For SGBS cells, siRNA transfections were conducted using the Neon Transfection System 100 µL Kit (Invitrogen, Darmstadt, Germany) with a pulse voltage of 1300 V, pulse width 20 ms, pulse number 2 and a cell density of 6 × 10[6 (link)] cells/ml [45 (link)]. A final concentration of 500 nM of AHCY-specific ON-TARGETplus SMARTpool siRNA or ON-TARGETplus control siRNA (Dharmacon, Lafayette, LA, USA) were used. After electroporation, SGBS cells were seeded at a final concentration of 1,000 cells/well in a 96-well format for proliferation analyses and at 100,000 cells/well in a 12-well format for adipocyte differentiation assay. Knockdown efficiency was confirmed using q-PCR and AHCY activity assay.
Protocol detail
SGBS Cell Knockdown of AHCY
Partial Protocol Preview
This section provides a glimpse into the protocol.
The remaining content is hidden due to licensing restrictions, but the full text is available at the following link:
Access Free Full Text.
Corresponding Organization :
Other organizations : University Hospital Leipzig, Helmholtz Zentrum München, Leipzig University
Top 5 similar protocols
Variable analysis
independent variables
- SiRNA transfection
- Pulse voltage (1300 V)
- Pulse width (20 ms)
- Pulse number (2)
- Cell density (6 × 10^6 cells/ml)
- SiRNA concentration (500 nM)
- Cell proliferation
- Adipocyte differentiation
- AHCY mRNA expression (knockdown efficiency)
- AHCY enzyme activity
- Neon Transfection System 100 µL Kit (Invitrogen, Darmstadt, Germany)
- ON-TARGETplus control siRNA (Dharmacon, Lafayette, LA, USA)
- Positive control: ON-TARGETplus SMARTpool siRNA targeting AHCY
- Negative control: ON-TARGETplus control siRNA
Annotations
Based on most similar protocols
Etiam vel ipsum. Morbi facilisis vestibulum nisl. Praesent cursus laoreet felis. Integer adipiscing pretium orci. Nulla facilisi. Quisque posuere bibendum purus. Nulla quam mauris, cursus eget, convallis ac, molestie non, enim. Aliquam congue. Quisque sagittis nonummy sapien. Proin molestie sem vitae urna. Maecenas lorem.
Sign up for free or login to display all annotations
As authors may omit details in methods from publication, our AI will look for missing critical information across the 5 most similar protocols.
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!