tf dna binding followed by emsa and deep sequencing

... Protocols for the preparation and hybridization of microarrays can be found in Additional file EMSA- Seq (TF- DNA binding followed by EMSA and deep sequencing) We chose DNA ligands that were 60-mers in ... each one a potential binding site) as DNA ligands in an EMSA assay, followed by DNA extraction, library preparation and deep sequencing of the DNA fraction that has been bound by a transcription ... DNA complex b EMSA free DNA Electrophoretic Mobility Shift Assay (EMSA) Protein -DNA Binding microarrays DNA- sequences bound by Transcription Factors (TFs) in vitro high binding affinity low binding...

Ngày tải lên: 09/08/2014, 23:20

... DH5a bacteria and mutations were confirmed using an automated DNA sequencing (Perkin-Elmer) Expression and purification of recombinant proteins Recombinant proteins were expressed and purified essentially ... cysteine and 0.3 mM cystine and refolding of the protein was accomplished by extensive dialysis against the same buffer Free cysteine residues were then blocked with iodoacetamide, followed by dialysis ... difluoride)-membrane and subjected to Western blot using MoAb 44 (A) Lane and show the Western blot, whereas lane and show the corresponding silver stained gels of two different C4BP preparations (lanes and 3, and...

Ngày tải lên: 17/03/2014, 09:20

... protein DnaA and this interaction is essential for DNA replication in Helicobacter [14,15] One protein that is central to the DNA replication, repair and recombination is single-stranded DNA binding ... towards ssDNA over dsDNA M13mp18 ssDNA or pUC18 dsDNA alone or a mixture of both was incubated either in the absence or presence of a different amount of HpSSB followed by separation of the DNA protein ... single-stranded DNA binding protein (T4 gp32) complexed to DNA Nature 376, 362–366 17 Webster G, Genschel J, Curth U, Urbanke C, Kang C & Hilgenfeld R (1997) A common core for binding single-stranded DNA: ...

Ngày tải lên: 30/03/2014, 02:20

... capping functions and that regulate telomere length [3] The TRF1, TRF2 and Pot1 subunits bind to telomeric DNA and to the other subunits of the complex, namely the TIN2, TPP1 and Rap1 proteins ... at the level of telomere itself, by several shelterin subunits, including TRF1, TIN2, TPP1, Pot1 and Rap1 For instance, Pot1, a single-stranded telomeric DNA- binding protein, behaves as a terminal ... were isolated, reverse transcribed and cDNA were analyzed by qPCR using primers for hTERT and PBGD Results are expressed as indicated in legend of table Standard deviations are from at least...

Ngày tải lên: 13/08/2014, 09:21

... when grammar rules change! Sign up to our newsletter here: englishgrammar.org (It's free) Powered by TCPDF (www.tcpdf.org) ...

Ngày tải lên: 17/07/2015, 20:31

... from the low-afnity and nonspecic DNA- binding state to the state characterized by high afnity and sequence specicity ể FEBS 2003 Conformational changes induced by cAMP and DNA binding to CRP (Eur ... apo-CRP structure induced by binding of cAMP and DNA by measuring uorescence resonance energy transfer (FRET) and uorescence anisotropy decay Two tryptophan residues, Trp13 and Trp85, were used as ... F and Fcor are uorescence intensity before and after the correction, and P and DA denote the initial sample absorbance at the excitation wavelength and the change in absorbance introduced by...

Ngày tải lên: 20/02/2014, 23:20

... forming the oligomeric structure and for DNA binding, by cloning an hbpR gene devoid of the A-domain, purifying this protein, and analyzing its DNA- binding characteristics and its activation capacity ... nucleotides and nucleotide motifs was characterized by site-directed mutagenesis and affinity binding by purified HbpR In several cases, DNaseI footprints were conducted to confirm the binding site ... of the binding region was not influenced by the change from four AT to four or six GC, was shown by mutants and (each with 3· GC and five mutations) Both mutants and were bound equally well by HbpR...

Ngày tải lên: 07/03/2014, 17:20

... M.SssI binding to DNA, but reduced M.HhaI binding by 1–2 orders of magnitude (Fig 6) Therefore, the bulky B[a]P residue positioned in the DNA minor groove severely inhibits DNA binding to M.HhaI by ... the DNA A Fig Bar graphs representing relative Kd (K rel ) and kcat (k rel ) values for binding and d cat methylation of DNA containing (+)-cis-B[a]PN2-dG and (+)-trans-B[a]P-N2-dG adducts by ... residues in the DNA duplexes, and are consistent with a base-flipping model of the dC target residue Binding and methylation studies The interactions of M.HhaI and M.SssI with DNA containing site-specifically...

Ngày tải lên: 19/02/2014, 00:20

... thymus DNA, and this was followed by electrophoresis on 5% polyacrylamide gel Lane contains only DNA without any protein; lanes 2, and correspond to DNA complexes with p53(1–363), p73d and p73b, ... one strand and two AG steps in the other The mdm2 target, whose interaction with p53 was most strongly affected by DNA cis-platination, contains GG, GGG and AG motifs in one strand and GG and GTG ... 0.06, all mdm2, PGM1 and p21 targets exhibited very weak p53 binding (about 4% for mdm2 and PGM1 and 10% for p21) Sensitivity of the sequence-specific p53 DNA binding to DNA cis-platination depends...

Ngày tải lên: 19/02/2014, 05:20

... 2-mercaptoethanol, and 10% (v ⁄ v) glycerol, and the flow rate was 0.5 mLÆmin)1 CD measurements DNAs The /X174 phage ssDNA and dsDNA used in the DNA binding and ATPase assays were purchased from New England ... these loops are actually involved in DNA binding Because aromatic residues are involved in the ssDNA binding by bacterial single stranded DNA- binding protein (SSB) and human replication protein A ... should affect the DNA binding of the protein The DNA- binding ability of these alanine-substituted Rad51 mutants was evaluated by a gel retardation technique and by measurements of the DNA- dependent...

Ngày tải lên: 19/02/2014, 06:20

... equilibrium binding of CI to O1 and O2 DNA respectively (J) Cooperative binding: the operator DNA contents in the shifted complexes and and in the unbound labeled O1O2 DNA were determined by scanning ... of O1 DNA Synthesis of O2 and O1O2 DNAs Synthesis of O2 DNA Synthesis of O3 DNA Synthesis of O3 DNA Synthesis of S aureus cspC DNA Synthesis of S aureus cspC DNA (Fig 2A) was synthesized by PCR ... O1, and 49 bp O2 DNAs (Fig 2A) were prepared by mixing and annealing primers PCR11 and PCR21 and IIa and IIb, respectively (Table 1) The cooperativity parameter for the binding of CI to O1O2 DNA...

Ngày tải lên: 07/03/2014, 00:20

... 15 Bandshifts were analyzed by subsequent electrophoresis on a 5% polyacrylamide gel Gels were run at 100 V for h, and dried, and products were visualized by autoradiography and quantified by a ... and then cleaved with EcoRI Restricted fragments A and B (to delete site I), C and D (to delete site II), E and F (to delete site III), G and H (to delete the perfect inverted repeat) and I and ... that the helix–turn–helix DNA- binding domains of SenR and ChrA share 61% amino acid identity (data not shown) The designed SenRD60 and SenRD65A proteins showed reduced DNA- binding affinity for up-furS...

Ngày tải lên: 07/03/2014, 10:20

... examine whether the DNA binding activity of dHAND could be affected by Akt phosphorylation The influence of phosphorylation by Akt on the DNA binding activity was examined by gel shift analysis ... heterodimers between dHAND mutants and ME2 were compared to that of wild type dHAND using probe E Protein amounts of dHAND, dHANDAsp-X, and dHAND-Asp used in this experiment were estimated by Western blotting ... Fig 8C, dHAND-Asp-X and -Asp almost lost their DNA binding activity These results indicated that the phosphorylation of Thr102, Thr112, and Ser114 inhibited DNA binding activity of dHAND In this...

Ngày tải lên: 07/03/2014, 16:20

... sequence boxes I-III (CSB), tRNA genes, L and H strand promoters (LSP and HSP), L and H strand transcription start sites (LSS and HSS) The L-strand sequence and position of the putative promoter ... OL and OH represent the origin of L and H strand replication, respectively LSS, L strand start site; HSS, H strand start site The map positions of the rRNA, tRNAs and mRNAs (ND1 to ND6, A6 and ... signal AATAAA, and also the sequences upstream and downstream of the canonical polyadenylation signal are important for protein binding to D-TERM DNA The binding specificity of D-TERM DNA and also the...

Ngày tải lên: 08/03/2014, 08:20

... analyzed by HPLC and the molecule induced by the viral infection was detected by MALDI-TOF MS (Fig 2B) The molecule was purified to homogeneity by gel permeation and reversed-phase chromatography and ... gradient and analyzed by electron microscopy as described [22] Briefly, 4000 flies were injected with DCV and collected and frozen after death Flies were crushed in 10 mM Tris/HCl (pH 7.5), followed by ... containing two layers of caesium chloride (5% and 40%), and ultracentrifuged for 16 h at 36 000 g at 15 °C The virus band was collected and DCV was recovered by centrifugation (25 000 g, h 30 min, 15...

Ngày tải lên: 08/03/2014, 08:20

... 30 and harvested by centrifugation for 10 at 2000 g Cells were solubilized by boiling in SDS sample buffer for and the extracted proteins were resolved by means of 7.5% SDS/ PAGE followed by ... lysed and the LBP was immunoprecipitated by anti-LBP Ig and autoradiographed (lane 6) Indirect immunoblotting revealed a 67-kDa protein band using laminin as the primary probe followed by treatment ... destroyed by heat denaturation (100 °C, min) indicating that the conformation of both the receptor and ligand are essential for binding Changes in pH of the binding buffer also had marked effect on binding...

Ngày tải lên: 08/03/2014, 23:20

... results of filter binding studies and electrophoretic mobility shift studies of RNA binding Direct comparison of the binding of BcBL1 and BcBL2 to B caldolyticus PyrR by the filter binding and gel shift ... non-specific binding to RNA cannot be excluded The binding of BcBL2 was described by a single tight binding curve although, in the presence of 0.5 mm GMP, the binding curve was broad and fitted less ... is favored by binding of either uridine nucleotides or binding of RNA itself in the case of BcBL1 The low affinity state is favored by the binding of guanosine nucleotides Thus, RNA binding involves...

Ngày tải lên: 16/03/2014, 06:20

... test) protein that binds single-stranded nucleic acids and is able to promote interstrand reannealing of DNA as well as RNA [35,36] hnRNP Al is phosphorylated by CK2 and PKA PKA phosphorylation suppresses ... single-strand DNA binding protein could promote or restrict the interaction of a conventional transcription factor with the promoter Thus, single-strand loop formation directed by constitutive and ... Radioactive bands were quantified and annealing percentage was determined by dividing the amount of annealed oligonucleotide (ds) by the total amount [annealed plus single-stranded (ss)] in each lane, and...

Ngày tải lên: 16/03/2014, 12:20

... of mM ATP (lane 6–10) and analysed by native PAGE (6% acrylamide) followed by silver staining Lane and has hRad51 (10 lM) without DNA and lane 11 had hRad52 (10 lM) with DNA The position of asterisk ... analyse DNA induced conformational changes on hRad51 protein hRad51 (25 lM) was incubated in binding buffer in the absence (lanes and 3) or presence of 75 lM ssDNA (lanes and 5) and mM ATP (lanes and ... cofactors and later subjected to centrifugation and the resulting pellet and supernatant were analysed by SDS PAGE followed by silver staining (C) Dynamic light scattering to study the effect of DNA...

Ngày tải lên: 16/03/2014, 14:20

... DNA binding In order to shed light on the specific character of DNA adducts of trans-PtHMP, we further examined the DNA binding properties of this new transplatin analogue and compared these binding ... prepare easily and precisely the samples of DNA modified by the platinum complex at a preselected value of rb The samples of DNA modified by new platinum compound and analyzed further by biophysical ... transcribed DNA strand in vitro is not affected by a nearby lesion in the opposite strand J Biol Chem 278, 19558–19564 28 Corda Y, Job C, Anin MF, Leng M & Job D (1991) Transcription by eucaryotic and...

Ngày tải lên: 16/03/2014, 14:20