Isolation, Culture and characterisation of Adipose Tissue-Derived Mesenchymal Stem Cells (ADMSCs) by Explant-Enzymatic Methods
Volume 8, Issue 4, Spring 2018, Pages 303-313
https://doi.org/10.52547/JCT.8.4.303
mh mohammadi mahdiabadi hasani, M Nabiuni, Kazem Parivar, Siamak Yari, Alireza Sahebi
Abstract Aim: In this study, we use combination of enzymatic and explant methods, for isolation and culture of ADMSCs.
Materials and Method: Adipose tissues dissected out from the abdominal region of male Wistar rats. Tissues were minced into small pieces (1-2 mm), and then treated with trypsin-EDTA 0.25% for 30 minutes at 37 ̊C in shaker-incubator. Enzymatic treated tissues were centrifuged and floating parts were cultured. Statistical analysis of the cells number was investigated using GraphPad Prismv6 software.
Results: Results showed that ADMSCs isolated with our methods have growth characteristics similar to conventional methods. ADMSCs were maintained up to 10th passage and exhibited a homogeneous population in terms of appearance and morphology. We demonstrated that ADMSCs by these combination methods have mesenchymal characteristic markers (positive for CD90, CD44, CD73, CD105 and negative for CD35, CD45, CD11b).
Conclusion: In the current study, we showed that the combination of enzymatic and explant methods creates a simple, inexpensive and reproducible method for isolation and culture of ADMSCs.
Induction and Optimization of Hairy Root Growth Condition for Valeriana officinalis L. Through Inoculation by Agrobacterium rhizogenes
Volume 5, Issue 1, Summer 2014, Pages 23-30
https://doi.org/10.52547/JCT.5.1.23
MR D, N A, M J, A S
Abstract Aim: In this study, with the aim of optimizing hairy root growth conditions, leaf, hypocotyl and root explants were inoculated by strain ‘A13’ of Agrobacterium rhizogenes. Then effect of four basal MS liquid media was considered with different combinations and temperature on rate of hairy roots growth.
Material and methods: In order to hairy root induction via spary and leaf disc methods, two strains of Agrobacterium rhizogenes (A13 and 9534) were used. For confirmation of transformation but not infection of hairy roots, PCR reaction was performed through specific amplification of rolB and virD genes. Obtained hairy roots in a basal liquid MS medium were cultured under different treatments in based on completely randomized design (CRD) with three replicates and dry weight of hairy roots was measured after two months.
Results: Hairy roots were observed in root, hypocotyl and leaf explants after 8 days. The highest percentage of transformation was found in leaf explants. The obtained results showed that the highest (0.19 gr) and the lowest(0.05 gr) rate of hairy root dry weight related to basal liquid MS medium containing 30 g/l sucrose in 35°C and basal MS liquid medium containing 0.2 gr ∕ l NAA hormone,respectively,.
Conclusion: The results indicate the main role of culture medium compounds and temperature in enhancing of hairy root biomass.