Lentiviral Knockdown of Human RAD51

Lentivirus particles producing either non-targeted control (C) shRNAs or those targeting human RAD51 (R) were purchased from Sigma Chemical Co., St Louis, MO, USA. Cells (2×10⁵ per well) were seeded into 24-well plates one day before transduction. On the day of transduction, medium was replaced with fresh medium containing hexadimethrine bromide at a final concentration of 8 μg/ml. Lentiviral particles (25 μl) were added to each well, mixed and incubated at 37 °C in a humidified incubator with 5% CO₂. After 16 h, the virus-containing medium was replaced with fresh medium and cells maintained in culture. On the next day, cells were trypsinized and transferred to 25 cm² flasks. After another 48 h, puromycin was added to the medium at a final concentration of 1 μg/ml. Suppression of HsRAD51 protein expression was confirmed by western blotting after 7 days of puromycin selection.

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.

Pal J., Bertheau R., Buon L., Qazi A., Batchu R., Bandyopadhyay S., Ali-Fehmi R., Beer D., Weaver D., Reis R.S., Goyal R., Huang Q., Munshi N, & Shammas M. (2011). Genomic evolution in Barrett’s adenocarcinoma cells: critical roles of elevated hsRAD51, homologous recombination and Alu sequences in the genome. Oncogene, 30(33), 3585-3598.

Publication 2011

Cells Hexadimethrine bromide Hsrad51 protein Human Lentivirus Puromycin Shrnas Virus

Corresponding Organization :

Other organizations : Dana-Farber Cancer Institute, The Barbara Ann Karmanos Cancer Institute, Wayne State University, University of Michigan–Ann Arbor, Central Arkansas Veterans Healthcare System, University of Arkansas for Medical Sciences, VA Boston Healthcare System, Harvard University

Top 5 similar protocols

Protocol cited in 7 other protocols

Variable analysis

independent variables

Lentivirus particles producing either non-targeted control (C) shRNAs or those targeting human RAD51 (R)

dependent variables

Suppression of HsRAD51 protein expression

control variables

Cells (2×10^5 per well) were seeded into 24-well plates one day before transduction
On the day of transduction, medium was replaced with fresh medium containing hexadimethrine bromide at a final concentration of 8 μg/ml
Lentiviral particles (25 μl) were added to each well, mixed and incubated at 37 °C in a humidified incubator with 5% CO2
After 16 h, the virus-containing medium was replaced with fresh medium and cells maintained in culture
On the next day, cells were trypsinized and transferred to 25 cm2 flasks
After another 48 h, puromycin was added to the medium at a final concentration of 1 μg/ml

positive control

Non-targeted control (C) shRNAs

negative control

None specified

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.

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?

Revolutionizing how scientists
search and build protocols!