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DNA methylation array of irradiated and/or lithium exposed iPSC-derived human neural progenitor cells

ABSTRACT: DNA isolation was performed following the DNeasy Blood and Tissue Kit (Qiagen). Cells were irradiated in an CIX2 Xstrahl X-Ray Irradiator with Aluminum filter at a focus-to-skin distance (FSD) of 40 cm and with an energy of 195kV, 10 mA, for and absorbed dose of 4 GY. To allow for assessment of irradiation damage, cells remained in the incubator for 1 hour. Pre-warmed DMEM/F-12 was mixed with the appropriate amount of LiCl powder (Sigma-Aldrich, Cat# L9650-100G), passed through a 0.22 μm filter, and supplemented with 1% Penicillin-Streptomycin, 1% N2, 0.1% B27, 0.1% EGF, and 0.1% FGF2 to produce 3mM Lithium Chloride (LiCl) hNSPC medium.

ORGANISM(S): Homo sapiens

PROVIDER: GSE236202 | GEO | 2023/06/30

REPOSITORIES: GEO

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Project description:Treatment of the seeds. The Solanum lycopersicum var. Money Maker wild type (MM) and fri1-1 mutant seeds were incubated on distilled-water saturated cotton wool in clear plastic boxes wrapped in black plastic sheets in darkness during 3 hours at 25°C. After that, the seeds were treated as follows: a) 24 hr of continuous irradiation with far-red light, FRc; b) 24 hr of FRc follow by a 10 min red light pulse (Rp), FRc+Rp; and c) complete darkness. After the light treatments, the seeds were incubated in darkness (25°C) until sampling. Red light (35umol/m2sec) was provided by 40W fluorescent lamps (Phillips 40/15) in combination with a red translucid acrylic. FRc light (100 umol/m2sec) was provided by a set of 150W incandescent internal reflector lamps (Phillips) in combination with a red acetate filter, six 2mm thick blue acrylic filters Paolini 2031 and 10cm water filter. Total RNA isolation. The seeds were grinded in liquid nitrogen and extracted with two volumes of extraction buffer (1% SDS, 6% p-aminosalycilic acid, 1% NaCl, 6% water-saturated phenol and 2% 2-mercaptoethanol) and two volumes of acid phenol:chloroform solution. The aqueous phase was sequentially extracted with one volume of acid phenol:chloroform solution and one volume of chloroform. The nucleic acids in the aqueous phase were precipitated with 0.15 volumes of 3M NaCl and 2.5 volumes of 100% ethanol at –80°C for 20 minutes. After centrifugation, the pellet was washed with 70% ethanol, resuspended in RNase-free water and precipitated with one volume of 4M LiCl over night at 4°C. The LiCl was removed after centrifugation and the pellet was rinsed with 2M LiCl and resuspended in RNase-free water. The total RNA was treated with RNase-free DNase (Promega) and cleaned up with mini spin columns (RNeasy Plant Mini Kit, Qiagen). The mRNA was amplificated with Message Amp II aRNA Amplification Kit (Ambion) and concentrated by vacumm centrifugation. Keywords: Direct comparison 21 hybs total

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DNA methylation array of irradiated and/or lithium exposed iPSC-derived human neural progenitor cells

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