Abstrakt
Regeneracja roślin poprzez pośrednią organogenezę pozwala na otrzymanie zróżnicowania genetycznego, które może zostać wykorzystane w kreowaniu nowych odmian. W pracy przedstawiono szybki i efektywny protokół jednoetapowego otrzymywania regenerantów stewii (Stevia rebaudiana Bertoni). Do otrzymania tkanki kalusowej oraz regeneracji pędów, jako eksplantaty pierwotne wykorzystano liście i międzywęźla, które wyłożono na pożywkę MS (Murashige and Skoog) wzbogaconą roślinnymi regulatorami wzrostu (PGR’s) NAA (1-Naphthaleneacetic acid) (2.0 mg/l), BA (6-Benzylaminopurine) (4.0 mg/l), 2,4-D (2,4-Dichlorophenoxyacetic) (2.0 mg/l). Tkanka kalusowa powstała na obu typach eksplantatów, jednakże ta pochodząca z na międzywęźlach była jedynie proliferująca. Z eksplantatów liściowych na zastosowanym podłożu już po 6 tygodniach trwania kultury uzyskano średnio 3.92 pędy z eksplantatu. Analizę zdolności morfogenetycznych otrzymanych regenerantów przeprowadzono na podłożu MS wzbogaconym o PGR’s - kinetin (0.25 mg/l) and BA (6-Benzylaminopurine) (0.5 mg/l). Ocena średniej liczby pędów, średniej długości pędów (cm) i średniej liczby węzłów na eksplantat wskazuje na zróżnicowanie fenotypowe otrzymanych regenerantów. Zastosowanie markerów RAPD (Randomly Amplified Polymorphic DNA) potwierdziło istnienie różnic także na poziomie DNA. Zaproponowany jednoetapowy, protokół regeneracji drogą pośredniej organogenezy indukuje zmienność somaklonalną Stevia rebaudiana Bertoni a uzyskane regeneranty po selekcji mogą być wykorzystane w hodowli twórczej tego gatunku.
Bibliografia
- Al-Taweel S.K., Azzam C.R., Khaled K.A., Abdel-Aziz R.M., 2021. Improvement of stevia (Stevia rebaudiana Bertoni) and steviol glycoside through traditional breeding and bio-technological approaches. SABRAO J. Breed. Genet. 53(1), 88–111.
- Asthana P., Jaiswal V.S., Jaiswal U., 2011. Micropropagation of Sapindustrifoliatus L. and assessment of genetic fidelity of micropropagated plants using RAPD analysis. Acta Phys-iol. Plant. 33, 1821–1829. https://doi.org/10.1007/s11738-011-0721-0
- Bairu M.W., Aremu A.O., Van Staden J., 2011. Somaclonal variation in plants: causes and detection methods. Plant Growth Regul. 63, 147–173. https://doi.org/10.1007/s10725-010-9554-x
- Cao X., Hammerschlag F.A., 2000. Improved shoot organogenesis from leaf explants of high-bush blueberry. HortScience 35(5), 945–947. https://doi.org/10.21273/HORTSCI.35.5.945
- Chacón-Morales P., Amaro-Luis J.M., Bahsas A., 2013. Isolation and characterization of (+)-mellein, the first isocoumarin reported in Stevia genus. Av. en Quim. 8, 145–151.
- Chester K., Tamboli E.T., Parveen R., Ahmad S., 2013. Genetic and metabolic diversity in Stevia rebaudiana using RAPD and HPTLC analysis. Pharm. Biol. 51, 771–777. https://doi.org/10.3109/13880209.2013.765898
- Doliński R., Jabłońska E., 2015. Mikrorozmnażanie stewii (Stevia rebaudiana Bert.) z eksplantatów węzłowych izolowanych z roślin wytworzonych in vitro. Agron. Sci. 70(4), 13–24.
- Doyle J.J., Doyle J.L., 1987. A rapid DNA isolation procedure for small quantities of fresh leaf tissue. Phytochem. Bull. 9, 11–15.
- Dyduch-Siemińska M., Najda A., Gawroński J., Balant S., Świca K., Żaba A., 2020. Stevia rebaudiana Bertoni, a source of high-potency natural sweetener-biochemical and genetic characterization. Molecules 25(4), 767. https://doi.org/10.3390/molecules25040767
- El-Zaidy M., Zayed M., Alharbi S., Doaigey A., Al Sahli A., Ammar R., 2014. Micropropaga-tion of seven Stevia rebaudiana Bert. genotypes via adult leaf explants. J. Pure Appl. Mi-crobiol. 8(2),1289–1298.
- Esmaeili F., Kahrizi D., Mansouri M., Yari K., Kazemi N., Ghaheri M., 2016. Cell dedifferentiation in Stevia rebauiana as a pharmaceutical and medicinal plant. J. Rep. Pharm. Sci. 5, 12–17.
- Ferrazzano G.F., Cantile T., Alcidi B., Coda M., Ingenito A., Zarrelli A., Di Fabio G., Pollio A., 2016. Is Stevia rebaudiana Bertoni a non cariogenic sweetener?. Molecules 21(1), 1–12. https://doi.org/10.3390/molecules21010038
- Gantait S., Das A., Banerjee J., 2018. Geographical distribution, botanical description and self-incompatibility mechanism of genus Stevia. Sugar Tech. 20, 1–9. https://doi.org/10.1007/s12355-017-0563-1
- Gantait S., Das A., Mandal N., 2015. Stevia: A comprehensive review on ethnopharmacological properties and in vitro regeneration. Sugar Tech. 17, 95–106. https://doi.org/10.1007/s12355-014-0316-3
- Grafi G., Barak S., 2014. Stress induces cell dedifferentiation in plants. Biochim. Biophys. Acta. 1849, 378–384. https://doi.org/10.1016/j.bbagrm.2014.07.015
- Gunasena M.D.K.M., Senarath W.T.P.S.K., 2019. In vitro plant regeneration of Stevia rebaudiana through indirect organogenesis. Int. J. Bot. Stud. 4(4), 199–203.
- Hammer Ø., Harper D.A.T., Ryan P.D., 2001. Past: paleontological statistics software package for education and data analysis. Palaeontol. Elect. 4, 9.
- Hassanen S.A., Khalil R.M.A., 2013. Biotechnological studies for improving of stevia (Stevia rebaudiana Bertoni) in vitro plantlets. Middle East J. Sci. Res. 14 (1), 93–106. https://doi.org/10.5829/idosi.mejsr.2013.14.1.1948
- Heinz D.J., Mee G.W.P., 1971. Morphologic, cytogenetic, and enzymatic variation in Saccharum species hybrid clones derived from callus tissue. Am. J. Bot. 58, 257–262. https://doi.org/10.1002/j.1537-2197.1971.tb09971.x
- Jaccard P., 1908. Nouvellesrechearches sur la distribution locale. Bull. Soc. Vaudoise Sci. Nat. 44, 223–270.
- Jain S.M., 2001. Tissue culture-derived variation in crop improvement. Euphytica 118, 153–166. https://doi.org/10.1023/A:1004124519479
- Janarthanam B., Gopalakrishnan M., Lakshmi Sai G., Sekar T., 2009. Plant regeneration from leaf derived callus of Stevia rebaudiana Bertoni. Plant Tissue Cult. Biotech. 19(2), 133–141. https://doi.org/10.3329/ptcb.v19i2.5430
- Javed R., Yucesan B., Zia M., Ekrem G., 2018. Elicitation of secondary metabolites in callus cultures of Stevia rebaudiana Bertoni grown under ZnO and CuO nanoparticles stress. Sugar Tech. 20(2), 194–201. https://doi.org/10.1007/s12355-017-0539-1
- Khan A., Jayanthi M., Gantasala N.P., Bhooshan N., Rao U., 2016. A rapid and efficient protocol for in vitro multiplication of genetically uniform Stevia rebaudiana (Bertoni). Indian J. Exp. Biol. 54, 477–481.
- Krishna H., Alizadeh M., Singh D., Singh U., Chauhan N., Eftekhari M., Sadh R.K., 2016. Somaclonal variations and their applications in horticultural crops improvement. Biotech. 6, 1–18. https://doi.org/10.1007/s13205-016-0389-7
- Majumder S., Rahman S.D., 2016. Micropropagation of Stevia rebaudiana Bertoni through direct and indirect organogenesis. J. Innov. Pharmac. Biol. Sci. 13(3), 47–56.
- Masri M.I., Amein M.M.M., Ranya M., Aziz A., Sayed D.O., 2019. Callogenesis and plant regeneration via in vitro culture of stevia rebaudiana explants. Egypt. J. Plant Breed. 23(1), 65–76.
- Modi A.R., Patil G., Kumar N., Singh A.S., Subhash N.A., 2012. Simple and efficient in vitro mass multiplication procedure for Stevia rebaudiana Bertoni and analysis of genetic fideli-ty of in vitro raised plants through RAPD. Sugar Tech. 14, 391–397. https://doi.org/10.1007/s12355-012-0169-6
- Moktaduzzaman M., Mahbubur-Rahman S.M., 2009. Regeneration of Stevia rebaudiana and analysis of somaclonal variation by RAPD. Biotechnology 8(4), 449–455. https://doi.org/10.3923/biotech.2009.449.455
- Mubarak M.H., Belal A.H., EL-Geddawy I.H., Eman I., Sarag E.L., Nasr M.I., 2008. Micropropagation of Stevia rebaudiana in vitro. Meeting the challenges of sugar crops and integrated industries in developing countries. Al Arish, Egypt, 293–298.
- Murashige T., Skoog F.A., 1962. Revised medium for rapid growth and bioassays with tobacco tissue culture. Physiol. Plant. 15, 473–497.
- Patel R. M., Shah R.R., 2009. Regeneration of stevia plant through callus culture. Indian. J. Pharm. Sci. 71(1), 46–50. https://doi.org/ 10.4103/0250-474X.51954
- Predieri S., 2001. Mutation induction and tissue culture in improving fruits. Plant Cell. Tissue Organ Cult. 64, 185–210. http://dx.doi.org/10.1023/A:1010623203554
- Preethi D., Sridhar T.M., Naidu C.V., 2011. Efficient protocol for indirect shoot regeneration from leaf explants of Stevia rebaudiana (Bert.) – an important calorie free biosweetner. J. Phytol. 3(5), 56–60.
- Razak U.N.A.A., Ong C.B., Yu T.S., Lau L.K., 2014. In vitro micropropagation of Stevia rebaudiana Bertoni in Malaysia. Braz. Arch. Biol. Technol. 1, 23–28. https://doi.org/10.1590/S1516-89132014000100004
- Samuel P., Ayoob K.T., Magnuson B.A., Wölwer-Rieck U., Jeppesen P.B., Rogers P.J., Row-land I., Mathews R., 2018. Stevia leaf to stevia sweetener: exploring its science, benefits, and future potential. J. Nutr. 148, 1186S–1205S. https://doi.org/10.1093/jn/nxy102
- Sarmah D., Sutradhar M., Singh B.K., 2017. Somaclonal variation and its’ application in ornamentals plants. Int. J. Pure Appl. Biosci. 5, 396–406. https://doi.org/10.18782/2320-7051.2762
- Shah S.H., Ali S., Jan S.A., Din J., Ali G.M., 2015. Callus induction, in vitro shoot regeneration and hairy root formation by the assessment of various plant growth regulators in tomato (Solanumlycopersicum Mill.). J. Anim. Plant Sci. 25, 528–538.
- Sharma N., Gauchan D.P., Dhakal A., Luitel A., Shakya S., Shakya R., 2015. Establishment of regenerative callus, cell suspension system and molecular characterization of Stevia rebaudiana Bertoni for the production of stevioside in in vitro. Int. J. Res. Appl. Sci. Eng. Technol. 3, 133–144.
- Sharma N., Kaur R., Era V., 2016. Potential of RAPD and ISSR markers for assessing genetic diversity among Stevia rebaudiana Bertoni accessions. Indian J. Biotechnol. 15, 95–100.
- Sikdar S.U., Zobayer N., Azim F., Ashrafuzzaman M., Prodhan S.H., 2012. An efficient callus initiation and direct regeneration of Stevia rebaudiana. Afr. 11(45), 10381–10387. https://doi.org/10.5897/AJB11.2363
- Singh M., Saharan V., Rajpurohit D., Sen Y., Joshi A., Sharma A., 2017. Thidiazuron induced direct shoot organogenesis in Stevia rebaudiana and assessment of clonal fidelity of regenerated plants by RAPD and ISSR. Int. J. Curr. Microbiol. Appl. Sci. 6, 1690–1702. https://doi.org/10.20546/ijcmas.2017.608.203D.
- Skirvin R.M., Norton M., McPheeters K.D., 1993. Somaclonal variation: has it proved useful for plant improvement. Acta Hortic. 336, 333–340.
- Soliman H.I.A., Metwali E.M.R., Almaghrabi O.A.H., 2014. Micropropagation of Stevia rebaudiana Betroni and assessment of genetic stability of in vitro regenerated plants using inter simple sequence repeat (ISSR) marker. Plant Biotechnol. 31, 249–256. https://doi.org/10.5511/plantbiotechnology.14.0707a
- Taleie N., Hamidoghli S., Hamidoghli Y., 2012. In vitro plantlet propagation of Stevia
- rebaudiana Bertoni. South-West. J. Hortic. Biol. Environ. 3(1), 99–108. https://doi.org/10.20546/ijcmas.2017.607.122
- Taware A., Mukadam S.S., Chavan A., Taware S.D., Ambedkar B., 2010. Comparative studies of in vitro and in vivo grown plants and callus of Stevia rebaudiana (Bertoni). Int. J. In-tegr. Biol. 9, 10–15.
- Thiyagarajan M., Venkatachalam P., 2012. Evaluation of the genetic fidelity of in vitro propagated natural sweetener plant (Stevia rebaudiana Bert.) using DNA-based markers. Plant Cell Biotechnol. Mol. Biol. 13, 93–98.
- Thiyagarajan M., Venkatachalam P., 2015. Assessment of genetic and biochemical diversity of Stevia rebaudiana Bertoni by DNA fingerprinting and HPLC analysis. Ann. Phytomedicine. 4, 79–85.
Downloads
Download data is not yet available.