Evaluation of Clonal Uniformity in Six Superior Cacao Clones Based on SSR Marker

Indah Sulistiyorini, Rubiyo Rubiyo, Sudarsono Sudarsono


Propagation of cacao plants is generally carried out vegetatively. Therefore, plants that are clonally propagated should be genetically uniform. Genetic uniformity in cacao clones is also very important information for germplasm conservation and in obtaining pure parental crosses. Evaluation of genetic uniformity can be seen through analysis using SSR markers. This study aimed to determine the genetic uniformity in six cacao clones using SSR markers. This experiment was conducted at IIBCRI Integrated Laboratory in Sukabumi and Plant Molecular Biology Laboratory, IPB Bogor, from September 2015 to December 2016.  Six cacao clones used (TSH 858, TSH 908, ICS 13, PA 300, GC 7 and UIT) are from Kalitelepak experimental station of  PTPN XII, Genteng District, Banyuwangi, East Java.  Ten samples were taken randomly to represent cacao clones. DNA amplification was carried out using 12 SSR markers. The result showed that 12 SSR markers generated 45 alleles with the number of alleles per locus was 3-4 alleles. The polymorphic information content (PIC) ranges from 0.370.67, which are identied as very informative molecular analysis in identifying the genetical uniformity of the evaluated cacao population. Six SSR loci generated variant alleles within both the TSH 858 and UIT clones, indicating there are off-type plants in these two samples. Clonal uniformity were detected for samples of the GC 7, ICS 13, PA 300 and TSH 908 clones. On the other hand, 8.33% of evaluated samples within the TSH 858 and UIT clones were off-type plants.


Cacao; clone; genetic uniformity; SSR markers

Full Text:

PDF (Indonesian)


Allen, G., Flores-Vergara, M., Krasynanski, S., Kumar, S., & Thompson, W. (2006). A modified protocol for rapid DNA isolation from plant tissues using cetyltrimethylammonium bromide. Nat. Prot, 1(5), 2320–2325. https://doi.org/doi:10.1038/nprot.2006.384

Ardiyani, F., & Yuliasmara, F. (2015). Perbanyakan tanaman kakao. In T. Wahyudi, Pujiyanto, & Misnawi (Eds.), Kakao :Sejarah, Botani, Proses Produksi, Pengolahan, dan Perdagangan (p. Hal 727). Yogyakarta: Gadjah Mada University Press.

Bandupriya, H. D. D., Iroshini, W. W. M. A., Perera, S. A. C. N., Vidhanaarachchi, V. R. M., Fernando, S. C., Santha, E. S., & Gunathilake, T. R. (2017). Genetic fidelity testing using ssr marker assay confirms trueness to type of micropropagated Coconut (L.) plantlets derived from unfertilized ovaries. The Open Plant Science Journal, 10(1), 46–54. https://doi.org/10.2174/1874294701710010046

Cahyaningsih, Y. F., Wiendi, N. M. A., & Toruan-Mathius, N. (2016). Deteksi kestabilan genetik ramet kelapa sawit hasil kultur in vitro menggunakan ssr detection of genetic stability in ramet of oil palm derived from in vitro culture by SSR. J.Agron. Indonesia, 44(1), 83–90.

Creste, S., Neto, A. T., & Figueira, A. (2001). Detection of single sequence repeat polymorphisms in denaturing polyacrylamide sequencing gels by silver staining. Plant Molecular Biology Reporter, 19(4), 299–306. https://doi.org/10.1007/BF02772828

Cruz-Martínez, V., Castellanos-Hernández, O. A., Acevedo-Hernández, G. J., Torres-Morán, M. I., Gutiérrez-Lomelí, M., Ruvalcaba-Ruiz, D., … Rodríguez-Sahagún, A. (2017). Genetic fidelity assessment in plants of Sechium edule regenerated via organogenesis. South African Journal of Botany, 112, 118–122. https://doi.org/10.1016/j.sajb.2017.05.020

Govindaraj, M., Vetriventhan, M., & Srinivasan, M. (2015). Importance of genetic diversity assessment in crop plants and its recent advances : An overview of its analytical perspectives. Genetics Research International, 2015, 1–15.

Izzah, N. K., Lee, J., Perumal, S., Park, J. Y., Ahn, K., Fu, D., … Yang, T.-J. (2013). Microsatellite-based analysis of genetic diversity in 91 commercial Brassica oleracea L . cultivars belonging to six varietal groups. Genet Resour Crop Evol, 60, 1967–1986. https://doi.org/10.1007/s10722-013-9966-3

Johnsiul, L., & Awang, A. (2016). Utilization of molecular markers to detect the authenticity of cocoa clones. International Journal of Agriculture, Forestry and Plantation, 3(June), 101–104.

Kalia, R. K., Rai, M. K., Kalia, S., Singh, R., & Dhawan, A. K. (2011). Microsatellite markers: An overview of the recent progress in plants. Euphytica, 177(3), 309–334. https://doi.org/10.1007/s10681-010-0286-9

Kurniasih, S., Rubiyo, Setiawan, A., Purwantara, A., & Sudarsono. (2011). Analisis keragaman genetik plasma nutfah kakao (Theobrtoma cacao L.) berdasarkan marka SSR. Jurnal Littri, 17(4), 156–162.

Lanaud, C., Risterucci, A. M., Pieretti, I., Falque, M., Bouet, A., & Lagoda, P. (1999). Isolation and characterization of microsatellites in Theobroma cacao L. Molecular Ecology, 8, 2141–2152.

Mahdi, A., Hare, K., Maliheh, E., Mohsen, M., & Mahdieh, M. (2015). Assessment of clonal fidelity in micropropagated horticultural plants. Journal of Chemical and Pharmaceutical Research , 7( 12 ), 977–990.

Marshall TC , Slate, J., Kruuk, L. E. B., & Pemberton, J. M. (1998). Statistical confidence for likelihood-based paternity inference in natural population. Mol. Ecol, 7(5), 639–655.

Mateescu, R. G., Zhang, Z., Tsai, K., Phavaphutanon, J., Burton-Wurster, N. I., Lust, G., … Todhunter, R. J. (2005). Analysis of allele fidelity, polymorphic information content, and density of microsatellites in a genome-wide screening for hip dysplasia in a crossbreed pedigree. Journal of Heredity, 96(7), 847–853. https://doi.org/10.1093/jhered/esi109

Nagy, S., Poczai, P., Cernák, I., Gorji, A. M., Hegedus, G., & Taller, J. (2012). PICcalc: An online program to calculate polymorphic information content for molecular genetic studies. Biochemical Genetics, 50(9–10), 670–672. https://doi.org/10.1007/s10528-012-9509-1

Nookaraju, A., & Agrawal, D. C. (2012). Genetic homogeneity of in vitro raised plants of grapevine cv . Crimson Seedless revealed by ISSR and microsatellite markers. South African Journal of Botany, 78, 302–306. https://doi.org/10.1016/j.sajb.2011.08.009

Pandey, R. N., Singh, S. P., Rastogi, J., Sharma, M. L., & Singh, R. K. (2012). Early assessment of genetic fidelity in sugarcane (Saccharum officinarum) plantlets regenerated through direct organogenesis with RAPD and SSR markers. Australian Journal of Crop Science, 6(4), 618–624.

Permentan. (2013). Standar Operasional Prosedur Penetapan Kebun Sumber Benih, Sertifikasi Benih, dan Evaluasi Kebun Sumber Benih Tanaman Kakao ((Theobroma cacao L.).

Perrier, X., & J.P., J.-C. (2010). DARwin software. http://darwin.cirad.fr/darwin.

Powell, W., Machray, G. C., & Provar, J. (1996). Polymorphism revealed by simple sequence r e p e a t s w. Trends in Plant Science, 1(7), 215–221.

Pugh, T., Fouet, O., Risterucci, A. M., Brottier, P., Abouladze, M., Deletrez, C., … Lanaud, C. (2004). A new cacao linkage map based on codominant marker: development and integration of 201 new microsatellite markers. Theor. Appl. Genet, 108, 1151–1161. https://doi.org/DOI 10.1007/s00122-003-1533-4

Rogrio, M. F. S., Uilson, V. L., Didier, C., Jose, L. P., Eline, M. L., Tamiles, B. M., & Karina, P. G. (2014). A protocol for large scale genomic DNA isolation for cacao genetics analysis. African Journal of Biotechnology, 13(7), 814–820. https://doi.org/10.5897/AJB2013.13181

Rubiyo, Izzah, N. K., Sulistiyorini, I., & Tresniawati, C. (2015). Evaluation of genetic diversity in cacao collected from Kolaka, Southeast Sulawesi, using SSR markers. Indonesian Journal of Agricultural Science, 16(2), 71–78.

Saunders, J. A., Mischke, Æ. S., & Leamy, Æ. E. A. (2004). Selection of international molecular standards for DNA fingerprinting of Theobroma cacao. Theoretical and Applied Genetics, 110(1), 41–47. https://doi.org/10.1007/s00122-004-1762-1

Singh, R., Nagappan, J., Tan, S., Panandam, J. M., & Cheah, S. (2007). Development of simple sequence repeat ( SSR ) markers for oil palm and their application in genetic mapping and fingerprinting of tissue culture clones. Asia Pacific Journal of Molecular Biology & Biotechnology, 15(3), 121-131.

Syukur, M., Sujiprihati, S., & Yunianti, R. (2015). Teknik Pemuliaan Tanaman. Jakarta: Penebar Swadaya.

Wicaksono, I. N. ., Rubiyo, Sukma, D., & Sudarsono. (2017). Analisis Keragaman genetik 28 nomor koleksi kakao (Theobroma cacao L.) berdasarkan marka SSR. Jurnal Tanaman Industri dan Penyegar, 4(1), 13–22.

DOI: http://dx.doi.org/10.21082/jtidp.v5n3.2018.p135-144


  • There are currently no refbacks.

Copyright (c) 2018 Jurnal Tanaman Industri dan Penyegar

Creative Commons License
This work is licensed under a Creative Commons Attribution-NonCommercial-ShareAlike 4.0 International License.

 Creative Commons License
This work is licensed under a Creative Commons Attribution-NonCommercial-ShareAlike 4.0 International License.


P-ISSN: 2356-1297
E-ISSN: 2528-7222
Accredited No.30/E/KPT/2018 on Oktober 24, 2018 by Ministry of Research, Technology and Higher Education of the Republic of Indonesia


Jurnal Tanaman Industri dan Penyegar (JTIDP) Editorial Office :

Indonesian Industrial and Beverage Crops Research Institute
Jl. Raya Pakuwon Km. 2, Parungkuda, Sukabumi 43357 Jawa Barat Indonesia
Telp : (0266) 6542181
Fax : (0266) 6542087
Email : jtidp@litbang.pertanian.go.iduppublikasi@gmail.com
Website : http://balittri.litbang.pertanian.go.id

View My Stats