"Regulatory" sequences in lacZ?

Iain Cartwright Iain.Cartwright at UC.EDU
Wed Oct 11 18:02:54 EST 1995


I am wondering if anyone has ever noticed unexpected transcriptional
activating abilities associated with transgenic lacZ genes (actually the
approximately 3 kb of lacZ coding sequences)?  We have generated numerous
lines of fly in which a minimal luciferase reporter (just a TATA box fused
to the luciferase gene) is joined immediately upstream of the TATA sequence
to the E. coli lacZ sequence.  When the lacZ gene (which is not hooked up to
a eukaryotic promoter and is not expressed - don't ask why we constructed
this, that's another story) is cloned in the divergent orientation relative
to luciferase (i.e. its 5' end adjacent to TATA) there is no, or very
little, expression of the luciferase reporter.  However, when the lacZ
sequence is inverted (so that its 3' end is now adjacent to the luciferase
TATA box) there  are high levels of luciferase activity (up to 100-fold
greater than in the previous orientation).  This effect would not seem to be
typical of a classical  enhancer, but acts as if the 3' end of the lacZ
coding sequence contains some cryptic promoter proximal-specific activating
sequence (we don't yet know if there is any tissue specificity to the
effect!).  This has been a consistent observation in up to ten different
transgenic lines of each type.  Perhaps interestingly, there is less than a
two-fold effect when testing the two constructs transiently in tissue
culture cells.  We could actually investigate this phenomenon further I
suppose, but I'd be more interested in hearing from anybody who has had
similar or related experiences, and who perhaps even knows where such a
cryptic sequence element might be located.  Thanks for reading this far!

Iain Cartwright




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