuquipata, Granja Experimental Bullcay, 2003. https://repositorio.iniap.gob.ec/handle/41000/2368

D. S. Correll, Flora of Perú, Volume VIII, Parte V-B, Number 2., Field Museum of Natural History, 1967.

P. M. Jørgensen, and S. León-Yánez, Catalogue of the Vascular Plants of Ecuador, Saint Louis: Missouri Botanical Garden Press., 1999.

T. Särkinen, M. Baden, P. Gonzáles, M. Cueva, L. L. Giacomin, D. M. Spooner, R. Simon, H. Juárez, P. Nina, J. Molina, and S. Knapp, “Listado anotado de Solanum L. (Solanaceae) en el Perú,†Rev. Peru Biol. vol. 22, pp. 003-062, Apr. 2015. http://www.scielo.org.pe/scielo.php?script=sci_arttext&pid=S1727-99332015000100001

L. de la Torre, H. Navarrete, P. Muriel, M. Macía, and H. Balslev, Eds., Enciclopedia de las Plantas Útiles del Ecuador, Quito & Aarhus: Herbario QCA de la Escuela de Ciencias Biológicas de la Pontificia Universidad Católica del Ecuador & Herbario AAU del Departamento de Ciencias Biológicas de la Universidad de Aarhus, 2008.

V. Van den Eynden, E. Cueva, and O. Cabrera, Plantas Silvestres Comestibles del Sur del Ecuador, Quito, Ecuador: Univ. Politécnica Salesiana/Abya - Yala,1998.

C. B. Heiser, “Solanum caripense y el origen de Solanum muricatum,†Revista Politécnica vol. 1, no. 3, pp. 5-11, 1969.

B. C. Murray, K. R. Hammett, and F. D. Grigg, “Seed set and breeding system in the pepino Solanum muricatum, Ait., Solanaceae,†Sci. Hortic. vol. 49, issues 1-2, pp. 83-92, Jan. 1992. https://agris.fao.org/agris-search/search.do?recordID=NL19920041933

A. Rodríguez-Burruezo, J. Prohens, and A. Fita, “Breeding strategies for improving the performance and fruit quality of the pepino (Solanum muricatum): A model for the enhancement of underutilized exotic fruits,†Food Res. Int. vol. 44, pp. 1927–1935, Aug. 2011.

C. B. Heiser, “Origin and variability of the pepino (Solanum muricatum): A preliminary report,†Baileya vol. 12, pp. 151-158, 1964.

G. J. Anderson, and L. M. Bernardello, “The relationships of Solanum cochoae (Solanaceae), a new species from Peruâ€. Novon vol. 1, pp. 127-133, Autumn 1991.

G. J. Anderson, C. T. Martine, J. Prohens, and F. Nuez, “Solanum perlongistylum and S. catilliflorum, new endemic peruvian species of Solanum, section Basarthrum, are close relatives of the domesticated pepino, S. muricatum,†Novon vol. 16, pp. 161-167, Jun. 2006.

J. Prohens, S. Soler, and F. Nuez, “The effects of thermotherapy and sodium hypochlorite treatments on pepino seed germination, a crucial step in breeding programmes,†Ann. Appl. Biol. vol. 134, pp. 299-305, May. 1999.

J. Morales, and I. Vaca, “Propagación in vitro de tzímbalo (Solanum caripense Dunal),†RTE vol. 29, pp. 89-104, Dec. 2016. http://www.rte.espol.edu.ec/index.php/tecnologica/article/view/547

S. L. Bithell, B. A. Mckenzie, G. W. Bourdot, and G. D. Hill, “Germination requirements of laboratory stored seeds of Solanum nigrum and Solanum physalifolium,†N. Z. Plant Prot. vol. 55, pp. 222-227, Aug. 2002.

M. Ibrahim, M. Munira, M. Kabir, A. Islam, and M. Miah, “Seed germination and graft compatibility of wild Solanum as rootstock of tomato,†Online J. Biol. Sci. vol. 1, no. 8, pp. 701-703, Aug. 2001.

A. Taab, “Seed dormancy and germination in Solanum nigrum and S. physalifolium as influenced by temperature conditions,†Doctoral thesis, Swedish Univ. of Agricultural Sciences, Upsala Sweden., 2009. https://pub.epsilon.slu.se/2064/

J. Prohens, A. Rodríguez-Burruezo, M. Cámara, E. Torija, and F. Nuez, “Morphological and physico-chemical characteristics of fruits of pepino (Solanum muricatum), wild relatives (S. caripense and S. tabanoense) and interspecific hybrids: Implications in pepino breeding,†Eur. J. Hortic. Sci. vol. 70, pp. 224-230, Oct. 2005.

J. Prohens, F. Herraiz, M. Raigón, S. Vilanova, M. García-Martínez, P. Gramazio, M. Plazas, and A. Rodríguez-Burruezo, “Fruit composition diversity in land races and modern pepino (Solanum muricatum) varieties and wild related species,†Food Chem. vol. 15, pp. 49-58, Jul. 2016.

F. J. Herraiz, S. Vilanova, I. Andújar, D. Torrent, M. Plazas, P. Gramazio, and J. Prohens, “Morphological and molecular characterization of local varieties, modern cultivars and wild relatives of an emerging vegetable crop, the pepino (Solanum muricatum), provides insight into its diversity, relationships and breeding history,†Euphytica vol. 206, pp. 301-318, Apr. 2015.

F. J. Herraiz, J. Blanca, P. Ziarsolo, P. Gramazio, M. Plazas, G. Anderson, J. Prohens, and S. Vilanova, “The first de novo transcriptome of pepino (Solanum muricatum): assembly, comprehensive analysis and comparison with the closely related species S. caripense, potato and tomato,†BMC Genomics vol. 321, pp. 1-17, May. 2016.

A. Lucca, “Búsqueda de genes candidatos que controlen QTLs involucrados en la resistencia al estrés hídrico mediante el análisis de perfiles transcripcionales en especies silvestres de Solanum,†Doctoral thesis, Univ. de Buenos Aires, Agentina., 2011. https://repositorio.inta.gob.ar/xmlui/handle/20.500.12123/6192

M. B. Mantilla, “Transformación genética de la naranjilla, Solanum quitoense, mediante Agrobacterium tumefaciens,†Graduate thesis, Univ. San Francisco de Quito, Cumbayá, Ecuador, 2008. http://repositorio.usfq.edu.ec/handle/23000/930

A. Rodríguez-Burruezo, J. Prohens, and F. Nuez, “Wild relatives can contribute to the improvement of fruit quality in pepino (Solanum muricatum),†Euphytica vol. 129, pp. 311–318, Feb. 2003.

S. Bustin, “Quantification of mRNA using real-time RT-PCR: Trends and problems,†J. Mol. Endocrinol. vol. 29, pp. 23–39, Sep. 2002.

C. Orlando, P. Pinzani, and M. Pazzagli, “Developments in quantitative PCR,†Clin. Chem. Lab. Med. vol. 36, pp. 255-269, May. 1998.

J. Vandesompele, K. De-Preter, F. Pattyn, B. Poppe, N. Van-Roy, A. De-Paepe, and F. Speleman, “Accurate normalization of real-time quantitative RT-PCR data by geometric averaging of multiple internal control genes,†Genome Biol. vol. 3, pp. 0034.1–0034.11, Jun. 2002.

P. Fernández, “Cuantificación mediante la técnica de PCR en tiempo real. Usos y aplicaciones,†Instituto de Biotecnología, CICVyA, INTA-Castelar, 2011.

K. Livak, and T, “Schmittgen. Analysis of relative gene expression data using Real-Time Quantitative PCR and the 2−ΔΔð¶ð‘‡ method,†Methods vol. 25, pp. 402-408, Dec. 2001.

T. Holton, and E. Cornish, “Genetics and biochemistry of anthocyanin biosynthesis,†The Plant Cell vol. 7, pp. 1071-1083, Jul. 1995

Y. Liu, Y. Tikunov, R. Schouten, L. Marcelis, R. Visser, and A. Bovy, “Anthocyanin biosynthesis and degradation mechanisms in Solanaceous vegetables: a review,†Front. Chem. vol. 6, pp. 1-17, Mar. 2018.

Y. Tanaka, and A. Ohmiya, “Seeing is believing engineering anthocyanin and carotenoid biosynthetic pathways,†Curr. Opin. Biotechnol. vol. 19, pp. 190–197, Apr. 2008.

L. Qi-Neng, and Y. Quing, “cDNA cloning and expression of anthocyanin biosynthetic genes in wild potato (Solanun pinnatisectum),†Afr. J. Biotechnol. vol. 5, pp. 811-818, May. 2006.

M. Jiang, Y. Liu, L. Ren, H. Lian, and H. Chen, “Molecular cloning and characterization of anthocyanin biosynthesis genes in eggplant (Solanum melongena L.),†Acta Physiol. Plant. vol. 38, Jun. 2016.

N. Martínez-Cruz, K. Arévalo-Niño, M. Verde-Star, C. Rivas-Morales, A. Oranday-Cárdenas, A. Núñez-González, and M. Morales-Rubio, “Antocianinas y actividad anti radicales libres de Rubus adenotrichus Schltdl (zarzamora),†Rev. Mex. Cienc. Farm. vol. 42, pp. 66-71, Aug. 2011.

P. Gramazio, J. Prohens, M. Plazas, I. Andújar, F. Herraiz, E. Castillo, S. Knapp, R. Meyer, and S. Vilanova, “Location of chlorogenic acid biosynthesis pathway and polyphenol oxidase genes in a new interspecific anchored linkage map of eggplant,†BMC Plant Biol. vol. 14, pp. 1-15, Dec. 2014.

M. Zaro, A. Vicente, A. Chaves, and A. Concellón, “Cambios en los antioxidantes fenólicos de berenjena violeta durante el desarrollo y almacenamiento refrigerado,†Rev. Iber. Tecnología Postcosecha vol. 17, pp. 86-92, 2016.

R. Payyavula, R. Shakya, V. Sengoda, J. Munyaneza, P. Swamy, and D. Navarre, “Synthesis and regulation of chlorogenic acid in potato: Rerouting phenylpropanoid flux in HQT-silenced lines,†Plant Biotechnol. J. vol. 13, pp. 551–564, 2015.

R. Niggeweg, A. Michael, and C. Martin, “Engineering plants with increased levels of the antioxidant chlorogenic acid,†Nat. Biotechnol. vol. 22, pp. 746–754, Jun. 2004.

C. André, R. Schafleitner, S. Legay, I. Lefèvre, C. Alvarado, G. Nomberto, L. Hoffmann, J. Hausman, Y. Larondelle, and D. Evers, “Gene expression changes related to the production of phenolic compounds in potato tubers grown under drought stress,†Phytochemistry vol. 70, pp. 1107–1116, Jun. 2009.

R. Robbins, “Phenolic acids in foods: An overview of analytical methodology,†J. Agric. Food Chem. vol. 51, pp. 2886–2887, May. 2003.

P. Glorio, Técnicas Modernas de Investigación en Ciencia de Alimentos (Aplicaciones de Biología Molecular y Uso de Lector de Micropozos), Univ. Nacional Agraria La Molina, Lima, Perú, 2010.

J. Morales, R. Blas, V. Chiluisa-Utreras, J. Flores, and G. Ortega, “Gene expression of flavanone 3-hydroxylase (F3H), anthocyanidin synthase (ANS), and p-coumaroyl ester 3-hydroxilase (C3H) in tzimbalo fruit,†in Proc. CIT 2020 - ESPE, 2020, paper 211, p. 10. https://cit2020.org/life-sciences/

IPGRI (International Plant Genetic Resources Institute)/COMAV (Instituto Universitario de Conservación y Mejora de la Agrodiversidad Valenciana), “Descriptors for pepino (Solanum muricatum),†IPGRI, Rome, Italy, 2004. https://agris.fao.org/agris-search/search.do?recordID=QJ2005000012

I. Madroñero, J. Rosero, L. Rodríguez, J. Navia, and C. Benavides, “Caracterización morfoagronómica de genotipos promisorios de papa criolla (Solanum tuberosum L. Grupo Andigenum) en Nariño,†Temas Agrarios vol. 18, pp. 50-66, Jul. - Dec. 2013.

M. Morales, G. Espinosa, R. Morales, B. Sánchez, A. Jiménez, and Y. Milián, “Caracterización morfológica y evaluación de resistencia a Fusarium oxysporum en especies silvestres del género Solanum sección Lycopersicon,†Rev. Colomb. Biotecnol. vol. 16, pp. 62-73, Jul. 2014.

Y. Zhang, Z. Hu, G. Chu, C. Huang, S. Tian, Z. Zhao, and G. Chen, “Anthocyanin accumulation and molecular analysis of anthocyanin biosynthesis-associated genes in eggplant (Solanum melongena L.),†J. Agric. Food Chem. vol. 62, pp. 2906−2912, Apr. 2014.

H. Feng, X. Huang, Q. Zhang, G. Wei, X. Wang, and Z. Kang, “Selection of suitable inner reference genes for relative quantification expression of microRNA in wheat,†Plant physiology and biochemistry: PPB/SFPV vol. 51, pp. 116–122, Feb. 2012.

F. Rivas, “Análisis de la expresión del gen PR-1, mediante la técnica de PCR en tiempo real (RT-PCR), en tomate (Solanum lycopersicum) infectado con Phytophtora infestans,†Graduate thesis, Univ. de las Fuerzas Armadas, Sangolquí, Ecuador, 2010. http://repositorio.espe.edu.ec/xmlui/handle/21000/2640

S. Ballou, K. Yun, C. Cheng, and B. de los Reyes, “Cold sensitivity gradient in tuber-bearing Solanum based on physiological and transcript profiles,†Crop Sci. vol. 47, pp. 2027-2035, 2007.

J. Stommel, G. Lightbourn, B. Winkel, and R. Griesbach, “Transcription factor families regulate the anthocyanin biosynthetic pathway in Capsicum annuum,†J. Amer. Soc. Hort. Sci. vol. 134, pp. 244-251, Mar. 2009.

J. Crisci, and M. López, “Introducción a la Teoría y Práctica de la Taxonomía Numérica,†Washington, DC, USA: Secretaría General de la Organización de los Estados Americanos. Programa Regional de Desarrollo Científico y Tecnológico, 1983.

R. Sokal, and C. Michener, “A statistical method for evaluating systematic relationships,†Univ. Kansas Sci. Bull. vol. 38, pp. 1409-1438, 1958.

C. Quispe, R. Mansilla, A. Chacón, and R. Blas, “Análisis de la variabilidad morfológica del “añu†Tropaeolum tuberosum Ruiz & Pavón procedente de nueve distritos de la región Cusco,†Ecol. Apl. vol. 14, pp. 211-222, Jul. – Dec. 2015.

D. Montgomery, Diseño y Análisis de Experimentos, Univ. Estatal de Arizona, Editorial Limusa Wiley, 2004.

R. Sokal, and F. Rohlf, “The comparison on dendrograms by objective methods,†Taxon vol. 11, pp. 33-40, Feb. 1962.

D. Torrent, “Caracterización morfológica y molecular en pepino dulce (Solanum muricatum) y especies silvestres relacionadas,†Graduate thesis, Univ. Politécnica, Valencia, Spain, 2014. https://riunet.upv.es/handle/10251/46141?show=full

F. J. Herráiz, “Desarrollo de herramientas morfológicas y genómicas para el estudio del pepino dulce (Solanum muricatum) y especies relacionadas. Caracterización de su valor nutracéutico,†Doctoral thesis. Univ. Politécnica, Valencia, Spain, 2015. https://riunet.upv.es/handle/10251/61962

F. Camarena, J. Chura, and R. Blas, Mejoramiento Genético y Biotecnológico de Plantas, Lima, Perú: Univ. Nacional Agraria La Molina/AGROBANCO, 2014. https://www.agrobanco.com.pe/agrosaber/linea-editorial/

Y. Tanaka, N. Sasaki, and A. Ohmiya, “Biosynthesis of plant pigments: anthocyanins, betalains and carotenoids,†Plant J. vol. 54, pp. 733–749, May. 2008.

L. Cseke, A. Kirakosyan, P. Kaufman, and M. Westfall, Handbook of Molecular and CellularMethods in Biology and Medicine, Boca Ratón, USA: CRC Press, 2011.

T. Little, and F. Hills, Agricultural Experimentation: Design and Analysis, Wiley, 1978.

J. Hellemans, and J. Vandesompele, “qPCR data analysis – unlocking the secret to successful results". In PCR Ttroubleshooting and Optimization: The Essential Guide, S. Kennedy, and N. Oswald, Eds., U. K.: Caister Academic Press, 2011.

E. España, “Respuesta del cultivo de pepino dulce (Solanum muricatum Ait) a la fertilización química mediante el sistema de parcelas de omisión en el cantón Ibarra, provincia de Imbabura,†Graduate thesis, Univ. Técnica de Babahoyo, Carchi, Ecuador, 2015. http://dspace.utb.edu.ec/handle/49000/1053

F. J. Herraiz, D. Villaño, M. Plazas, S. Vilanova, F. Ferreres, J. Prohens, and D. Moreno, “Phenolic profile and biological activities of the pepino (Solanum muricatum) fruit and its wild relative S. caripense,†Int. J. Mol. Sci. vol. 17, pp. 1-15, Mar. 2016.




DOI: http://dx.doi.org/10.18517/ijaseit.11.2.13681

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