The line is available with the following restriction: 1. The cell line was deposited at the ATCC by Rockefeller University and is provided for research purposes only. Neither the cell line nor the products derived from it may be sold or used for commercial purposes. Nor can the cells be distributed to third parties for purposes of sale, or producing for sale, cells or their products. The cells are provided as a service to the research community. They are provided without warranty of merchantability or fitness for a particular purpose or any other warranty, expressed or implied. 2. Any proposed commercial use of the cells, or their products, must first be negotiated with Cell Genesys, 500 Forbes Boulevard, South San Francisco, CA 94080 Attn: Robert H. Tidwell; Senior Vice President, Corporate Development.

SV40 T antigen [45408]

The 293T/17 cell line is a derivative of the 293T (293tsA1609neo) cell line. 293T is a highly transfectable derivative of the 293 cell line into which the temperature sensitive gene for SV40 T-antigen was inserted. 293T cells were cloned and the clones tested with the pBND and pZAP vectors to obtain a line capable of producing high titers of infectious retrovirus, 293T/17. These cells constitutively express the simian virus 40 (SV40) large T antigen, and clone 17 was selected specifically for its high transfectability. 293T/17 cells were cotransfected with the pCRIPenv- and the pCRIPgag-2 vectors to obtain the ANJOU 65 (see ATCC CRL-11269) cell line. ANJOU 65 cells were cotransfected with the pCRIPgag-2 and pGPT2E vectors to obtain the BOSC 23 (see ATCC CRL-11270) ecotropic envelope-expression packaging cell line. ANJOU 65 cells were also cotransfected with the pCRIPAMgag vector along with a plasmid expressing the gpt resistance gene to obtain the Bing (see ATCC CRL-11554) amphotropic envelope-expression packaging cell line.

Protocol:

1. Remove and discard culture medium.
2. Briefly rinse the cell layer with 0.25% (w/v) Trypsin- 0.53 mM EDTA solution to remove all traces of serum that contains trypsin inhibitor.
3. Add 2.0 to 3.0 ml of Trypsin-EDTA solution to flask and observe cells under an inverted microscope until cell layer is dispersed (usually within 5 to 15 minutes).
   Note: To avoid clumping do not agitate the cells by hitting or shaking the flask while waiting for the cells to detach. Cells that are difficult to detach may be placed at 37°C to facilitate dispersal.
4. Add 6.0 to 8.0 ml of complete growth medium and aspirate cells by gently pipetting.
5. Add appropriate aliquots of the cell suspension to new culture vessels.
6. Incubate cultures at 37°C.

Subcultivation ratio: A subcultivation ratio of 1:4 to 1:8 is recommended

Medium renewal: Every 2 to 3 days

Recommended medium (without the additional supplements or serum described under ATCC Medium): ATCC 30-2002
recommended serum: ATCC 30-2020
derivative: ATCC CRL-11269