Advertisement

Functional & Integrative Genomics

, Volume 18, Issue 4, pp 439–455 | Cite as

GA3 application in grapes (Vitis vinifera L.) modulates different sets of genes at cluster emergence, full bloom, and berry stage as revealed by RNA sequence-based transcriptome analysis

  • Anuradha Upadhyay
  • Smita Maske
  • Satisha Jogaiah
  • Narendra Y. Kadoo
  • Vidya S. Gupta
Original Article

Abstract

In grapes (Vitis vinifera L.), exogenous gibberellic acid (GA3) is applied at different stages of bunch development to achieve desirable bunch shape and berry size in seedless grapes used for table purpose. RNA sequence-based transcriptome analysis was used to understand the mechanism of GA3 action at cluster emergence, full bloom, and berry stage in table grape variety Thompson Seedless. At cluster emergence, rachis samples were collected at 6 and 24 h after application of GA3, whereas flower clusters and berry samples were collected at 6, 24, and 48 h after application at full bloom and 3–4 mm berry stages. Seven hundred thirty-three genes were differentially expressed in GA3-treated samples. At rachis and flower cluster stage respectively, 126 and 264 genes were found to be significantly differentially expressed within 6 h of GA3 application. The number of DEG reduced considerably at 24 h. However, at berry stage, major changes occurred even at 24 h and a number of DEGs at 6 and 24 h were 174 and 191, respectively. As compared to upregulated genes, larger numbers of genes were downregulated. Stage-specific response to the GA3 application was observed as evident from the unique set of DEGs at each stage and only a few common genes among three stages. Among the DEGs, 67 were transcription factors. Functional categorization and enrichment analysis revealed that several transcripts involved in sucrose and hexose metabolism, hormone and secondary metabolism, and abiotic and biotic stimuli were enriched in response to application of GA3. A high correlation was recorded for real-time PCR and transcriptome data for selected DEGs, thus indicating the robustness of transcriptome data obtained in this study for understanding the GA3 response at different stages of berry development in grape. Chromosomal localization of DEGs and identification of polymorphic microsatellite markers in selected genes have potential for their use in breeding for varieties with improved bunch architecture.

Keywords

Vitis vinifera GA3 response Bunch architecture RNA seq Transcription factors Microsatellite markers 

Notes

Acknowledgements

This work was funded by the Department of Biotechnology, Government of India, New Delhi, under the grant no. BT/PR4856/PBD/16/967/2012.

Supplementary material

10142_2018_605_MOESM1_ESM.xlsx (91 kb)
ESM 1 (XLSX 90 kb)

References

  1. Achard P, Cheng H, De Grauwe L, Decat J, Schoutteten H, Moritz T, Van Der Straeten D, Peng J, Harberd NP (2006) Integration of plant responses to environmentally activated phytohormonal signals. Science 311:91–94.  https://doi.org/10.1126/science.1118642 CrossRefPubMedGoogle Scholar
  2. Berthet S, Demont-Caulet N, Pollet B, Bidzinski P, Cézard L, Le Bris P, Borrega N, Hervé J, Blondet E, Balzergue S, Lapierre C, Jouanin L (2011) Disruption of LACCASE4 and 17 results in tissue-specific alterations to lignification of Arabidopsis thaliana stems. Plant Cell 23:1124–1137.  https://doi.org/10.1105/tpc.110.082792 CrossRefPubMedPubMedCentralGoogle Scholar
  3. Bourquin V, Nishikubo N, Abe H, Brumer H, Denman S, Eklund M, Christiernin M, Teeri TT, Sundberg B, Mellerowicz EJ (2002) Xyloglucan endotransglycosylases have a function during the formation of secondary cell walls of vascular tissues. Plant Cell 14:3073–3088.  https://doi.org/10.1105/tpc.007773 CrossRefPubMedPubMedCentralGoogle Scholar
  4. Burdiak P, Rusaczonek A, Witoń D, Głów D, Karpiński S (2015) Cysteine-rich receptor-like kinase CRK5 as a regulator of growth, development, and ultraviolet radiation responses in Arabidopsis thaliana. J Exp Bot 66:3325–3337.  https://doi.org/10.1093/jxb/erv143 CrossRefPubMedPubMedCentralGoogle Scholar
  5. Chai L, Li Y, Chen S, Perl A, Zhao F, Ma H (2014) RNA sequencing reveals high resolution expression change of major plant hormone pathway genes after young seedless grape berries treated with gibberellin. Plant Sci 229:215–224.  https://doi.org/10.1016/j.plantsci.2014.09.010 CrossRefPubMedGoogle Scholar
  6. Chen L-Q, Hou B-H, Lalonde S, Takanaga H, Hartung ML, Qu X-Q, Guo W-J, Kim J-G, Underwood W, Chaudhuri B, Chermak D, Antony G, White FF, Somerville SC, Mudgett MB, Frommer WB (2010) Sugar transporters for intercellular exchange and nutrition of pathogens. Nature 468:527–532 http://www.nature.com/nature/journal/v468/n7323/abs/nature09606.html#supplementary-information CrossRefPubMedPubMedCentralGoogle Scholar
  7. Chen X, Lu S, Wang Y, Zhang X, Lv B, Luo L, Xi D, Shen J, Ma H, Ming F (2015) OsNAC2 encoding a NAC transcription factor that affects plant height through mediating the gibberellic acid pathway in rice. Plant J 82:302–314.  https://doi.org/10.1111/tpj.12819 CrossRefPubMedGoogle Scholar
  8. Cheng C, Jiao C, Singer SD, Gao M, Xu X, Zhou Y, Li Z, Fei Z, Wang Y, Wang X (2015) Gibberellin-induced changes in the transcriptome of grapevine (Vitis labrusca × V. vinifera) cv. Kyoho flowers. BMC Genomics 16:128.  https://doi.org/10.1186/s12864-015-1324-8 CrossRefPubMedPubMedCentralGoogle Scholar
  9. Choi Y-J, Hur YY, Jung S-M, Kim S-H, Noh J-H, Park S-J, Park K-S, Yun H-K (2013) Transcriptional analysis of Dehydrin1 genes responsive to dehydrating stress in grapevines. Hortic Environ Biotechnol 54:272–279.  https://doi.org/10.1007/s13580-013-0094-y CrossRefGoogle Scholar
  10. Chundawat BS, Singh R (1980) Effect of growth regulators on phalsa (Grewia asiatica L.) I. Growth and fruiting. Indian J Hortic 37:124–131Google Scholar
  11. Correa J, Mamani M, Munoz-Espinoza C, Laborie D, Munoz C, Pinto M, Hinrichsen P (2014) Heritability and identification of QTLs and underlying candidate genes associated with the architecture of the grapevine cluster (Vitis vinifera L.) Theor Appl Genet 127:1143–1162.  https://doi.org/10.1007/s00122-014-2286-y CrossRefPubMedGoogle Scholar
  12. Correa J, Ravest G, Laborie D, Mamani M, Torres E, Muñoz C, Pinto M, Hinrichsen P (2015) Quantitative trait loci for the response to gibberellic acid of berry size and seed mass in tablegrape (Vitis vinifera L.) Aust J Grape Wine Res 21:496–507.  https://doi.org/10.1111/ajgw.12141 CrossRefGoogle Scholar
  13. De Smet I, Signora L, Beeckman T, Inzé D, Foyer CH, Zhang H (2003) An abscisic acid-sensitive checkpoint in lateral root development of Arabidopsis. Plant J 33:543–555.  https://doi.org/10.1046/j.1365-313X.2003.01652.x CrossRefPubMedGoogle Scholar
  14. Derbyshire P, McCann MC, Roberts K (2007) Restricted cell elongation in Arabidopsis hypocotyls is associated with a reduced average pectin esterification level. BMC Plant Biol 7:31.  https://doi.org/10.1186/1471-2229-7-31 CrossRefPubMedPubMedCentralGoogle Scholar
  15. Dokoozlian NK, Peacock WL (2001) Gibberellic acid applied at bloom reduces fruit set and improves size of ‘Crimson Seedless’ table grapes. Hortscience 36:706–709Google Scholar
  16. Espinoza A, Contreras D, Orellana M, Pérez R, Aguirre C, Castro A, Riquelme A, Fichet T, Pinto M, Hinrichsen P (2009) Modulation by gibberellic acid of aquaporin genes expression during berry development of grapevine (Vitis vinifera L.) Acta Hortic 827:355–362CrossRefGoogle Scholar
  17. Estornell LH, Agustí J, Merelo P, Talón M, Tadeo FR (2013) Elucidating mechanisms underlying organ abscission. Plant Sci 199-200:48–60.  https://doi.org/10.1016/j.plantsci.2012.10.008 CrossRefPubMedGoogle Scholar
  18. Farrokhi N, Burton RA, Brownfield L, Hrmova M, Wilson SM, Bacic A, Fincher GB (2006) Plant cell wall biosynthesis: genetic, biochemical and functional genomics approaches to the identification of key genes. Plant Biotechnol J 4:145–167.  https://doi.org/10.1111/j.1467-7652.2005.00169.x CrossRefPubMedGoogle Scholar
  19. Finkelstein R, Reeves W, Ariizumi T, Steber C (2008) Molecular aspects of seed dormancy. Annu Rev Plant Biol 59:387–415.  https://doi.org/10.1146/annurev.arplant.59.032607.092740 CrossRefPubMedGoogle Scholar
  20. Fu X, Harberd NP (2003) Auxin promotes arabidopsis root growth by modulating gibberellin response. Nature 421:740–743CrossRefPubMedGoogle Scholar
  21. Fukazawa J, Nakata M, Ito T, Yamaguchi S, Takahashi Y (2010) The transcription factor RSG regulates negative feedback of NtGA20ox1 encoding GA 20-oxidase. Plant J 62:1035–1045.  https://doi.org/10.1111/j.1365-313X.2010.04215.x PubMedGoogle Scholar
  22. Grienenberger E, Besseau S, Geoffroy P, Debayle D, Heintz D, Lapierre C, Pollet B, Heitz T, Legrand M (2009) A BAHD acyltransferase is expressed in the tapetum of Arabidopsis anthers and is involved in the synthesis of hydroxycinnamoyl spermidines. Plant J 58:246–259.  https://doi.org/10.1111/j.1365-313X.2008.03773.x CrossRefPubMedGoogle Scholar
  23. Heinrich M, Hettenhausen C, Lange T, Wünsche H, Fang J, Baldwin IT, Wu J (2013) High levels of jasmonic acid antagonize the biosynthesis of gibberellins and inhibit the growth of Nicotiana attenuata stems. Plant J 73:591–606.  https://doi.org/10.1111/tpj.12058 CrossRefPubMedGoogle Scholar
  24. Jan A, Komatsu S (2006) Functional characterization of gibberellin-regulated genes in rice using microarray system. Genomics Proteomics Bioinformatics 4:137–144.  https://doi.org/10.1016/S1672-0229(06)60026-0 CrossRefPubMedPubMedCentralGoogle Scholar
  25. Kasimatis AN, Swanson FH, Vilas E, Peacock WL, Leavitt GM (1979) The relation of bloom-applied gibberellic acid to the yield and quality of Thompson Seedless raisins. Am J Enol Vitic 30:224–226Google Scholar
  26. Kushiro T, Okamoto M, Nakabayashi K, Yamagishi K, Kitamura S, Asami T, Hirai N, Koshiba T, Kamiya Y, Nambara E (2004) The Arabidopsis cytochrome P450 CYP707A encodes ABA 8′-hydroxylases: key enzymes in ABA catabolism. EMBO J 23:1647–1656CrossRefPubMedPubMedCentralGoogle Scholar
  27. Lashbrooke JG, Young PR, Dockrall SJ, Vasanth K, Vivier MA (2013) Functional characterisation of three members of the Vitis vinifera L. carotenoid cleavage dioxygenase gene family. BMC Plant Biol 13:1–17.  https://doi.org/10.1186/1471-2229-13-156 CrossRefGoogle Scholar
  28. Lee C, Teng Q, Zhong R, Ye Z-H (2012) Arabidopsis GUX proteins are glucuronyltransferases responsible for the addition of glucuronic acid side chains onto xylan. Plant Cell Physiol 53:1204–1216.  https://doi.org/10.1093/pcp/pcs064 CrossRefPubMedGoogle Scholar
  29. Li R, Wang W, Wang W, Li F, Wang Q, Xu Y, Wang S (2015) Overexpression of a cysteine proteinase inhibitor gene from Jatropha curcas confers enhanced tolerance to salinity stress. Electron J Biotechnol 18:368–375.  https://doi.org/10.1016/j.ejbt.2015.08.002 CrossRefGoogle Scholar
  30. Li J, Yu X, Lou Y, Wang L, Slovin JP, Xu W, Wang S, Zhang C (2015) Proteomic analysis of the effects of gibberellin on increased fruit sink strength in Asian pear (Pyrus pyrifolia). Sci Hortic 195:25–36.  https://doi.org/10.1016/j.scienta.2015.08.035 CrossRefGoogle Scholar
  31. Liu X, Baird WV (2003) The ribosomal small-subunit protein S28 gene from Helianthus annuus (Asteraceae) is down-regulated in response to drought, high salinity, and abscisic acid. Am J Bot 90:526–531.  https://doi.org/10.3732/ajb.90.4.526 CrossRefPubMedGoogle Scholar
  32. Looney NE, Wood DF (1977) Some cluster thinning and gibberellic acid effects on fruit set, berry size, vine growth and yield of de Chaunac grapes. Can J Plant Sci (Ottawa) 57:653–659CrossRefGoogle Scholar
  33. Lu K, Liang S, Wu Z, Bi C, Yu Y-T, Ma Y, Wang X-F, Zhang D-P (2016) Overexpression of an Arabidopsis cysteine-rich receptor-like protein kinase, CRK5, enhances abscisic acid sensitivity and confers drought tolerance. J Exp Bot 67:5009–5027.  https://doi.org/10.1093/jxb/erw266 CrossRefPubMedPubMedCentralGoogle Scholar
  34. Magome H, Yamaguchi S, Hanada A, Kamiya Y, Oda K (2004) dwarf and delayed-flowering 1, a novel Arabidopsis mutant deficient in gibberellin biosynthesis because of overexpression of a putative AP2 transcription factor. Plant J 37:720–729.  https://doi.org/10.1111/j.1365-313X.2003.01998.x CrossRefPubMedGoogle Scholar
  35. Mao D, Yu F, Li J, Tan D, Li J, Liu Y, Li X, Dong M, Chen L, Li D (2015) FERONIA receptor kinase interacts with S-adenosylmethionine synthetase and suppresses S-adenosylmethionine production and ethylene biosynthesis in Arabidopsis. Plant Cell Environ 38:2566–2574CrossRefPubMedGoogle Scholar
  36. May P (2000) From bud to berry, with special reference to inflorescence and bunch morphology in Vitis vinifera L. Aust J Grape Wine Res 6:82–98.  https://doi.org/10.1111/j.1755-0238.2000.tb00166.x CrossRefGoogle Scholar
  37. Molitor D, Rothmeier M, Behr M, Fischer S, Hoffmann L, Evers D (2011) Crop cultural and chemical methods to control grey mould on grapes. Vitis 50Google Scholar
  38. Molitor D, Behr M, Hoffmann L, Evers D (2012a) Benefits and drawbacks of pre-bloom applications of gibberellic acid (GA3) for stem elongation in Sauvignon Blanc. South Afr J Enol Vitic Stellenbosch 33:198–202Google Scholar
  39. Molitor D, Behr M, Hoffmann L, Evers D (2012b) Impact of grape cluster division on cluster morphology and bunch rot epidemic. Am J Enol Vitic 63:508–514.  https://doi.org/10.5344/ajev.2012.12041 CrossRefGoogle Scholar
  40. Mosesian RM, Nelson KE (1968) Effect on ‘Thompson Seedless’ fruit of gibberellic acid bloom sprays and double girdling. Am J Enol Vitic 19:37–46Google Scholar
  41. Nakata M, Yuasa T, Takahashi Y, Ishida S (2009) CDPK1, a calcium-dependent protein kinase, regulates transcriptional activator RSG in response to gibberellins. Plant Signal Behav 4:372–374CrossRefPubMedPubMedCentralGoogle Scholar
  42. Oulkar DP, Banerjee K, Ghaste MS, Ramteke SD, Naik DG, Patil SB, Jadhav MR, Adsule PG (2011) Multiresidue analysis of multiclass plant growth regulators in grapes by liquid chromatography/tandem mass spectrometry. J AOAC Int 94:968–977CrossRefPubMedGoogle Scholar
  43. Pérez FJ, Gómez M (2000) Possible role of soluble invertase in the gibberellic acid berry-sizing effect in sultana grape. Plant Growth Regul 30:111–116.  https://doi.org/10.1023/a:1006318306115 CrossRefGoogle Scholar
  44. Petti C, Hirano K, Stork J, DeBolt S (2015) Mapping of a cellulose-deficient mutant named dwarf1-1 in sorghum bicolor to the green revolution gene gibberellin20-oxidase reveals a positive regulatory association between gibberellin and cellulose biosynthesis. Plant Physiol 169:705–716.  https://doi.org/10.1104/pp.15.00928 CrossRefPubMedPubMedCentralGoogle Scholar
  45. Puig S (2014) Function and regulation of the plant COPT family of high-affinity copper transport proteins. Adv Bot 2014:9–9.  https://doi.org/10.1155/2014/476917 Google Scholar
  46. Richter R, Behringer C, Müller IK, Schwechheimer C (2010) The GATA-type transcription factors GNC and GNL/CGA1 repress gibberellin signaling downstream from DELLA proteins and PHYTOCHROME-INTERACTING FACTORS. Genes Dev 24:2093–2104.  https://doi.org/10.1101/gad.594910 CrossRefPubMedPubMedCentralGoogle Scholar
  47. Robertson M (2004) Two transcription factors are negative regulators of gibberellin response in the HvSPY-signaling pathway in barley aleurone. Plant Physiol 136:2747–2761.  https://doi.org/10.1104/pp.104.041665 CrossRefPubMedPubMedCentralGoogle Scholar
  48. Roper TR, Williams LE (1989) Net CO2 assimilation and carbohydrate partitioning of grapevine leaves in response to trunk girdling and gibberellic acid application. Plant Physiol 89:1136–1140CrossRefPubMedPubMedCentralGoogle Scholar
  49. Schulz P, Herde M, Romeis T (2013) Calcium-dependent protein kinases: hubs in plant stress signaling and development. Plant Physiol 163:523–530.  https://doi.org/10.1104/pp.113.222539 CrossRefPubMedPubMedCentralGoogle Scholar
  50. Shavrukov YN, Dry IB, Thomas MR (2004) Inflorescence and bunch architecture development in Vitis vinifera L. Aust J Grape Wine Res 10:116–124.  https://doi.org/10.1111/j.1755-0238.2004.tb00014.x CrossRefGoogle Scholar
  51. Street IH, Shah PK, Smith AM, Avery N, Neff MM (2008) The AT-hook-containing proteins SOB3/AHL29 and ESC/AHL27 are negative modulators of hypocotyl growth in Arabidopsis. Plant J 54:1–14.  https://doi.org/10.1111/j.1365-313X.2007.03393.x CrossRefPubMedGoogle Scholar
  52. Tello J, Aguirrezábal R, Hernáiz S, Larreina B, Montemayor MI, Vaquero E, Ibáñez J (2015) Multicultivar and multivariate study of the natural variation for grapevine bunch compactness. Aust J Grape Wine Res 21:277–289.  https://doi.org/10.1111/ajgw.12121 CrossRefGoogle Scholar
  53. Tello J, Torres-Pérez R, Grimplet J, Ibáñez J (2016) Association analysis of grapevine bunch traits using a comprehensive approach. Theor Appl Genet 129:227–242.  https://doi.org/10.1007/s00122-015-2623-9 CrossRefPubMedGoogle Scholar
  54. Ugare B, Banerjee K, Ramteke SD, Pradhan S, Oulkar DP, Utture SC, Adsule PG (2013) Dissipation kinetics of forchlorfenuron, 6-benzyl aminopurine, gibberellic acid and ethephon residues in table grapes (Vitis vinifera). Food Chem 141:4208–4214.  https://doi.org/10.1016/j.foodchem.2013.06.111 CrossRefPubMedGoogle Scholar
  55. Untergasser A, Cutcutache I, Koressaar T, Ye J, Faircloth BC, Remm M, Rozen SG (2012) Primer3—new capabilities and interfaces. Nucleic Acids Res 40:e115.  https://doi.org/10.1093/nar/gks596 CrossRefPubMedPubMedCentralGoogle Scholar
  56. Upadhyay A, Aher LB, Shinde MP, Mundankar KY, Datre A, Karibasappa GS (2013) Microsatellite analysis to rationalize grape germplasm in India and development of a molecular database. Plant Genet Resour 11:225–233.  https://doi.org/10.1017/S1479262113000117 CrossRefGoogle Scholar
  57. Upadhyay A, Jogaiah S, Maske SR, Kadoo NY, Gupta VS (2015) Expression of stable reference genes and SPINDLY gene in response to gibberellic acid application at different stages of grapevine development. Biol Plant 59:436–444.  https://doi.org/10.1007/s10535-015-0521-2 CrossRefGoogle Scholar
  58. Wang Z, Zhao F, Zhao X, Ge H, Chai L, Chen S, Perl A, Ma H (2012) Proteomic analysis of berry-sizing effect of GA3 on seedless Vitis vinifera L. Proteomics 12:86–94.  https://doi.org/10.1002/pmic.201000668 CrossRefPubMedGoogle Scholar
  59. Weaver RJ (1958) Effect of gibberellic acid on fruit set and berry enlargement in seedless grapes of Vitis vinifera. Nature 181:851–852CrossRefGoogle Scholar
  60. Weaver RJ (1975) Effect of time of application of potassium gibberellate on cluster development of ‘Zinfandel’ grapes. Vitis 14:97–102Google Scholar
  61. Weaver RJ, McCune S (1959) Effect of gibberellin on seedless Vitis vinifera. Hilgardia 29:247–275.  https://doi.org/10.3733/hilg.v29n06p247 CrossRefGoogle Scholar
  62. Woodger FJ, Millar A, Murray F, Jacobsen JV, Gubler F (2003) The role of GAMYB transcription factors in GA-regulated gene expression. J Plant Growth Regul 22:176–184.  https://doi.org/10.1007/s00344-003-0025-8 CrossRefGoogle Scholar
  63. Wu A-M, Lv S-Y, Liu J-Y (2007) Functional analysis of a cotton glucuronosyltransferase promoter in transgenic tobaccos. Cell Res 17:174–183CrossRefPubMedGoogle Scholar
  64. Xie C, Mao X, Huang J, Ding Y, Wu J, Dong S, Kong L, Gao G, Li CY, Wei L (2011) KOBAS 2.0: a web server for annotation and identification of enriched pathways and diseases. Nucleic Acids Res 39:W316–W322.  https://doi.org/10.1093/nar/gkr483 CrossRefPubMedPubMedCentralGoogle Scholar
  65. Zavala JA, Casteel CL, Nabity PD, Berenbaum MR, DeLucia EH (2009) Role of cysteine proteinase inhibitors in preference of Japanese beetles (Popillia japonica) for soybean (Glycine max) leaves of different ages and grown under elevated CO2. Oecologia 161:35–41.  https://doi.org/10.1007/s00442-009-1360-7 CrossRefPubMedGoogle Scholar
  66. Zeng H, Xu L, Singh A, Wang H, Du L, Poovaiah BW (2015) Involvement of calmodulin and calmodulin-like proteins in plant responses to abiotic stresses. Front Plant Sci 6:600.  https://doi.org/10.3389/fpls.2015.00600 PubMedPubMedCentralGoogle Scholar
  67. Zhang S, Cai Z, Wang X (2009) The primary signaling outputs of brassinosteroids are regulated by abscisic acid signaling. Proc Natl Acad Sci 106:4543–4548.  https://doi.org/10.1073/pnas.0900349106 CrossRefPubMedPubMedCentralGoogle Scholar
  68. Zhao J, Favero DS, Peng H, Neff MM (2013) Arabidopsis thaliana AHL family modulates hypocotyl growth redundantly by interacting with each other via the PPC/DUF296 domain. Proc Natl Acad Sci 110:E4688–E4697.  https://doi.org/10.1073/pnas.1219277110 CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2018

Authors and Affiliations

  • Anuradha Upadhyay
    • 1
  • Smita Maske
    • 1
  • Satisha Jogaiah
    • 1
    • 2
  • Narendra Y. Kadoo
    • 3
  • Vidya S. Gupta
    • 3
  1. 1.ICAR-National Research Centre for GrapesPuneIndia
  2. 2.ICAR-Indian Institute of Horticulture ResearchBengaluruIndia
  3. 3.CSIR-National Chemical LaboratoryPuneIndia

Personalised recommendations