Lentiviral Transduction of Breast Cancer Cells

CRYβB2 coding sequence was cloned into a lentivirus vector (Addgene) using the Gateway Technology System (Thermo Fisher). MCF10A, MCF10AT1, and DCIS.COM cells overexpressing luciferase and CRYβB2 were generated following lentivirus infection. For immunofluorescence, MCF10AT1 cells were infected with lentivirus containing the CRYβB2 sequence tagged with the myc-DDK (flag) sequence (Origene). For CRISPR knockout, nucleolin and CRYβB2 guide RNAs were designed using sgRNA online web page from Broad Institute and cloned into Lenticrispr V2 (Addgene). 293T cells were transfected with the lentivirus constructs using Lipofectamine and viruses were used to infect cancer cells [50 (link)].

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.

Yan Y., Narayan A., Cho S., Cheng Z., Liu J.O., Zhu H., Wang G., Wharram B., Lisok A., Brummet M., Saeki H., Huang T., Gabrielson K., Gabrielson E., Cope L., Kanaan Y.M., Afsari A., Naab T., Yfantis H.G., Ambs S., Pomper M.G., Sukumar S, & Merino V.F. (2021). CRYβB2 enhances tumorigenesis through upregulation of nucleolin in triple negative breast cancer. Oncogene, 40(38), 5752-5763.

Publication 2021

lentivirus vector Lenticrispr V2

293t cells Cancer Cells Coding sequence Crispr Dcis Immunofluorescence Lentivirus Lentivirus infection Lipofectamine Luciferase Nucleolin Rnas Vector Viruses

Corresponding Organization :

Other organizations : Johns Hopkins Medicine, Johns Hopkins University, Huazhong University of Science and Technology, Union Hospital, Howard University, Veterans Health Administration, National Institutes of Health, National Cancer Institute

Top 5 similar protocols

Variable analysis

independent variables

Overexpression of CRYβB2 in MCF10A, MCF10AT1, and DCIS.COM cells
CRISPR knockout of nucleolin and CRYβB2

dependent variables

Cell phenotypes

control variables

Overexpression of luciferase in MCF10A, MCF10AT1, and DCIS.COM cells
Transfection of 293T cells with lentivirus constructs

positive controls

Overexpression of luciferase in MCF10A, MCF10AT1, and DCIS.COM cells

negative controls

Transfection of 293T cells with lentivirus constructs

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!