Micropropagation and assessment of genetic stability of Dendrobium transparens Wall. Ex Lindl. using RAPD and ISSR markers

Abstract

IntroductionDendrobium species have been widely used for many health disorders since ancient times. However, due to unrelenting collection to meet the increasing demand for their use in medication and other health products, the natural habitats of medicinal Dendrobium transparens have been devastated and are on the verge of extinction.MethodsAn efficient in-vitro propagation protocol for Dendrobium transparens using seed derived protocorms was established and genetic homogeneity of the in-vitro regenerants and the wild plant was studied. ResultsThe maximum seed germination was observed in Full strength Murashige and Skoog medium (FMS). Induction of protocorms were achieved on basal as well as half-strength MS medium. The highest number of shoot (11.9 shoots/explant) was achieved in half MS medium fortified with 100 mL/L coconut water in addition with Benzyl amino purine (BAP) 1 mg/L and Kinetin 2 mg/L. Further, elongated shoots were transferred to full and half strength MS root initiating medium supplemented with different concentration of auxins. However, a maximum of (8.3 ± 0.6, 4.9 ± 0.1 cm) roots were achieved in full MS medium fortified with 100 mL/L coconut water and Napthalene acetic acid (NAA) 1.5 mg/L. Ten rapid Random Amplified Polymorphic DNA (RAPD) and Inter Simple Sequence Repeats (ISSR) primers were used to analyze genetic stability among in-vitro and mother plant. RAPD primers produced a total of 23 fragments while ISSR primers produced a total of 16 fragments. ConclusionThe amplified bands of all the samples of in-vitro plants were similar to bands of mother plant. The present research reported here is indicating the applicability of tissue culture for true-to-type plant production and conservation of D. transperens.

Efficient regeneration of in vitro derived plants and genetic fidelity assessment of Phalaenopsis orchid

“…bananas by Alizadeh and Singh (2009), gerberas by Bhatia et al (2011), Dendrobium densiflorum by Mohanty and Das (2013), Spilanthes calva by Razaq et al (2013), and Rauvolfia hookeri by Ranjush and Gangaprasad (2014) cleared the genetic fidelity test using RAPD primers. But when clonality tests were performed on the mother and in vitro micropropagated seedlings, two of the ten selected RAPD primers showed clear monomorphic bands (Joshi et al, 2023). It was determined that genetic clonality was preserved by taking into consideration the monomorphic banding pattern that the mother and the in vitro cloned plant both displayed.…”

Genetic similarity among Dendrobium species from Indonesia using RAPD markers

“…In this study we used five RAPD primers from the 20 primers (Operon Technology Ltd) that were selected previously. The five selected primers, namely OPD-8 (Dendrobium (Choopeng et al 2019); OPA-7 (Dendrobium (Tikendra et al 2019a), Coelogyne (Hartati 2020), Phaius (Hartati et al 2021), Rhynchostylis retusa (Oliya et al 2021), Dendrobium (Joshi et al 2023)); OPB-12, OPA-13 and OPB-18 (Coelogyne (Hartati 2020), Phaius (Hartati and Samanhudi 2022) have been shown to produce polymorphic bands in orchids. PCR reaction was performed with a total volume of 15 µL containing 0.2 n MdNTP; 1 x reaction buffer; 2mM MgCl2; DNA sample 25 ng; 1 pmol; and 1 unit of Taq DNA polymerase (Promega) using a Thermocycler (Takara) for 45 cycles.…”