New biological trends on cell and callus growth and azadirachtin production in Azadirachta indica

2021

Preprint

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Neem is the medicinal plant used as antimalarial, antibacterial, antiviral, insecticide, and antimicrobial. This study aimed to investigate and prediction the effect of yeast extract and sampling time on cell growth, secondary metabolites synthesis, SQS1 and MOF1 genes expression by response surface methodology. The highest fresh and dry cell weights were 580.25 g/L and 21.01 g/L obtained 6 days after using 100 mg/L yeast extract. The highest azadirachtin accumulation and production were 16.08 mg/g DW and 219.78 mg/L obtained 2 and 4 days after using 25 mg/L yeast extract. Maximum mevalonic acid accumulation and production were 1.75 mg/g DW and 23.77 mg/L observed 2 days after application of 50 mg/L yeast extract. The highest amount of squalene accumulation (0.22 mg/g DW) and production (4.53 mg/L) were achieved 4 days after using 50 mg/L yeast extract. Prediction results showed that the highest azadirachtin accumulation (13.61 mg/g DW) and production (190.50 mg/L), mevalonic acid accumulation (0.50 mg/g DW) and production (5.57 mg/L), and squalene accumulation (0.30 mg/g DW) obtained by using 245 mg/L yeast extract for 2 days, 71 mg/L yeast extract for 2 days, 200 mg/L yeast extract for 4.96 days, without yeast extract for 6.54 days and 4 days, respectively. Also, predicted that the highest squalene production is achieved by long-term exposure to high concentrations of yeast extract. The qRT-PCR analysis showed the maximum relative gene expression of SQS1 and MOF1 obtained by using 150 mg/L yeast extract for 4 days and 25 mg/L yeast extract for 2 days, respectively.

Section: Plant Materials and Cell Suspension Culture Establishmentmentioning

confidence: 99%

Improvement and prediction of secondary metabolites production under yeast extract elicitation of Azadirachta indica cell suspension culture using response surface methodology

2021

Preprint

Self Cite

“…Then cultures were maintained in the growth chamber at 25 ± 2 °C in the dark. The friable calli were transferred to the liquid MS medium with the same concentrations of picloram and kinetin and kept on a rotary shaker at 110 rpm and 26 ± 2 °C in the dark and sub-cultured every 12 days (Farjaminezhad and Garoosi 2019 ).…”

Section: Methodsmentioning

confidence: 99%

“…The stock solution of yeast extract (Merck, Germany) was prepared by dissolving of yeast extract in distilled water and then filtered using a 0.22 µm syringe filter. According to our previous study growth curve (Farjaminezhad and Garoosi 2019 ), eight days after cell culture different concentrations of yeast extract including 0, 25, 50, 100, 150 and 200 mg/L were added to cell suspension cultures. The cultures were kept on a rotary shaker at 110 rpm and 26 ± 2 °C in the dark and sampling was performed after 2, 4, 6, 8, 10 and 12 days of each treatment.…”

Section: Methodsmentioning

confidence: 99%

Improvement and prediction of secondary metabolites production under yeast extract elicitation of Azadirachta indica cell suspension culture using response surface methodology

2021

AMB Expr

Self Cite

Neem is a medicinal plant used as antimalarial, antibacterial, antiviral, insecticide, and antimicrobial drug. This study aimed to investigate and predict the effect of yeast extract and sampling time on cell growth, secondary metabolites synthesis, SQS1 and MOF1 genes expression by response surface methodology. The highest fresh and dry cell weights were 580.25 g/L and 21.01 g/L, respectively obtained 6 days after using 100 mg/L yeast extract. The highest azadirachtin accumulation and production were 16.08 mg/g DW and 219.78 mg/L obtained 2 and 4 days, respectively after using 25 mg/L yeast extract. Maximum mevalonic acid accumulation (1.75 mg/g DW) and production (23.77 mg/L) were observed 2 days after application of 50 mg/L yeast extract. The highest amount of squalene accumulation (0.22 mg/g DW) and production (4.53 mg/L) were achieved 4 days after using 50 mg/L yeast extract. Prediction results exhibited the highest azadirachtin accumulation (13.61 mg/g DW) and production (190.50 mg/L), mevalonic acid accumulation (0.50 mg/g DW) and production (5.57 mg/L), and squalene accumulation (0.30 mg/g DW) by using 245 mg/L yeast extract for 2 days, 71 mg/L yeast extract for 2 days, 200 mg/L yeast extract for 4.96 days, without yeast extract for 6.54 days and 4 days, respectively. Also, it was predicted that the highest squalene production is achieved by long-term exposure to high concentrations of yeast extract. The qRT-PCR analysis displayed the maximum relative gene expression of SQS1 and MOF1 by using 150 and 25 mg/L yeast extract for 4 and 2 days treatment.

“…The leaves of neem collected from Bandar Abbas city of Iran and surface sterilized with ethanol and sodium hypochlorite and cultured on MS medium containing 1 mg/L picloram and 2 mg/L kinetin and maintained in the growth chamber at 25 ± 2 °C in the dark. The friable calli were transferred to the liquid MS medium with same concentrations of picloram and kinetin and kept on a rotary shaker at 110 rpm with 26 ± 2 °C in the dark and sub-cultured every 12 days (Farjaminezhad and Garoosi 2019).…”

Section: Plant Materials and Cell Suspension Culture Establishmentmentioning

confidence: 99%

“…The cell suspension cultures were transferred to 100 mL Erlenmeyer asks containing 25 mL liquid MS medium supplemented with 1 mg/L picloram and 2 mg/L kinetin with an initial cell density of 2.6 × 10 5 (SCV = 8%). According to the our previous study growth curve (Farjaminezhad and Garoosi 2019), eight days after culture different concentrations of yeast extract including 0, 25, 50, 100, 150 and 200 mg/L were added to cell suspension culture (Merck, Germany). The cultures were kept on a rotary shaker at 110 rpm with 26 ± 2 °C in the dark and sampling were performed after 2, 4, 6, 8, 10 and 12 days.…”

Section: Elicitation With Yeast Extractmentioning

confidence: 99%

Improvement and Prediction of Secondary Metabolites Biosynthesis Under Yeast Extract Elicitation of Neem Cell Suspension Culture Using Response Surface Methodology

2021

Preprint

Self Cite

Neem is the medicinal plant used as antimalarial, antibacterial, antiviral, insecticide and antimicrobial. The aim of this study was investigate effect of yeast extract and sampling time on cell growth, secondary metabolites synthesis, SQS1 and MOF1 genes expression and prediction of secondary metabolites synthesis by response surface methodology. Highest fresh and dry cell weight were 580.25 g/L and 21.01 g/L obtained 6 days after addition 100 mg/L yeast extract. Highest azadirachtin accumulation and production were 16.08 mg/g DW and 219.78 mg/L obtained at 25 mg/L yeast extract for 2 and 4 days. Maximum mevalonic acid accumulation and production were 1.746 mg/g DW and 23.77 mg/L observed 2 days after addition 50 mg/L yeast extract addition. Highest amount of squalene accumulation (0.215 mg/g DW) and production (4.526 mg/L) were achieved 4 days after application of 50 mg/L yeast extract. Prediction results showed that highest azadirachtin accumulation (13.607 mg/g DW) and production (190.50 mg/L), mevalonic acid accumulation (0.500 mg/g DW) and production (5.57 mg/L) and squalene accumulation (0.301 mg/g DW) obtained by using 245 mg/L yeast extract for 2 days, 71 mg/L yeast extract for 2 days, 200 mg/L yeast extract for 4.96 days, whitout yeast extract for 6.54 days and 4 days, respectively and predicted that highest squalene production is achieved by long-term exposure to high concentrations of yeast extract. qRT-PCR analysis showed the maximum relative gene expression of SQS1 and MOF1 by addition 150 mg/L yeast extract for 4 days and 25 mg/L yeast extract for 2 days, respectively.