A robust genetic transformation protocol to obtain transgenic shoots of Solanum tuberosum L. cultivar ‘Kufri Chipsona 1’

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The genetic transformation of plants is an important biotechnological tool used for crop improvement for many decades. The present study was focussed to investigate various factors affecting genetic transformation of potato cultivar ‘Kufri Chipsona 1’. It was observed that explants pre-cultured for 2 days on MS2 medium (MS medium containing 10 µM silver nitrate, 10 µM BA, 15 µM GA3), injured with a surgical blade and co-cultivated with Agrobacterium tumefaciens strain EHA105 [O.D600 (0.6)] for 2 days results in maximum transient β-glucuronidase (GUS) expression. The addition of 100 µM acetosyringone in MS2 medium also increased rate of transient GUS expression in both the explants. Clumps of putative transgenic shoots were regenerated using the optimised culture conditions from leaf and internodal explants. The stable integration of T-DNA was established using histochemical staining for GUS and amplification of DNA fragment specific to nptII and uidA genes. Within the clumps, around 67.85% of shoots showed uniform GUS expression in all the tissues and about 32.15% shoots show intermittent GUS expression establishing chimeric nature. Uniform GUS staining of the tissue was used as initial marker of non-chimeric transgenic shoots. Quantitative expression of nptII transgene was found to be directly proportional to uniformity of GUS staining in transgenic shoots. The present investigation indicated that manipulation of culture conditions and the medium composition may help to get transgenic shoots with uniform expression of transgene in all the tissues of potato cultivar ‘Kufri Chipsona 1’.

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  1. Aggarwal D, Kumar A, Reddy MS (2011) Agrobacterium tumefaciens mediated genetic transformation of selected elite clone(s) of Eucalyptus tereticornis. Acta Physiol Plant 33:1603–1611.

  2. Alvarez R, Ordas RJ (2007) Improved genetic transformation protocol for cork oak (Quercus suber L.). Plant Cell Tiss Organ Cult 91:45–52.

  3. An G, Watson BD, Chiang CC (1986) Transformation of tobacco, tomato, potato and Arabidopsis thaliana using binary Ti vector system. Plant Physiol 81:301–305.

  4. APEDA (2003) Agricultural & processed food products export development authority post harvest manual for export of potatoes

  5. Bairwa A, Venkatasalam EP, Sudha R, Umamaheswari R, Sharma S, Singh BP et al (2016) Managementof late blight disease in Kharif potato at Karnataka. Potato J 43:173–181

  6. Banerjee AK, Prat S, Hannapel DJ (2006) Efficient production of transgenic potato (S. tuberosum L. ssp. andigena) plants via Agrobacterium tumefaciens mediated transformation. Plant Sci 170:732–738.

  7. Barrell PJ, Meiyalaghan S, Jacobs JME, Conner AJ (2013) Applications of biotechnology and genomics in potato improvement. Plant Biotechnol J 11:907–920.

  8. Beaujean A, Sangwan RS, Lecardonnel A, Sangwan-Norreel BS (1998) Agrobacterium-mediated transformation of three economically important potato cultivars using sliced internodal explants: an efficient protocol of transformation. J Exp Bot 49:1589–1595.

  9. Bhowmik SSD, Cheng AY, Long H, Tan GZH, Hoang TML, Karbaschi MR et al (2019) Robust genetic transformation system to obtain non-chimeric transgenic chickpea. Front Plant Sci 10:524

  10. Breseghello F, Coelho ASG (2013) Traditional and modern plant breeding methods with examples in rice (Oryza sativa L.). J Agric Food Chem 61:8277–8286.

  11. Chakraborty S, Chakraborty N, Datta A (2000) Increased nutritive value of transgenic potato by expressing a non-allergenic seed albumin gene from Amaranthus hypochondriacus. Proc Natl Acad Sci USA 97:3724–3729.

  12. Chakravarty B, Pruski GW, Flinn B, Gustafson V, Sharon R (2007) Genetic transformation in potato: approaches and strategies. Am Potato J 84:301–311.

  13. Chang S, Puryear J, Cairney J (1993) A simple and efficient method for isolating RNA from pine trees. Plant Mol Biol Rep 11:113–116.

  14. Chateau S, Sangwan RS, Sangwan-Norreel BS (2000) Competance of Arabidopsis thaliana genotypes and mutants for Agrobacterium tumefaciens-mediated gene transfer: role of phytohormones. J Exp Bot 51:1961–1968.

  15. Chen P-Y, Wang C-K, Soong S-C, To K-Y (2003) Complete sequence of the binary vector pBI121 and its applications in cloning T-DNA insertion from transgenic plants. Mol Breed 11:287–293.

  16. Conner AJ, Williams MK, Abernethy DJ, Fletcher PJ, Genet RA (1994) Field performance of transgenic potatoes. N Z J Crop Hortic 22:361–371.

  17. Conner AJ, Jacobs JME, Genet RA (1997) Transgenic potatoes versus “traditional” potatoes: what’s the difference? In: McLean GD, Waterhouse PM, Evans G, Gibbs MJ (eds) Commercialization of transgenic crops: risk, benefit and trade considerations. Cooperative Research Centre for Plant Science and Bureau of Resource Sciences, Canberra, pp 23–36

  18. Dale PJ, Hampson KK (1995) An assessment of morphogenic and transformation efficiency in a range of varieties of potato (Solanum tuberosum). Euphytica 85:101–108.

  19. De Block M (1988) Genotype-independent leaf disk transformation of potato (Solanum tuberosum) using Agrobacterium tumefaciens. Theor Appl Genet 76:767–774.

  20. De Vetten N, Wolters AM, Raemakers K, van der Meer I, ter Stege R, Heeres E, Heeres P, Visser R (2003) A transformation method for obtaining marker-free plants of a cross-pollinating and vegetatively propagated crop. Nat Biotechnol 21:439–442

  21. Doyle JJ, Doyle JL (1990) Isolation of plant DNA from fresh tissue. Focus 12:13–15

  22. Faize M, Faize L, Burgos L (2010) Using quantitative real-time PCR to detect chimeras in transgenic tobacco and apricot and to monitor their dissociation. BMC Biotechnol 10:53.

  23. FAOSTAT (2019) Food and Agriculture organisation of the United Nations (;2019). Accessed 4 Oct 2019

  24. Farhanah A, Suharsono S, Wattimena GA, Widyastuti U (2017) Genetic engineering of potato plant (Solanum tuberosum L.) cv. JalaIpam with MmPMA gene encoding plasma membrane H+-ATPase. Pak J Biotechnol 14:37–42

  25. Fatahillah, Suharsono S, Astuti UW (2016) Genetic transformation of potato (Solanum tuberosum L.) cv. Nooksack with FBPase/ClRan1 genes mediated by Agrobacterium tumefaciens. Pak J Biotechnol 13:187–192

  26. Feher A, Felfoldi K, Preiszner J (1991) PEG- mediated transformation of leaf protoplast of Solanum tuberosum L. cultivar. Plant Cell Tiss Organ Cult 27:105–114.

  27. Felcher KJ, Douches DS, Kirk WW, Hammerschmidt R (2003) Expression of a fungal glucose oxidase gene in three potato cultivars with different susceptibility to late blight (Phytophthora infestans Mont. deBary). J Am Soc Hortic Sci 128:238–245.

  28. Godwin I, Todd G, Ford-Lloyd B, Newbury HJ (1991) The effect of acetosyringone and pH on Agrobacterium-mediated transformation vary according to plant species. Plant Cell Rep 9:671–675.

  29. Gohlke J, Deeken R (2014) Plant responses to Agrobacterium tumefaciens and crown gall development. Front Plant Sci 5:155.

  30. GOI (2018) Monthly report on potato. Horticulture statistic division. Department of Agriculture, Cooperation and farmers welfare

  31. Han EH, Goo YM, Lee MK, Lee SW (2015) An efficient transformation method for a potato (Solanum tuberosum L. var. Atlantic). J Plant Biotechnol 42:77–82.

  32. Heeres P, Schippers-Rozenboom M, Jacobsen E, Visser RGF (2002) Transformation of a large number of potato varieties: genotype dependent variation in efficiency and somaclonal variability. Euphytica 124:13–22.

  33. Hess KM, Dudley MW, Lynn DG, Joerger RD, Binns AN (1991) Mechanism of phenolic activation of Agrobacterium virulence genes: development of a specific inhibitor of bacterial sensor/response systems. PNAS 88:7854–7858

  34. Hood EE, Helmer GL, Fraley RT, Chilton MD (1986) The hypervirulence of Agrobacterium tumefaciens A281 is encoded in a region of pTiBo542 outside of T-DNA. J Bacteriol 168:1291–1301.

  35. Jefferson RA, Kavanagh TA, Bevan MW (1987) GUS fusions: β-glucuronidase as a sensitive and versatile gene fusion marker in higher plants. EMBO J 6:3901–3907

  36. Jongedijk E, De Schutter AAJM, Stolte T, Van den Elzen PJM, Cornelissen BJC (1992) Increased resistance to potato virus-x and preservation of cultivar properties in transgenic potato under field conditions. Bio/Technology 10:422–429.

  37. Jube S, Borthakur D (2009) Development of an Agrobacterium mediated transformation protocol for the tree-legume Leucaena leucocephala using immature zygotic embryos. Plant Cell Tiss Organ Cult 96:325–333.

  38. Kamrani M, Ebadi A, Shiri M (2015) Effect of explant, genotype and plant growth regulators on regeneration and Agrobacterium-mediated transformation of potato. J Agron 14:227–233.

  39. Kaur A, Reddy MS, Kumar A (2017) Efficient, one step and cultivar independent shoot organogenesis of potato. Physiol Mol Biol-Plant 23:461–469.

  40. Keeling KM, Brooks DA, Hopwood JJ, Li P, Thompson JN, Bedwell DM (2001) Gentamicin-mediated suppression of Hurler syndrome stop mutations restores a low level of alpha-Liduronidase activity and reduces lysosomal glycosaminoglycan accumulation. Hum Mol Genet 10:291–299.

  41. Khatun A, Hasan MM, Bachchu MAA, Moniruzzaman M, Nasiruddin KM (2012) Agrobacterium-mediated Genetic transformation of Potato (Solanum tuberosum L.) var. Cardinal and Heera. The Agriculturists 10:81–86.

  42. Kumar V, Luthra SK, Bhardwaj V, Singh BP (2014) Indian potato varieties and their salient features. Technical bulletin no. 78. ICAR-Central Potato Research Institute, Shimla, India

  43. Kurtz DI (1974) Fidelity of protein synthesis with chicken embryo mitochondrial and cytoplasmic ribosomes. Biochemistry 13:572–577.

  44. Li S, Zhen C, Xu W, Wang C, Cheng Y (2017) Simple, rapid and efficient transformation of genotype Nisqually-1: a basic tool for the first sequenced model tree. Sci Rep 7:2638.

  45. Lopez SJ, Kumar RR, Pius PK, Muraleedharan N (2014) Agrobacterium tumefaciens-mediated genetic transformation in Tea (Camellia sinensis [L.] O. Kuntze). Plant Mol Biol Rep 22:201a–201j.

  46. Maheswaran GM, Welander M, Hutchinson JF, Graham MW, Richards D (1992) Transformation of apple rootstock M26 with Agrobacterium tumefaciens. J Plant Physiol 139:560–568.

  47. Millam S (2004) Agrobacterium-mediated transformation of potato. In: Curtis I (ed) Transgenic crops of the world. Springer, Dordrecht, pp 257–269.

  48. Molla MM, Nasiruddin KM, Al-Amin M et al (2011) Effect of growth regulators on direct regeneration of potato. Int Proc Chem Biol Environ Eng 12:205–210

  49. Murashige T, Skoog F (1962) A revised medium for rapid growth and bio assays with tobacco tissue cultures. Physiol Plant 15:473–497.

  50. Niu X, Li X, Veronese P, Bressan RA, Hasegawa SC, Weller PM (2000) Factors affecting Agrobacterium tumefaciens-mediated transformation of pepper mint. Plant Cell Rep 19:304–310.

  51. Padilla IM, Burgos L (2010) Aminoglycoside antibiotics: structure, functions and effects on in vitro plant culture and genetic transformation protocols. Plant Cell Rep 29:1203–1213.

  52. Rakosy-Tican E, Aurori CM, Dijkstra C, Thieme R, Aurori A, Davey MR (2007) The usefulness of the gfp reporter gene for monitoring Agrobacterium-mediated transformation of potato dihaploid and tetraploid genotypes. Plant Cell Rep 26:661–671.

  53. Rana RK, Pandit A, Pandey SK (2009) Profitability analysis of Kufri Chipsona-1 cultivation in Uttar Pradesh of India. Potato J 36:166–172

  54. Sadawarti M, Patel K, Samadhiya RK, Gupta PK, Singh SP, Gupta VK, Roy S, Chakrabarti SK, Verma D (2018) Evaluation of table and processing varieties of potato (Solanum tuberosum L.) FOR North-Central India. Int J Chem Stud 6:823–833

  55. Sawahel WA (2002) The production of transgenic potato plants expressing human alpha interferon using lipofectin-mediated transformation. Cell Mol Biol Lett 7:19–29

  56. Shin DY, Seong ES, Na JK, Yoo JH, Kang WH, Ghimire BK et al (2011) Conditions for regeneration and transformation of Solanum tuberosum cultivars using the cotton glutathione S-transferase (Gh-5) gene. Afr J Biotechnol 10:15135–15141.

  57. Si HJ, Xie CH, Liu J (2003) An efficient protocol for Agrobacterium mediated transformation with microtuber and the introduction of an antisense class I gene into potato. Acta Agron Sin 29:801–805

  58. Sidorov VA, Kasten D, Pang S, Hajdukiewicz PTJ, Staub JM, Nehra NS (1999) Stable chloroplast transformation in potato: use of green fluorescent protein as a plastid marker. Plant J 19:209–216.

  59. Snyder GW, Belknap WR (1993) A modified method for routine Agrobacterium mediated transformation of in vitro grown potato microtubers. Plant Cell Rep 12:324–327.

  60. Stiekema WJ, Heidekamp F, Louwerse JD, Verhoeven HA, Dijkhuis P (1988) Introduction of foreign genes into potato cultivars Bintje and Desiree using an Agrobacterium tumefaciens binary vector. Plant Cell Rep 7:47–50.

  61. Subramoni S, Nathoo N, Klimov E, Yuan ZC (2014) Agrobacterium tumefaciens responses to plant-derived signaling molecules. Front Plant Sci 5:322.

  62. Tang X, Zhang N, Si H, Calderon-Urrea A (2017) Selection and validation of reference genes for RT-qPCR analysis in potato under abiotic stress. Plant Methods 13:85–93.

  63. Terakami S, Matsuta N, Yamamoto T, Sugaya S, Gemma H, Soejima J (2007) Agrobacterium-mediated transformation of the dwarf pomegranate (Punica granatum L.). Plant Cell Rep 26:1243–1251.

  64. Tournier V, Grat S, Marque C, El Kayal W, Penchel R, de Andrade G et al (2003) An efficient procedure to stably introduce genes into an economically important pulp tree (Eucalyptus grandis X Eucalyptus urophylla). Transgenic Res 12:403–411

  65. Trujillo C, Rodriguez-Arango E, Jaramillo S, Hoyos R, Orduz S, Arango R (2001) One-step transformation of two Andean potato cultivars (Solanum tuberosum L. subsp. andigena). Plant Cell Rep 20:637–641.

  66. Valkov VT, Gargano D, Manna C, Formisano G, Dix PJ, Gray JC et al (2011) High efficiency plastid transformation in potato and regulation of transgene expression in leaves and tubers by alternative 5′ and 3′ regulatory sequences. Transgenic Res 20:137–151.

  67. Veale MA, Slabbert MM, Emmenes LV (2012) Agrobacterium-mediated transformation of potato cv. Mnandi for resistance to the potato tuber moth (Phthorimaea operculella). S Afr J Bot 80:67–74.

  68. Visser RGF, Jacobsen E, Hesseling-Meinders A, Schans MJ, Witholt B, Feenstra WJ (1989) Transformation of homozygous diploid potato with an Agrobacterium tumefaciens binary vector system by adventitious shoot regeneration on leaf and stem segments. Plant Mol Biol 12:329–337.

  69. Volkov RA, Komarova NY, Panchuk II, Hemleben V (2003) Molecular evolution of rDNA external transcribed spacer and phylogeny of sect. Petota (genus Solanum). Mol Phylogenet Evol 29:187–202.

  70. Weisburg WA, Barns SM, Pelletier DA, Lane DJ (1991) 16S rDNA amplification for phylogenetic study. J Bacteriol 173:697–703.

  71. Wendt T, Doohan F, Winckelmann D, Mullins E (2011) Gene transfer into Solanum tuberosum via Rhizobium spp. Transgenic Res 20:377–386.

  72. Wendt T, Doohan F, Mullins E (2012) Production of Phytophthora infestans-resistant potato (Solanum tuberosum) utilising Ensifer adhaerens OV14. Transgenic Res 21:567–578.

  73. Yasmeen A (2009) An improved protocol for the regeneration and transformation of tomato (cv. Rio Grande). Acta Physiol Plant 31:1271–1277.

  74. Yevtushenko DP, Misra S (2010) Efficient Agrobacterium-mediated transformation of commercial hybrid poplar Populus nigra L. X P. maximowiczii A. Henry. Plant Cell Rep 29:211–229.

  75. Zambre M, Terryn N, Clercq JD, De Buck S, Dillen W, Van Montagu M et al (2003) Light strongly promotes gene transfer from Agrobacterium tumefaciens to plant cells. Planta 216:580–586.

  76. Zhong G (2007) Establishment of high efficient regenration system of Lilium and Agrobacterium-mediated genetic transformation. Ph.D. thesis, Southwest University Press

  77. Zuker A, Ahroni A, Tzfira T, Ben-Meir H, Vainstein A (1999) Wounding by bombardment yields highly yields highly efficient Agrobacterium-mediated transformation of carnation (Dianthus caryophyllus L.). Mol Breed 5:367–375.

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Kaur, A., Guleria, S., Reddy, M.S. et al. A robust genetic transformation protocol to obtain transgenic shoots of Solanum tuberosum L. cultivar ‘Kufri Chipsona 1’. Physiol Mol Biol Plants (2020) doi:10.1007/s12298-019-00747-4

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  • Agrobacterium tumefaciens
  • Pre-culture period
  • Co-cultivation
  • Injury
  • Chimeric shoots