RESEARCH PAPER
Faithful transmission and expression of transgenes via somatic embryos
of transgenic cassava plants at the sixth cycle of vegetative propagation
1
Department of Crop Production and Protection, Faculty of Agriculture, Obafemi Awolowo University, Ile-Ife, Nigeria
Submission date: 2016-02-18
Final revision date: 2016-05-03
Acceptance date: 2016-05-17
Publication date: 2016-07-20
BioTechnologia 2016;97(2):65-77
KEYWORDS
TOPICS
ABSTRACT
The transmission of transgenes via somatic embryos from one vegetative cycle to another in cassava (Manihot esculenta Crantz) has not been well studied to date. This study examined somatic embryogenesis and regeneration from transgenic cassava plants expressing the β -glucuronidase gene (GUS) under the control of a cassava vein mosaic promoter (CsVMV) at the sixth cycle of vegetative propagation. Primary, secondary and cyclic somatic embryos were induced from an axillary bud and immature leaf lobe explants. Plantlet regeneration via shoot organogenesis was examined in the cotyledons of somatic embryos. Histochemical, polymerase chain reaction (PCR) and reverse transcriptase PCR analyzes were used to confirm the presence of the GUS gene in regenerated plants. The frequencies of primary, secondary and cyclic somatic embryos from immature leaf lobes were significantly greater (P < 0.05) than those of axillary buds. The transient expression of the GUS gene was detected in all types of somatic embryos from both axillary bud and immature leaf lobe explants. Shoot induction from cotyledons of somatic embryos produced from axillary buds was 27.0% greater than that of leaf lobes, with selection performed with the use of kanamycin. In regenerated transgenic plants (grown from somatic embryos obtained from immature leaf lobes), a non-uniform expression of GUS was observed in three chimeric lines assessed by histochemical and molecular analyzes. In regenerated transgenic plants obtained from somatic embryos from axillary buds, uniform and high expression of GUS was observed in one line in all tested tissues and in most cell types of leaves, stems, petioles, roots and tubers, thus showing the faithful transmission of the GUS transgene under the control of CsVMV promoter via a somatic embryo.
REFERENCES (36)
1.
Bull S.E, Ndunguru J., Gruissem W., Beeching J.R., Vanderschuren H. (2011) Cassava constraints to production and the transfer of biotechnology to African laboratories. Plant Cell Rep. 30: 677-679.
2.
Ceballos H., Iglesias C.A., Pereze J.C., Dixon A.G.O. (2004) Cassava breeding: opportunities and challenges. Plant Mol. Biol. 56: 503-516.
3.
Dellaporta S.L., Wood J., Hicks J.B. (1983) A plant DNA mini preparation version II. Plant Biotech. Mol. Biol. Rep. 1: 19- 21.
4.
El-Sharkawy M.A. (2004) Cassava biology and physiology. Plant Mol. Biol. 56: 481-501.
5.
FAO (2008) Cassava for food and energy security. FAO Media Centre, Rome. www.fao.org/newsroom/en/news/2008/ 1000899/index.html. Accessed Oct 2010.
6.
Gantait S., Kundu S., Ali N., Sahu N.C. (2015) Synthetic seed production of medicinal plants: a review on influence of explants, encapsulation agent and matrix. Acta Physiol. Plant. 37: 98-107.
7.
Hankoua B.B., Ng S.Y.C., Fawole I., Pouti-Kaerlas J., Pillay M., Dixon A.G.O. (2005) Regeneration of a wide range of African cassava genotypes via shoot organogenesis from cotyledons of maturing somatic embryos and conformity of field-established regenerants. Plant Cell Tiss. Organ Cult. 82: 221-231.
8.
Hankoua B.B., Taylor N.J., Ng S.Y.C., Fawole I., Puonti-Kaerlas J., Padmanabhan C., Fondong V.N. (2006) Production of the first transgenic cassava in Africa via direct shoot organogenesis from friable embryogenic calli and germination of maturing somatic embryos. Afri. J. Biotech. 5: 1700-1712.
9.
Ibrahim A.B. Heredia F., Pinheiro C., Aragao F., Campos F. (2008) Optimization of somatic embryogenesis and selection regimes for particle bombardment of friable embryogenic callus and somatic cotyledons of cassava (Manihot esculenta Crantz). Afr. J. Biotech.16: 2790-2797.
10.
Ihemere U., Ariaz-Garzon D., Lawrence S., Sayre R. (2006) Genetic modification of cassava for enhanced starch production. Plant Biotech. J. 4: 453-465.
11.
Ingelbrecht I., Dixon A.G.O., Raji A., Opabode J.T. (2010) Molecular enhancement of cassava (Manihot esculenta Crantz) for resistance to the cassava brown streak disease. Final Technical Report submitted to Rockefeller Foundation, New York, USA: 30.
12.
Ipekci Z., Gozukirmizi N. (2011) Direct somatic embryogenesis and synthetic seed production from Paulownia elongate. Plant Cell Rep. 22: 16-24.
13.
Jefferson R.A., Kavanagh T.A., Bevan M. (1987) GUS fusions: Beta-glucuronidase as a sensitive and versatile gene fusion marker in higher plants. EMBO J. 6: 3901-3907.
14.
Jørgensen K., Bak S., Busk P.K., Sørensen C., Olsen C.E. (2005) Cassava plants with a depleted cyanogenic glucoside content in leaves and tubers. Distribution of cyanogenic glucosides, their site of synthesis and transport, and blockage of the biosynthesis by RNA interference technology. Plant Physiol. 139: 363-337.
15.
Kulus D., Zalewska M. (2014) In vitro plant recovery from alginate encapsulated Chrysanthemum × grandiflorum (Ramat) Kitam shoot tips. Propagat. Ornament. Plants 14(1): 3-12.
16.
Li H.Q., Guo J.Y, Huang Y.W., Liang C.Y., Liu H.X., Potrykus I., Puonti-Kaerlas J. (1998) Regeneration of cassava plants via shoot organogenesis. Plant Cell Rep. 17: 410-414.
17.
Liu W., Liang Z., Sibbald S., Hunter D., Tian L. (2013) Preservation and faithful expression of transgene via artificial seeds in alfalfa. PLoS ONE8(5), e56699. doi: 10.1371/ journal.
18.
Nassar N., Ortiz R. (2010) Breeding cassava to feed the poor. Sci. Amer. 302: 78-84.
19.
Nweke F.I., Spencer D.D.C., Lynam J.K. (2002) The cassava transformation. Michigan State University Press, East Lansing.
20.
Nyaboga E.N., Njiru J.M., Tripathi L. (2015) Factors influencing somatic embryogenesis, regeneration, and Agrobacterium- mediated transformation of cassava (Manihot esculenta Crantz) cultivar TME14. Front Plant Sci. 6: 411-424.
21.
Murashige T., Skoog F. (1962) A revised medium for rapid growth and bioassays with tobacco tissue cultures. Physiol. Plant. 15: 473-497.
22.
Opabode J.T. (2006) Agrobacterium-mediated transformation of plants:emerging factors that influence efficiency. Biotech. Mol. Biology Rev. 1: 12-20.
23.
Opabode J.T., Oyelakin O.O., Akinyemiju O.A., Ingelbrecht I.L. (2013) Primary somatic embryos from axillary meristems and immature leaf lobes of selected African cassava varieties. Brit. Biotech. J. 3: 263-273.
24.
Opabode J.T, Oyelakin O.O., Akinyemiju O.A. Ingelbrecht I. L. (2014) Influence of type and age of primary somatic embryo on secondary and cyclic somatic embryogenesis of cassava (Manihot esculenta Crantz). Brit. Biotech. J. 4: 254-269.
25.
Oyelakin O.O., Opabode J.T., Raji A.A., Ingelbrecht I.L. (2015) A cassava vein mosaic virus promoter cassette induces high and stable gene expression in clonally propagated transgenic cassava (Manihot esculenta Crantz). South Afr. J. Bot. 97: 184-190.
26.
Puonti-Kaerlas J. (1998) Cassava Biotechnology. Biotech. Gen. Eng. Rev. 15: 329-336.
27.
Reddy M.C., Murthy K.S.R., Pullaih T. (2012) Synthetic seeds: A review in agriculture and forestry. Afr. J. Biotech. 11(78): 14254-14275.
28.
SAS Institute (2002). Statistical Analysis System “SAS/ STAT” User’s Guide, Version 6, Fourth edition; Vol. 1. The SAS Institute Inc., Gary, N.C.
29.
Schreuder M.M., Raemakers C.J.J.M., Jacobsen E., Visser R.G.F. (2001) Eficient production of transgenic plants by Agrobacterium – mediated transformation of cassava (Manihot esculenta Crantz). Euphytica 120: 35-45.
30.
Sukmadjaja D., Widhiastuti H. (2011) Effects of plant growth regulators on shoot multiplication and root induction of cassava varieties culture. Biotropia 18: 50-60.
31.
Rossin C.B., Rey M.E.C. (2011) Effect of explant source and auxins on somatic embryogenesis of selected cassava (Manihot esculenta Crantz) cultivars. South Afr. J. Bot. 77: 59-65.
32.
Taylor N.J., Chavarriaga P., Raemakers K., Siritunga D., Zhang P. (2004) Development and application of transgenic technologies in cassava. Plant Mol. Biol. 56: 671- 678.
33.
Verdaguer B., deKochko A., Beachy R.N., Fauquet C. (1996) Isolation and expression in transgenic tobacco and rice plants of the Cassava vein mosaic virus (CVMV) promoter. Plant Mol. Biol. 31: 1129-1139.
34.
Zhang P., Pounti-Kaerlas I. (2004) Regeneration of transgenic cassava from transformed embryogenic tissues. Meth. Mol. Biol. 286: 1025-1049.
35.
Zhang P., Potrykus I., Puonti-Kaerlas J. (2000) Efficient production of transgenic cassava using negative and positive selection. Transg. Res. 9: 405-415.
36.
Zhang P., Vanderschuren H., Futterer J., Gruissem W. (2005) Resistance to cassava mosaic disease in transgenic cassava expressing antisense RNAs targeting virus replication genes. Plant Biotech. J. 3: 385-397.
Share
RELATED ARTICLE
| eISSN: | 2353-9461 |
| ISSN: | 0860-7796 |
We process personal data collected when visiting the website. The function of obtaining information about users and their behavior is carried out by voluntarily entered information in forms and saving cookies in end devices. Data, including cookies, are used to provide services, improve the user experience and to analyze the traffic in accordance with the Privacy policy. Data are also collected and processed by Google Analytics tool (more).
You can change cookies settings in your browser. Restricted use of cookies in the browser configuration may affect some functionalities of the website.
You can change cookies settings in your browser. Restricted use of cookies in the browser configuration may affect some functionalities of the website.
