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Dose-dependent and quantitative PCR amplifications of each fraction obtained from the ChIP procedure were performed.
The ChIP procedure was carried out following the manufacturer's instructions (Upstate Biotechnology) with some modifications.
The ChIP procedure was performed as described previously [39] with AGS cells grown to 70 80% confluence.
The chromatin immunoprecipitation (ChIP) procedure was performed using a modification of an assay kit (Upstate Biotechnology, Charlottesville,VA) according to the manufacturer's instructions [28].
To investigate whether holo TFIIH specifically binds to UV-damaged DNA in vivo, we performed a modified ChIP procedure, which was recently used for detecting the specific binding of yeast Rad26, human CSA and CSB to DNA lesions [13], [44].
After centrifugation, the supernatant was diluted 20 times with re-ChIP buffer (1% Triton X-100, 2 mM EDTA, 150 mM NaCl, 20 mM Tris-HCl, [pH 8.1]) and subjected again to the ChIP procedure.
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At least four 15 cm dishes of cells were used per chromatin immunoprecipitation (ChIP) and samples were processed according to standard ChIP procedures [ 36].
ChIP procedures were performed as described previously [ 68] with at least duplicate biological samples (Hmt1-myc; n=2, pan-acetyl-H4 and H3K36me2; n=3).
The chipping procedure was carried out in March 2009 using on the same header both the standard chipping drum and a special on purpose drum designed by CRA-ING to obtain coarser particles.
For Hrp1 and Hrp3 binding studies we used the ChIP-chip procedure described by Kurdistani et al. (2002) [33].
To provide evidence that our seq-ChIP-chip procedure resulted in enrichments coming from both ChIPs when assayed sequentially, we also performed two control seq-ChIP-chip experiments.
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