Sterilization protocols and the effect of plant growth regulators on callus induction and secondary metabolites production in in vitro cultures Melia azedarach L.

Fatemeh Ahmadpoor,Nasser Zare,Rasool Asghari,Parisa Sheikhzadeh

doi:10.1186/s13568-022-01343-8

Fatemeh Ahmadpoor, Nasser Zare + Show 2 more

Open Access

https://doi.org/10.1186/s13568-022-01343-8

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Journal: AMB Express	Publication Date: Jan 10, 2022
Citations: 9	License type: open-access

Affiliation: University of Mohaghegh Ardabili

Abstract

Melia azedarach L. is a valuable source of antioxidants and secondary metabolites. This study is a first extensive report about the effect of different serialization protocols and plant growth regulators (PGRs) on explant disinfection efficiency, callus induction and secondary metabolites production and accumulation in callus cultures of M. azedarach L. In this regard, the effect of plant growth regulators on callus induction and secondary metabolites production were examined. In addition, different sterilization agents were evaluated for disinfection of chinaberry leaf explants. The results showed that the lowest percentage of explant contamination and browning with the highest percentage of callus induction and callus growth obtained with explants pretreated with benomyl (2 g/L) for 2 h and sterilized with 7% H2O2 for 10 min and NaOCl 2% (without pH adjustment) for 12 min. Although adjusting the pH of NaOCl to pH = 7 and 10 significantly reduced the microbial contamination and increased the percentage of contamination-free cultures of M. azedarach L., adversely influenced the explant viability and callus induction and growth. The highest percentage of callus induction obtained on the MS medium containing 3 mg/L NAA/2,4-D and 1 or 3 mg/L Kin/BAP, and the highest callus yield (1804.833 mg/explant) belonged to the MS medium supplemented with 5 mg/L 2,4-D and 5 mg/L Kin. The callus cultures grown on the MS medium supplemented with 3 mg/L NAA and 1 mg/L Kin produced the highest amount of Quercetin (2.06 mg/g fresh weight), Rutin (5.56 mg/g fresh weight) and Kaempferol (1.84 mg/g fresh weight).

Highlights

Melia azedarach L. is one of the most valuable pharmaceutical plants
The reports on tissue culture of woody plants indicate that surface disinfection was not efficient to produce clean cultures, and always is accompanied with a high level of bacterial and fungal contamination (Pence 2005)
The results indicated that the percentage of bacterial and fungal contamination, clean explants, explants viability and browning and callus induction and growth were significantly influenced by disinfection methods

Summary

Introduction

Melia azedarach L. (known as Chinaberry) is one of the most valuable pharmaceutical plants. In vitro fungal and bacterial contaminations are one of the most important limiting factors, in woody plants. In order to overcome fungal and bacterial contaminations in the disinfection stage, different surface sterilizing agents including sodium and calcium hypochlorite, ethanol, mercuric chloride, Ahmadpoor et al AMB Express (2022) 12:3 silver nitrate, antibiotics, and fungicide were used in laboratories worldwide. Assareh and Sardabi (2005) reported that among calcium and sodium hypochlorite and mercuric chloride agents used for surface sterilization of Ziziphus spina-christi explants, Ca(OCl) (5% for 20 min) was the most efficient in explants sterilization. The reports on tissue culture of woody plants indicate that surface disinfection was not efficient to produce clean cultures, and always is accompanied with a high level of bacterial and fungal contamination (Pence 2005)

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Conclusion