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Fig. 3. HP1 associates with the silenced hsp26 and hsp70 reporter
genes. Chromatin immunoprecipitation experiments were performed to determine
whether the lac-hsp26-hsp70 transposon in stocks carrying the
lacI-HP1 expressor and the transposon (+ lacI-HP1) or the reporter transposon
alone (- lacI-HP1) was associated with HP1. Polyclonal HP1 antibodies were
used and polyclonal GFP antibodies served as a negative control. (A) Primer
sets corresponding to unique sequences within the reporter genes were used to
PCR amplify the immunoprecipitated material. The amount of immunoprecipitated
material (designated % input) was quantitated by dividing its signal intensity
by the signal intensity generated from a 1:100 dilution of the input material.
(B) The limits of HP1 spreading were determined using primer sets
corresponding to sequences over the reporter transposon in stock hsp26-4D5.
Primers corresponding to sequences located 1.9 (hsp26), 3.7
(hsp70) and 10.1 kb from the 3' end of the lac repeat
array and 0.5 and 3.1 kb from the 5' end of the lac repeat
array were assayed for HP1 association. (C) The amount of HP1 associated
material (% input) determined from immunoprecipitation with anti-HP1
antibodies, with or without lacI-HP1 expression, was compared at each primer
site (n=3). A probability (P value) of less than 0.05 was
considered to be a significant change.
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