Efficient in vitro plant regeneration from leaf-derived callus and genetic fidelity assessment of an endemic medicinal plant Ranunculus wallichianus Wight & Arnn by using RAPD and ISSR markers

Abstract

Ranunculus wallichianus is a medicinally important plant and an endemic species to Western Ghats of South India. An e cient and reliable indirect regeneration protocol system for R. wallichianus was developed from leaf explants in the present investigation. Leaf explants were cultured on both fullstrength and half-strength MS (Murashige & Skoog) medium supplemented with different concentrations (1.0 mg L − 1 to 3.0 mg L − 1 ) of 2,4-D and NAA. Among the different concentrations tested, the highest percentage of yellowish green compact nodular callus formation was observed on half-strength MS medium with 2.0 mg L − 1 of 2, 4-D. Then, the in vitro raised organogenic callus was cultured on half strength MS medium containing various concentrations (1.0 mg L − 1 to 3.0 mg L − 1 ) of BA, KIN and TDZ with 0.5 mg L − 1 NAA and 10% CW for in vitro shoot regeneration. The highest percentage of regeneration response (97%) and maximum number of shoots formation (11.1 ± 0.13 shoots/culture with 9.2 ± 0.35 cm mean shoot length) were obtained from MS medium containing 2.5 mg L − 1 BA with 0.5 mg L − 1 NAA and 10% CW. The well elongated in vitro raised shoots were rooted in half strength MS medium with 2.5 mg L − 1 IBA + 250 mg L − 1 activated charcoal shows high frequency of root formation. The well rooted plantlets were successfully hardened and acclimatized with the survival rate of 94%. Clonal delity of in vitro raised plantlets was assessed by using DNA based RAPD and ISSR molecular markers. The total of 56 and 47 monomorphic bands were obtained from RAPD and ISSR markers respectively. This present in vitro propagation protocol system could be an effective for the conservation of R. wallichianus with their genetic purity and its further investigations.

High Frequency Direct Organogenesis, Genetic Homogeneity, Chemical Characterization and Leaf Ultra-Structural Study of Regenerants in Diplocyclos palmatus (L.) C. Jeffrey

“…Analysis of morphological, molecular, and biochemical attributes in micropropagated plants assure the long-term stability of the acclimatization procedure. Random amplified polymorphic DNA (RAPD) and inter simple sequence repeat (ISSR), both of which are DNA-based molecular markers, are highly repeatable marking systems that have been widely used to preserve the genetic integrity of in vitro-propagated plants [31][32][33][34][35][36][37]. The present investigation was carried out to develop a micropropagation protocol with improved results in terms of shoot formation, rooting and acclimatization.…”

An efficient in vitro organogenesis protocol for the endangered relic tree species Bretschneidera sinensis and genetic fidelity assessment using DNA markers

“…Our protocol is controlled and reliable, as adventitious shoots transplanted more than 10 times are consistent with the genetic stability of first-generation donor material. Our findings align with similar genetic fidelity achievements in species like Cornus alba , Thunbergia coccinea Wall., Manglietiastrum sinicum , Ranunculus wallichianus , and Haplophyllum tuberculatum ( Erişen et al., 2020 ; Srinivasan et al., 2021 ; Alsafran et al., 2022 ; Sultana et al., 2022 ; Luo et al., 2023 ), where direct organogenesis shoots exhibited consistent genetic profiles, as verified by ISSR and RAPD markers ( Backiyarani et al, 2021 ).…”

The Comprehensive In Vitro Propagation and Genetic Homogeneity Analysis of Cryptocoryne crispatula var. yunnanensis: Addressing the Conservation Concerns for an Endangered Species in China and the Mekong Basin

“…As reported by Priyadharshini et al (2020), ISSR markers have been used to successfully identify 35 distinct, reproducible DNA bands that ranged in size from 500 to 2,500 base pairs and demonstrated a monomorphic pattern. Furthermore, the utility of ISSR markers extends to other species, such as Ficus carica, Prunus salicina and Ranunculus wallichianus (Ling et al, 2018;Thakur et al, 2021;Srinivasan et al, 2021).…”