In vitro cultures and regeneration of Bienertia sinuspersici (Chenopodiaceae) under increasing concentrations of sodium chloride and carbon dioxide

Abstract

To study the developmental transition of chloroplasts from C3 to C4 photosynthesis in the terrestrial single-cell C4 species Bienertia sinuspersici, a regeneration protocol was developed. Stem explant material developed callus either with or without red nodular structures (RNS) when cultured on Murashige–Skoog (MS) salts and vitamins, supplemented with 5 mM phosphate, plus 1 mg L−1 dichloropenoxy-acetic acid (2,4-D), and 87 mM sucrose (Stage 1 media). Only calli having RNS were able to regenerate plantlets. MS media plus phosphate was used throughout regeneration, with the Stage 2 media containing 2 mg L−1 6-benzylaminopurine, 43 mM sucrose and 1.5% soluble starch. Stage 3 media had no hormones or organic sources of carbon, and cultures were grown under ambient (~400 ppm) versus CO2 enrichment (1.2% CO2). When calli without RNS were cultured under Stage 3 conditions with 1.2% CO2, there was an increase in growth, protein content, and photosystem II yield, while structural and biochemical analyses indicated the cells in the calli had C3 type photosynthesis. CO2 enrichment during growth of RNS during Stage 3 had a large effect on regeneration success, increasing efficiency of shoot and root development, size of plantlets, leaf soluble protein, and chlorophyll concentration. Anatomical analysis of plantlets, which developed under 1.2% CO2, showed leaves developed C4 type chlorenchyma cells, including expression of key C4 biochemical enzymes. Increasing salinity in the media, from 0 to 200 mM NaCl, increased tissue osmolality, average plantlet area and regeneration success, but did not affect protein or chlorophyll content.

This is a preview of subscription content, log in to check access.

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5

References

  1. Akhani H, Barroca J, Koteyeva N, Voznesenskaya E, Franceschi V, Edwards G, Ghaffari SM, Ziegler H (2005) Bienertia sinuspersici (Chenopodiaceae): a new species from southwest Asia and discovery of a third terrestrial C4 plant without Kranz anatomy. Syst Bot 30:290–301

    Article  Google Scholar 

  2. Aragon CE, Escalona M, Rodrigues R, Canal MJ, Capote I, Pina D, Gonzalez-Olmedo J (2010) Effect of sucrose, light, and carbon dioxide on plantain micropropagation in temporary immersion bioreactors. In Vitro Cell Dev Biol-Plant 46:89–94

    Article  CAS  Google Scholar 

  3. Arnold V, Fletcher RA (1986) Stimulation of chlorophyll synthesis by benzyladenine and potassium in excised and intact cucumber cotyledons. Physiol Plant 68:169–174

    Article  CAS  Google Scholar 

  4. Bajji M, Kinet JM, Lutts S (1998) Salt stress effects on roots and leaves of Atriplex halimus L. and their corresponding callus cultures. Plant Sci 137:131–142

    Article  CAS  Google Scholar 

  5. Berlyn MB, Zelitch I (1975) Photoautotrophic growth and photosynthesis in tobacco callus cells. Plant Physiol 56:752–756

    PubMed  Article  CAS  Google Scholar 

  6. Boyd CN, Franceschi VR, Chuong SDX, Akhani H, Kiirats O, Smith M, Edwards GE (2006) Flowers of Bienertia cycloptera and Suaeda aralocaspica (Chenopodiaceae) complete their life cycle performing single-cell C4 photosynthesis. Funct Plant Biol 34:268–281

    Article  Google Scholar 

  7. Bradford MM (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72:248–254

    PubMed  Article  CAS  Google Scholar 

  8. Burger J, Edwards GE (1996) Photosynthetic efficiency, and photodamage by UV and visible radiation, in red versus green leaf Coleus varieties. Plant Cell Physiol 37:395–399

    Google Scholar 

  9. Chen Y, Dribnenki P (2004) Effect of medium osmotic potential on callus induction and shoot regeneration in flax anther culture. Plant Cell Rep 23:272–276

    PubMed  Article  CAS  Google Scholar 

  10. Chu CC, Wang CS, Sun CS, Hus C, Ying KC, Chu CY, Bi FY (1975) Establishment of an efficient medium for anther culture of rice through comparative experiments on the nitrogen sources. Sci Sin 18:659–668

    Google Scholar 

  11. Chuong SDX, Franceschi VR, Edwards GE (2006) The cytoskeleton maintains organelle partitioning required for single-cell C4 photosynthesis in Chenopodiaceae species. Plant Cell 18:2207–2223

    PubMed  Article  CAS  Google Scholar 

  12. Colombo SL, Andreo CS, Chollet R (1998) The interaction of shikimic acid and protein phosphorylation with PEP carboxylase from the C4 dicot Amaranthus viridis. Phytochem 48:55–59

    Article  CAS  Google Scholar 

  13. D’Halluin K, Bossut M, Bonne E, Mazur B, Leemans J, Botterman J (1992) Transformation of sugarbeet (Beta vulgaris I.) and evaluation of herbicide resistance in transgenic plants. Nat Biotech 10:309–314

    Article  Google Scholar 

  14. Dolezal K, Astot C, Hanus J, Holub J, Peters W, Beck E, Strnad M, Sandberg G (2001) Identification of aromatic cytokinins in suspension cultured photoautotrophic cells of Chenopodium rubrum by capillary liquid chromatography/frit-fast atom bombardment mass spectrometry. Plant Growth Regul 36:181–189

    Article  Google Scholar 

  15. Duff SMG, Moorhead GBG, Lefebvre DD, Plaxton WC (1989) Phosphate starvation inducible ‘bypasses’ of adenylate and phosphate dependent glycolytic enzymes in Brassica nigra suspension cells. Plant Physiol 90:1275–1278

    PubMed  Article  CAS  Google Scholar 

  16. Edwards GE, Franceschi VR, Voznesenskaya EV (2004) Single-cell C4 photosynthesis versus the dual-cell (Kranz) paradigm. Ann Rev Plant Biol 55:173–196

    Article  CAS  Google Scholar 

  17. Eisa S, Koyro HW, Kogel KH, Imani J (2005) Induction of somatic embryogenesis in cultured cells of Chenopodium quinoa. Plant Cell Tiss Organ Cult 81:243–246

    Article  Google Scholar 

  18. Flowers TJ, Colmer TD (2008) Tansley review: salinity tolerance in halophytes. New Phytol 179:945–963

    PubMed  Article  CAS  Google Scholar 

  19. Geekiyanage S, Takase T, Watanabe S, Fukai S, Kiyosue T (2006) The combined effect of photoperiod, light intensity and GA3 on adventitious shoot regeneration from cotyledons of spinach (Spinacia oleracea L.). Plant Biotech 23:431–435

    Article  CAS  Google Scholar 

  20. von Hedenstrom H, Breckle SW (1974) Obligate halophytes? A test with tissue culture methods. Z Pflanzenphysiol 74:183–185

    Google Scholar 

  21. Husemann W, Barz W (1977) Photoautotrophic growth and photosynthesis in cell suspension cultures of Chenopodium rubrum. Physiol Plant 40:77–81

    Article  Google Scholar 

  22. Jang JC, Sheen J (1994) Sugar sensing in higher plants. Plant Cell 6:1665–1679

    PubMed  Article  CAS  Google Scholar 

  23. Kanai R, Edwards GE (1999) The biochemistry of C4 photosynthesis. In: Sage RF, Monson RK (eds) C4 plant biology. Academic Press, San Diego, pp 49–80

    Google Scholar 

  24. Kennedy RA, Barnes JE, Laetsch WM (1977) Photosynthesis in C4 plant tissue cultures: significance of Kranz anatomy to C4 acid metabolism in C4 plants. Plant Physiol 59:600–603

    PubMed  Article  CAS  Google Scholar 

  25. Kinoshita I, Tsuji H (1984) Benzyladenine-induced increase in DNA content per chloroplast in intact bean leaves. Plant Physiol 76:575–578

    PubMed  Article  CAS  Google Scholar 

  26. Knudson LL, Tibbitts TW, Edwards GE (1977) Measurement of ozone injury by determination of leaf chlorophyll concentration. Plant Physiol 60:606–608

    PubMed  Article  CAS  Google Scholar 

  27. Komai F, Okuse I, Harada T (1996) Somatic embryogenesis and plant regeneration in culture of root segments of spinach (Spinacia oleracea L.). Plant Sci 113:203–208

    Article  CAS  Google Scholar 

  28. Krapp A, Hofmann B, Schafer C, Stitt M (1993) Regulation of the expression of rbcS and other photosynthetic genes by carbohydrates: a mechanism for the ‘sink regulation’ of photosynthesis? Plant J 3:817–828

    Article  CAS  Google Scholar 

  29. Krause GH, Weis E (1991) Chlorophyll fluorescence and photosynthesis: the basics. Ann Rev Plant Physiol Plant Mol Biol 42:313–349

    Article  CAS  Google Scholar 

  30. Laemmli UK (1970) Cleavage of structural proteins during the assembly of the head of Bacteriophage T4. Nature 227:680–686

    PubMed  Article  CAS  Google Scholar 

  31. Laetsch WM (1974) The C4 syndrome: a structural analysis. Ann Rev Plant Physiol 25:27–52

    Article  CAS  Google Scholar 

  32. Laetsch WM, Kortschak HP (1972) Chloroplast structure and function in tissue cultures of a C4 plant. Plant Physiol 49:1021–1023

    PubMed  Article  CAS  Google Scholar 

  33. Lara MV, Offermann S, Smith M, Okita TW, Andreo CS, Edwards GE (2008) Leaf development in the single-cell C4 system in Bienertia sinuspersici: expression of genes and peptide levels for C4 metabolism in relation to chlorenchyma structure under different light conditions. Plant Physiol 148:593–610

    PubMed  Article  CAS  Google Scholar 

  34. Lavergne D, Nato A, Dupuis JM, Pean M, Chagvardieff P (1992) Evidence for the expression of morphological and biochemical characteristics of C3-photosynthesis in chlorophyllous callus cultures of Zea mays. Physiol Plant 84:292–300

    Article  CAS  Google Scholar 

  35. Leisner CP, Cousins AB, Offermann S, Okita TW, Edwards GE (2010) The effects of salinity of photosynthesis and growth of the single-cell C4 species Bienertia sinuspersici (Chenopodiaceae). Photosynth Res 106:201–214

    PubMed  Article  CAS  Google Scholar 

  36. Long JJ, Wang JL, Berry JO (1994) Cloning and analysis of the C4 photosynthetic NAD-dependent malic enzyme of Amaranth mitochondria. J Biol Chem 269:2827–2833

    PubMed  CAS  Google Scholar 

  37. Milivojevic S, Mitrovic A, Culafic L (2005) Somatic embryogenesis in Chenopodium rubrum and Chenopodium murale in vitro. Biol Plant 49:35–39

    Article  Google Scholar 

  38. Moghaieb REA, Saneoka H, Fujita K (2004) Effect of salinity on osmotic adjustment, glycinebetaine accumulation and the betaine aldehyde dehydrogenase gene expression in two halophytic plants, Salicornia europaea and Suaeda maritima. Plant Sci 166:1345–1349

    Article  CAS  Google Scholar 

  39. Mol J, Jenkins G, Schafer E, Weiss D (1996) Signal perception, transduction, and gene expression involved in anthocyanin biosynthesis. Crit Rev Plant Sci 15:525–557

    CAS  Google Scholar 

  40. Mudalige RG, Longstreth DJ (2006) Effects of salinity on photosynthetic characteristics in photomixotrophic cell-suspension cultures from Alternanthera philoxeroides. Plant Cell Tiss Organ Cult 84:301–308

    Article  CAS  Google Scholar 

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

    Article  CAS  Google Scholar 

  42. Murota K, Ohshita Y, Watanabe A, Aso S, Sato F, Yamada Y (1994) Changes related to salt tolerance in thylakoid membranes of photoautotrophically cultured green tobacco cells. Plant Cell Physiol 35:107–113

    CAS  Google Scholar 

  43. Nguyen QT, Kozai T (1998) Environmental effects of the growth of plantlets in micropropagation. Environ Control Biol 36:59–75

    Google Scholar 

  44. Nguyen QT, Kozai T (2001) Growth of in vitro banana (Musa spp.) shoots under photomixotrophic and photoautotrophic conditions. In Vitro Cell Dev Biol Plant 37:824–829

    Article  Google Scholar 

  45. Offermann S, Okita TW, Edwards GE (2011) Resolving the compartmentation and function of C4 photosynthesis in the single-cell C4 species Bienertia sinuspersici. Plant Physiol 155:1612–1628

    Google Scholar 

  46. Park J, Okita TW, Edwards GE (2009a) Salt tolerant mechanisms in single-cell C4 species Bienertia sinuspersici and Suaeda aralocaspica (Chenopodiaceae). Plant Sci 176:616–626

    Article  CAS  Google Scholar 

  47. Park J, Knoblauch M, Okita TW, Edwards GE (2009b) Structural changes in the vacuole and cytoskeleton are key to development of the two cytoplasmic domains supporting single-cell C4 photosynthesis in Bienertia sinuspersici. Planta 229:369–382

    PubMed  Article  CAS  Google Scholar 

  48. Rogers SMD, Ogren WL, Widholm JM (1987) Photosynthetic characteristics of a photoautotrophic cell suspension culture of soybean. Plant Physiol 84:1451–1456

    PubMed  Article  CAS  Google Scholar 

  49. Seeni S, Gnanam A (1983) Photosynthesis in cell suspension culture of a C4 plant, Gisekia pharnaceoides L. Plant Cell Physiol 24:1033–1041

    CAS  Google Scholar 

  50. Sha Valli Khan PS, Kozai T, Nguyen QT, Kubota C, Dhawan V (2002) Growth and net photosynthetic rates of Eucalyptus tereticornis Smith under photomixotrophic and various photoautotrophic micropropagation conditions. Plant Cell Tiss Organ Cult 71:141–146

    Article  CAS  Google Scholar 

  51. Sha Valli Khan PS, Kozai T, Nguyen QT, Kubota C, Dhawan V (2003) Growth and water relations of Paulownia fortunei under photomixotrophic and photoautotrophic conditions. Biol Plant 46:161–166

    Article  Google Scholar 

  52. Shi XL, Han HP, Shi WL, Li YX (2006) NaCl and TDZ are two key factor for the improvement of in vitro regeneration rate of Salicornia europaea L. J Integr Plant Biol 48:1185–1189

    Article  CAS  Google Scholar 

  53. Singh A, Chikara J, Pandya JB (2004) Plant regeneration from callus cultures in Suaeda nudiflora (Wild.) Moq. Ind J Biotech 3:454–456

    CAS  Google Scholar 

  54. Smith MK, McComb JA (1981) Effect of NaCl on the growth of whole plants and their corresponding callus cultures. Aust J Plant Physiol 8:267–275

    Article  CAS  Google Scholar 

  55. Stefaniak B, Wozny A, Li V (2003) Plant micropropagation and callus induction of some annual Salsola species. Biol Plant 46:305–308

    Article  Google Scholar 

  56. Uchida A, Nagamiya K, Takabe T (2003) Transformation of Atriplex gmelini plants from callus lines using Agrobacterium tumefaciens. Plant Cell Tiss Organ Cult 75:151–157

    Article  CAS  Google Scholar 

  57. Usuda H, Kanai R, Takeuchi M (1971) Comparison of carbon dioxide fixation and the fine structure in various assimilatory tissues of Amaranthus retroflexus L. Plant Cell Physiol 12:917–930

    CAS  Google Scholar 

  58. Voznesenskaya EV, Koteyeva NK, Chuong SDX, Akhani H, Edwards GE, Franceschi VR (2005) Differentiation of cellular and biochemical features of the single-cell C4 syndrome during leaf development in Bienertia cycloptera (Chenopodiaceae). Am J Bot 91:1784–1795

    Article  Google Scholar 

  59. Wu Z, Chen LJ, Long YJ (2009) Analysis of ultrastructure and reactive oxygen species of hyperhydric garlic (Allium sativum L.) shoots. In Vitro Cell Dev Biol Plant 45:483–490

    Google Scholar 

  60. Xu C, Blair LC, Rogers SMD, Govindjee, Widholm JM (1988) Characteristics of five new photoautotrophic suspension cultures including two Amaranthus species and a cotton strain growing on ambient CO2 levels. Plant Physiol 88:1297–1302

  61. Yen HE, Grimes HD, Edwards GE (1995) The effects of high salinity, water-deficit, and abscisic acid on phosphoenolpyruvate carboxylase activity and proline accumulation in Mesembryanthemum crystallinum cell - cultures. J Plant Physiol 145:557–564

    CAS  Google Scholar 

  62. Yen HE, Chen YC, Yen SK, Lin JH (1999) Sugar uptake by photomixotrophic soybean cultures. Bot Bull Acad Sin 40:147–152

    CAS  Google Scholar 

  63. Zhao SZ, Ruan Y, Sun HZ, Wang BS (2008) Highly efficient Agrobacterium-based transformation system for callus cells of the C3 halophyte Suaeda salsa. Acta Physiol Plant 30:729–736

    Article  Google Scholar 

  64. Zhao SZ, Sun HZ, Chen M, Wang BS (2010) Light-regulated betacyanin accumulation in euhalophyte Suaeda salsa calli. Plant Cell Tissue Organ Cult 102:99–107

    Article  CAS  Google Scholar 

  65. Ziegler P, Scheibe R (1989) Greening and growth of suspension-cultured cells of Chenopodium rubrum under conditions of heterotrophic and autotrophic nutrition. Plant Cell Environ 12:725–735

    Article  CAS  Google Scholar 

Download references

Acknowledgments

This material is based upon work supported by the National Science Foundation under Grant IBN-0641232. Seeds of Bienertia sinuspersici were kindly provided by Dr. Abdulrahman Alsirhan, Kuwait. We thank the WSU Plant Transformation Center, the Franceschi Microscopy and Imaging Center of Washington State University for use of facilities and for staff assistance, C. Cody for plant growth management, and N. Koteyeva for assistance with graphics.

Author information

Affiliations

Authors

Corresponding author

Correspondence to Gerald E. Edwards.

Additional information

Communicated by F. Sato.

Electronic supplementary material

Below is the link to the electronic supplementary material.

Supplementary material 1 (DOC 1218 kb)

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Rosnow, J., Offermann, S., Park, J. et al. In vitro cultures and regeneration of Bienertia sinuspersici (Chenopodiaceae) under increasing concentrations of sodium chloride and carbon dioxide. Plant Cell Rep 30, 1541–1553 (2011). https://doi.org/10.1007/s00299-011-1067-1

Download citation

Keywords

  • Callus
  • C4 photosynthesis
  • CO2 enrichment
  • Halophyte
  • Regeneration