I have made a construct in an MSCV back-bone were DsRed and the Neo-r gene,
separated by an IRES, is under the control of the LTR regions. I used
Phoenix cells for the first transfection to get transient viral production,
and made GPG- Am12- and PG13- producer cell lines. All lines were intensely
red by microscope. However, when I try to test the viral titer from the
Am12 and PG13 producer lines on human cell lines (HeLa- and HT1080-cells) I
get no red cells at all. The same vector with eGFP works fine (and
integrates) in HeLa and HT1080 as well as in human CD34+ umbilical cord
blood cells. I have had DsRed transduced HeLa and HT1080 cells going for 14
days and can not se any red cells by microscope or FACS (producerlines are
99% pos. by FACS). (I have just started selection with G418 to make sure
that the cells have actually been transduced).
Does anyone have a good explanation, or a suggestion what to do next?
Thankful for any suggestions!
Ann Brun
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Ann Brun, MSc
Section for Molecular Medicine & Gene Therapy, WNC
Lund University
Sölvegatan 17
S-223 62 LUND, Sweden
Phone +46-46-222 05 92
Fax +46-46-222 05 68
e-mail Ann.Brun at molmed.lu.se
(internpost: hämtställe 57)
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