IPSCs were karyotyped in metaphase by the Shivanand R. Patil Cytogenetics and Molecular Laboratory at the University of Iowa using Leica Microsystems metaphase scanning platform and CytoVision version 7.7 software. Cells were grown in vitro and arrested at metaphase with colcemid. Chromosomes were stained by the G-banding method, counted and structurally evaluated for the presence or absence of detectable rearrangements. At least 20 cells were analyzed for each iPSC line. For TaqMan hPSC ScoreCard™ analysis, total RNA was isolated using the NucleoSpin RNA purification kit (Takara Bio, San Jose, CA). cDNA was generated from 1 µg of RNA using VILO cDNA synthesis kit (Thermo Fisher Scientific). cDNA was added to a TaqMan hPSC scorecard plate (Thermo Fisher Scientific) and amplified using a QuantStudio 6 Flex real-time PCR system. Results were analyzed using Thermo Fisher’s cloud-based analysis suite.
Nucleospin rna purification kit
Manufactured by Takara Bio
Sourced in United States
The NucleoSpin RNA purification kit is a product designed to efficiently isolate and purify RNA from a variety of sample types, including cells, tissues, and body fluids. The kit utilizes a silica-membrane technology to capture and wash RNA, while removing contaminants and inhibitors. The purified RNA can be used in various downstream applications, such as RT-PCR, Northern blotting, and RNA sequencing.
Automatically generated - may contain errors
Lab products found in correlation
Quantstudio 6 flex real time pcr system, by Thermo Fisher Scientific (2 mentions)
Vilo cdna synthesis kit, by Thermo Fisher Scientific (1 mentions)
Abi prism 7900ht, by Thermo Fisher Scientific (1 mentions)
Kicqstart sybr green qpcr readymix, by Merck Group (1 mentions)
Rnaiso plus, by Takara Bio (1 mentions)
Agilent 2100 bioanalyzer, by Agilent Technologies (1 mentions)
4 protocols using nucleospin rna purification kit
1
iPSC Karyotyping and Pluripotency Evaluation
2
Assessing Gene Expression by qPCR
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Cells were grown to 80% confluence and then treated with 1 μM I-BET151, 1 μM alisertib, both drugs at 1 μM, or vehicle control for 24 hours. Total RNA was extracted from the cells using NucleoSpin RNA purification kit (Takara Bio USA), and 1 μg of RNA was used for cDNA synthesis using Maxima RT cDNA First Strand Synthesis kit (ThermoFisher Scientific). qPCR was performed using KiCqStart SYBR Green qPCR ReadyMix (Sigma-Aldrich, St. Louis, MO) using the ABI PRISM 7900HT thermal cycler (ThermoFisher Scientific), with relative quantitation by the ddCt method as previously described [31] (link). Experiments were performed with technical duplicates on each plate and in three independent experiments, with the relative expression of each experiment used to calculate expression and standard deviation, plotted on each graph.
3
Measuring SNCA mRNA Stability in HeLa Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
HeLa Tet-On cells were plated and transfected with siSRP54 and pCS2-SNCA plasmid as described above. Cells were treated with DMSO (control) or 8 µg/mL of Actinomycin D 12 h after plasmid transfection for 0, 4, 6, 8, and 10 h. Total RNA was extracted and purified using the NucleoSpin RNA purification kit (Takara Bio USA, 740955) at the indicated time points after treatment. OmpA mRNA (20 ng) was added prior to purification and used for normalization [19 (link)]. Relative mRNA levels were quantified using RT-qPCR, as described below.
4
Cortical RNA Extraction and Purification
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Total RNAs were extracted from the whole ipsilateral cortex with RNAiso Plus (Takara Bio., Japan) and cleaned with NucleoSpin RNA purification kit (Takara Bio., Japan) with on-column DNase I digestion. RNA Integrity Numbers (RIN) was assessed using Agilent 2100 Bioanalyzer (Agilent Technologies, Palo Alto, CA). This indicator was > 8 in all the samples.
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!