2) Fig  7 R 2 for regressions of F v/F m(λex,λem) of simulated

2). Fig. 7 R 2 for regressions of F v/F m(λex,λem) of simulated

communities against F v/F m(470,683) and F v/F m(590,683) of respectively algal and cyanobacterial subpopulations. These plots represent cross sections of the excitation–emission regression matrix of Fig. 6: a the 683-nm emission line, b the 470-nm excitation line, and c the 590-nm excitation line. Key excitation–emission BIX 1294 cost pairs are indicated by the numeric markers corresponding to Figs. 6 and 8 The data underlying the optimal excitation/emission pairs identified from Figs. 6 and 7 are presented in Fig. 8 with corresponding regression statistics. Figure 8a confirms that community F v/F m(470,683) is strongly driven by the algal F v/F m and was FHPI concentration highly insensitive to the fluorescence of the cyanobacteria in the simulated communities. Only the case for equal Selleck Mocetinostat absorption in the algal and cyanobacterial subpopulations is shown here, but when the community composition was skewed to 90% in favour of the cyanobacteria, community F v/F m(470,683) remained a good (relative error <10%) predictor of algal F v/F m(470,683) in 92% of cases. The fluorescence emission of the cyanobacterial

fraction was too low at this excitation/emission pair to influence community variable fluorescence, even when mixed with algal cultures of low (variable) fluorescence. Fig. 8 Case plots underlying the linear regression analyses of community F v/F m(λex,λem) versus algal and cyanobacterial F v/F m(470,683) and F v/F m(590,683), respectively. a–c correspond to the key excitation–emission pairs highlighted with numerical markers in Fig. 6. a F v/F m(470,683), sensitive to algal but not cyanobacterial F v/F m, b F v/F m(590,683), with stronger correspondence to cyanobacterial compared to algal F v/F m and c F v/F m(590,650), strongly related to cyanobacterial F v/F m(590,683) >0.4. Colours and symbols correspond to Fig. 7, drawn black lines mark unity. The discrete distribution of the subcommunity F v/F m values is caused by

the limited number of cultures used to simulate community F v/F m matrices Under red–orange illumination centred at 590 nm (Fig. 8b) we note a better correlation of community and cyanobacterial F v/F m (R 2 = 0.54). Farnesyltransferase The relatively low slope and high offset of this regression were clearly caused by the inclusion of cases where cyanobacterial subpopulations with low F v/F m were mixed with algae with higher F v/F m, a result of a wider spread of F v/F m in the cyanobacterial cultures compared to the algae (Fig. 3). The regression results for the algal fraction under emission at 590 nm were clearly worse with R 2 = 0.18. The variable fluorescence originating from PBS pigments (F v/F m(590,650)) was lower than F v/F m(590,683) while the relation between community and cyanobacterial F v/F m was strong for cyanobacteria cultures with F v/F m >0.42 (Fig. 8c).

The shape and properties of the synthesized particles are highly

The shape and properties of the synthesized particles are highly dependent on the starting material used in the alkaline precipitation method (i.e., nitrates vs. chlorides vs. sulfates) [7]. However, thermal decomposition suffers from the drawback of using relatively toxic precursors in the syntheses. Thermal decomposition methods use toxic metallic precursors such as iron pentacarbonyl (Fe(CO)5) and other organic solvents for the process of synthesis [1, 4, 7]. There is much interest currently in alternative methods of nanoparticle synthesis, which use relatively non-toxic starting precursors and are environmentally friendly. It is now possible to prepare nanoparticles using

much less toxic chemical precursors, such as iron fatty acids [2, 8–10]. These so-called green synthesis methods are much less toxic CH5424802 price and can produce relatively stable and uniform magnetic nanoparticles [8, 10]. Superparamagnetic iron-platinum particles (SIPPs) produced using such methods are seen to maintain their relative stability in solutions [2, KU55933 order 8, 9]. Uniformity of size and shape of nanoparticles are important for issues related

to biocompatibility as a widely Ilomastat varying size range may lead to non-uniform behavior of the nanoparticles both in vitro and in vivo [11]. The general reaction for the synthesis of magnetic nanoparticles using a green method of synthesis is described as follows. The iron precursor of

the reaction is in the form of iron fatty acids (Fe-fatty acid). The second component of the bimetallic nanoparticle is a platinum precursor in the form of platinum acetylacetonate or Pt(acac)2. The solvent of the reaction is octadecene (ODE) or tetracosane (TCA). A fourth component of the reaction is the use of fatty amines and fatty acids as ligands. Fatty amines, in the form of octadecylamine (ODA), are carbon-18 single chain fatty amines that play a critical role in the stabilization of the nanocrystal in the early stages of synthesis [10]. Moreover, fatty amines can act as both the solvent and the ligand, reducing the number of chemicals needed to produce the alloy Calpain nanocrystals. In this report, we focus on the open question of the role played by the fatty amine in the formation of the bimetallic FePt nanocrystal. More specifically, we compare the effect of varying lengths of fatty amine ligands on the shape, structure, uniformity, composition, and magnetic properties of the synthesized magnetic FePt nanoparticles. Methods Materials used for synthesis Iron nitrate nonahydrate (Fe(NO3)3 · 9H2O) and Pt(acac)2 were purchased from Sigma (St. Louis, MO, USA). Additionally, all of the ligands including ODA, 1-hexadecylamine (HDA), 1-tetradecylamine (TDA), and 1-dodecylamine (DDA) were purchased from Sigma (St.

See Figure 1 for abbreviations Further kinetic analysis showed t

See Figure 1 for abbreviations. Further kinetic analysis showed that the K m value towards NAM was 5.81 mM, and the V max was at

400 nmol/min/mg protein. The kinetic data indicated that xapA in E. coli was much less efficient in using NAM to synthesize NR than using typical substrate (K m at 5.81 mM on NAM vs. 72 μM on xanthosine) [37], or when compared with other NAD+ salvaging enzymes (e.g., K m values at 70 μM and 2 μM for pncA and pncB on NAM and NA, respectively) [39, 40], but similar to those of deoD www.selleckchem.com/products/gs-9973.html (PNP-I) from calf and E. coli (i.e., 1.48 mM and 0.62 mM, respectively) in converting the non-typical substrate NR to NAM [38]. The contribution of xapA in NAD+ salvaging was also confirmed in bacterial mutants cultured in M9/NAM medium, in which the consumption of extracellular NAM by the triple-deletion (BW25113ΔnadCΔpncAΔxapA) was reduced by 95% in comparison to that by the double-deletion BW25113ΔnadCΔpncA (Figure 5A). The consumption of extracellular NAM was restored when vector expressing xapA (but not the EGFP control) was reintroduced to the triple-deletion (Figure 5A). The level of intracellular NAD+ was detectable in BW25113ΔnadCΔpncA (150 ng), GF120918 ic50 but virtually undetectable in BW25113ΔnadCΔpncAΔxapA (Figure 5B). Again, the intracellular NAD+ level could be restored by reintroducing xapA into the triple-deletion,

but not by EGFP (Figure 5B). Figure 5 Consumption of extracellular NAM (A) to form intracellular NAD + (B) by four strains of Escherichia many coli derived from BW25113 cultured in M9/NAM medium until the strain BW25113Δ nadC Δ pncA reached the mid-log phase. Strain 1, BW25113ΔnadCΔpncA; strain 2, BW25113ΔnadCΔpncAΔxapA; strain 3, BW25113ΔnadCΔpncAΔxapA/pBAD-xapA;

and strain 4, BW25113ΔnadCΔpncAΔxapA/pBAD-EGFP. Discussion Contribution of xapA to an alternative NAD+ salvage pathway from NAM Xanthosine phosphorylase (xapA, EC is a second purine nucleoside phosphorylase (PNP-II) in E. coli. Similar to PNP-I (deoD), it mainly functions in the purine metabolism by carrying out both phosphorylation and synthesis of purine and purine deoxy-/ribonucleosides [41]. Here we first obtained genetic evidence that xapA was probably involved in NAD+ salvage in E. coli. We also provided more direct biochemical evidences that xapA was able to synthesize NR from NAM. Both bacterial growth experiments and enzyme kinetic data indicated that xapA used NAM in a much less efficient way than using its typical substrates (i.e., purine analogs), suggesting that NAM ACP-196 served only as a non-typical substrate, which was comparable to the PNP-I. Therefore, the capability to convert NAM to NR appeared to be a “side effect” for xapA. However, such a side-effect was sufficient to maintain the survival of E. coli by feeding NAM into the salvage pathway III when all other NAD+ synthetic pathways were unavailable and only NAM was present in the minimal medium.

Full details of the methods are given in Additional File 3 The e

Full details of the methods are given in Additional File 3. The expression of tight junction-related genes differentially expressed from the microarray analysis was confirmed using qRT-PCR. The expression of seven target genes relative to three reference genes was assessed using the standard curve method. The reference genes (GAPD, SDHA and YWHAZ) were chosen based on the findings Selleckchem URMC-099 of Vandesompele et al [52] and their log ratios in the microarray data (close to 1; not differentially expressed). Five target genes (ZO-1, ZO-2,

OCLN, CGN and ACTB) were chosen from the tight junction-related genes that were differentially expressed (all up-regulated) in the microarray analysis. The two other target genes, GJA7 and CLDN3, were chosen to be included because they were down-regulated and not differentially expressed, respectively,

in the microarray analysis. The analysis was carried out as described in Additional File 3 and the data was analysed using Relative Expression Software Tool 2008 (version 2.0.7) with efficiency correction [53]. Fluorescent microscopy Caco-2 cells were grown on Lab Tek II Chamber Slides with Permanox™ coating (Nalge Nunc International Corp, Naperville, IL, USA) for 6 days until confluent. Caco-2 cells were treated with L. plantarum MB452 (OD 600 nm 0.9) or control media for 8 hours (n = 4 per treatment per antibody). After treatment, Caco-2 cells were rinsed twice with PI3K inhibitor PBS, fixed in either 4% (w/v) paraformaldehyde for 20 minutes (for CGN and ZO-1) or ice cold 70% ethanol (for ZO-2 and OCLN), quenched with 50 mM NH4Cl (in PBS) for 15 minutes, and blocked with blocking solution (2%

(v/v) foetal bovine serum, 1% sheep serum albumin, 0.1% Triton X-100, 0.05% Tween 20 in PBS, pH 7.2) for 20 minutes. Caco-2 cells were then immuno-stained with the primary antibodies (2.5 µg/mL rabbit Terminal deoxynucleotidyl transferase anti-ZO-1, 1.25 µg/mL rabbit anti-ZO-2, 2.5 µg/mL rabbit anti-occludin, 1 µg/mL rabbit anti-cingulin; Zymed, Invitrogen, NZ) in blocking solution for 1 hour, followed by a PBS wash (0.1% Triton X-100, 0.05% Tween 20 in PBS) to reduce non-specific staining, and the secondary antibody, Alexa Fluor 488 goat anti-rabbit IgG (5 µg/mL for ZO-2, 10 µg/mL for rest; Invitrogen, NZ) in blocking solution for 1 hour. The slides were imaged with a fluorescent microscope (Leica DM2500 microscope, Leica DFC420C camera) with the following settings: exposure 1.1 ms, saturation 2.25, gamma 1.52, gain 8.4× and magnification 40×. The images were viewed using LAS Image Overlay software (Leica Application Suite v1.8.2). Acknowledgements This work was funded by the AgResearch Internal Investment Fund. RCA is funded by a New Zealand Foundation of Research, Science and Technology Postdoctoral Fellowship (AGRX0602). The authors LY294002 acknowledge the contribution of Kelly Armstrong (fluorescent microscopy) and Paul Maclean (gene ontology and KEGG pathway analysis).

Therefore, replication of all mycoplasma plasmids is likely to be

Therefore, replication of all mycoplasma plasmids is likely to be driven through a rolling-circle mechanism by a Rep protein of the pMV158 family type. Mosaic structure of the mycoplasma plasmids is indicative of recombination events In spite of a conserved structure, multiple pair-wise DNA sequence comparisons indicated that mycoplasma plasmids are in fact a mosaic of rep, dso, copG, and sso blocks. This was evidenced by the occurrence of several local regions of homology detected by using the BLAST program (Figure 5). Pairs of plasmids that show a high level of identity for the Rep sequence (e.g. pKMK1 and check details pMG1B-1; pMG2D-1 and pMG2B-1) do not necessarily share a high degree of identity

for the region upstream of copG. Interestingly, high sequence identity for the region spanning sso was found to be indicative of plasmids being hosted by the same mycoplasma species. For instance, the following plasmid-pairs, pADB201 and pKMK1, pMG1B-1 and pMG2D-1, and pMG2B-1 and pMG2F-2 were isolated from Mmc, Mcc, and M. yeatsii, respectively (Figure 5). This result is consistent

with the fact that during replication this region interacts with chromosome-encoded components [18]. Further degrees of mosaicism were found in particular cases such as for pMG2D-1, in which two putative dso showing sequence heterogeneity are found. Other BMS202 examples of genetic variability are the small size of pBG7AU and the unusual location of the dso in pMG2F-2. Such a mosaic structure is clearly indicative of successive recombination

events between replicons. Figure 5 Analysis of plasmid Rabusertib supplier content of Mycoplasma yeatsii type strain GIH (TS). A. Agarose gel electrophoresis of total DNA. Lanes were loaded after twofold dilution series of the DNA preparation obtained as described in Methods. Bands corresponding to the chromosome and the 2 plasmids are identified. Lane M, DNA ladder. B. Estimated plasmid copy number of pMyBK1 and pMG2B-1 as estimated by gel assay (Panel A) and relative real-time PCR as described in Methods. pMyBK1 is a unique representative of a new replicon family As indicated above, M. yeatsii strain GIH TS was the only strain that yielded a banding pattern of extrachromosomal DNA that suggested the presence of two distinct Lck plasmids (Figure 5A). The small plasmid, pMG2B-1, was shown to belong to the pMV158 family like all other mycoplasma plasmids (Figure 3). In contrast, the larger plasmid (3,432 bp) named pMyBK1 (GenBank Accession number EU429323; [25]) has a genetic organization that sets it apart from the other mycoplasma plasmids. Initial database searches using pMyBK1 sequence as a query indicated low identity with other plasmids and prompted us to further analyze this plasmid that might represent a new family of replicons. First, the relative copy number of each plasmid of M.

Table 1 Characteristic of the patients of four major treatment re

(%) 43(55.1) 108(53.2) 118(54.4) 15(53.6) Female, no. (%) 35(44.9) 95(46.8) 99(45.6) 13(46.4) ECOG, no. (%)

        0 62(79.5) 168(82.8) 175(80.7) 24(85.7) 1 13(16.7) 31(15.2) 35(16.1) 3(10.7) 2 3(3.8) 4(2.0) 7(3.2) 1(3.6) Stage, no. (%)         CP 70(89.7) 184(90.7) 154(71.0) 21(75.0) AP 6(7.7) 12(5.9) 25(11.5) 4(14.3) BC 2(2.6) 7(3.4) 38(17.5) 3(10.7) Interval since diagnosis, mo         Median 0.5 28 13 7.5 Range 0-2 0-96 0-116 2-36 White-cell count (× 109/L)         Median 25.6 31.2 28.9 21.2 Range 2.2-667 7.5-540 11.2-760 9.0-350 Hemoglobin AZD5153 nmr (× g/L)         Median 120 123 115 128 Range 68-177 56-170 66-188 70-175 Platelet count (× 109/L)         Median 345 485 520 398 Range 25-2520 21-3540 9-7050 45-2950 Peripheral-blood blasts, % (Range)         CP 5(0-12) 4.5(0-14) 3(0-11) 4(0-9) AP 7(2-21) 9(0-22) 4(0-29) 12(5-19) BC 38(21-55) 36(15-60) 33(18-80) 34(15-53) Peripheral-blood basophils, % (Range)         CP 3(0-32) 5(0-36) 6(0-23) 4(0-20) AP 4(0-15) 5(0-10) 3(0-11) 5(1-9) BC 7(5-9) 4(0-12) 6(0-18) 9(3-15) Splenomegaly, no.

(%)         Any splenomegaly 21(26.9) 61(30.0) 75(34.6) 3(10.7) At least 10 cm 8(10.3) 28(13.8) 32(14.7) 1(3.6) CP = chronic phase, AP = accelerated phase, BC = blast crisis, Rabusertib in vivo HU = hydroxyurea, HSCT = hematopoietic stem cell transplant. a On monotherapy of HU; b On IFN-α(+Ara-C) without further imatinib or HSCT; c on imatinib (excluding those of < 3 mo medication due to economic issues, transplantation and adverse events). Table 2 Treatment Efficacy in CML-CP by Regimen   HU IFN(+Ara-C) Imatinib HSCT   n = 70(%) n = 184(%) n = 154(%) n = 21(%) CHR n(%) 44(62.9) 139(75.5) 142(92.2) 17(81.0) MCyR n(%) 0 37(20.1) 116(75.3) 15(71.4) CCyR n(%) 0 Orotidine 5′-phosphate decarboxylase 29(15.8) 99(64.3) 15(71.4) ND① 47(67.1) 43(23.4) 5(3.2) 0 CHR = complete hematologic response, MCyR

= major cytogenetic response, CCyR = complete cytogenetic response. aND: without examination during the treatment. Comparison of overall survival (OS) and progression-free survival (PFS) OS and PFS for the major regimens (IFN-α, imatinib and HSCT) were EPZ015938 mw compared in CP patients, and the results showed that both OS and PFS were significantly higher in the imatinib group compared to the IFN-α and HSCT groups (Figure 2). Estimated three-year and five-year OS rates were 88.2 ± 2.9% and 85.1 ± 3.2%, respectively, in patients who received imatinib; 74.7 ± 9.9% and 62.3 ± 14.1%, respectively, in the HSCT group; 83.8 ± 3.1% and 51.2 ± 3.4%, respectively, in the IFN-α group (P = 0.0075).

Working standards were made by diluting a 2 mM stock solution of

Working standards were made by diluting a 2 mM stock solution of the malondialdehyde precursor TEP with 80% ethanol supplemented with 2% of the antioxidant BHT to suppress the decomposition of lipid peroxides during the assay. Working concentrations of 0-50 μM were prepared for the lichens and 0-8 μM for the algae. Lichen thalli were homogenized on ice with 1 ml of deionized water and URMC-099 nmr centrifuged at 16,060 × g for 10 min. Supernatants were frozen at -20°C for NOx analysis, and the pellets resuspended in 500 μl ethanol-BHT. Algae were homogenized directly in

500 μl of ethanol-BHT with glass fragments (approx. 1 mm diameter) and strong vortexing for 30 min. Subsequently, 900

μM of TBA (2.57 × 10-2M), TCA (9.18 × 10-1M), and HCl (3.20 M) working solution was added to each sample and to the standards. The samples and standards were vortexed in a Vortex Labnet NSC 683864 nmr ×100 for 5 min at 3,000 rpm and then placed in a 70°C water bath for 30 min. Afterwards, the samples and standards were vortexed again, cooled on ice, and centrifuged at 10,060 × g for 10 min. The absorbance of supernatants was measured at 532 nm (A 532) in a Spectronic Genesys8 spectrophotometer. The absorbance at 600 nm (A 600) was then measured and this value was subtracted from the A 532 to eliminate the interferences of soluble sugars in the samples [35]. NO end-products determination To estimate NO generation, NO oxidation end-products (nitrate and nitrite) were measured in the soluble fraction of the samples using a GSK458 cell line Skalar autoanalyzer,

model SAN++. The automated determination of nitrate see more and nitrite is based on the cadmium reduction method: the sample is passed through a column containing granulated copper-cadmium to reduce nitrate to nitrite. The nitrite (that originally present plus that obtained from the reduction of nitrate) concentration is determined by its diazotization with sulfanilamide followed by coupling with N-(1-naphthyl)ethylenediamine dihydrochloride to form a highly colored azo dye, the absorbance of which is measured at 540 nm. This is the most commonly used method to analyze NO production and is known as the Griess reaction [23]. Statistics At least three samples for each treatment and each incubation time were prepared. Four assays were carried out on four different days for the lichens and on three different days for the algae. Data were analyzed for significance with Student’s t-test or by ANOVA. Results Bright-field micrographs showing the general anatomy of Ramalina farinacea are presented in Figure 1. The photobiont layer is located in the medulla and is surrounded by dispersed fungal hyphae, which become densely packed in the cortex of the lichen. Figure 1 Anatomy of Ramalina farinacea. Thalli of R.

We next made quantitative measurements of the cellular uptake of

We next made quantitative measurements of the HDAC inhibitor cellular uptake of different PEG-CS-NPs formulations using flow cytometry. The mean fluorescence intensities (MFIs) of the cells after 4 h of incubation with different PEG-CS-NPs formulations were shown in Figure 7. The MFI should be directly correlated with the mean

number of NPs taken up per cell. The MFI of HeLa cells treated with the FITC-(FA + PEG)-CS-NPs was significantly higher than the FITC-PEG-CS-NPs, and even the MFI of HeLa cells treated with the FITC-(MTX + PEG)-CS-NPs was also significantly higher than the FITC-(FA + PEG)-CS-NPs. These results also supported HSP inhibitor the idea of the targeting effect of both the FITC-(FA + PEG)-CS-NPs and FITC-(MTX + PEG)-CS-NPs to HeLa cells. The presence of excess of the free FA efficiently inhibited the cellular uptake of FITC-(MTX + PEG)-CS-NPs, which confirmed that the (MTX + PEG)-CS-NPs enter the cells through the FA receptor-mediated endocytosis. Figure 6 In vitro cellular uptake of the (MTX + PEG)-CS-NPs. Laser scanning confocal microscopy images of (A) HeLa cells incubated with the FITC-PEG-CS-NPs. (B)

HeLa cells incubated with the FITC-(FA + PEG)-CS-NPs. NU7026 ic50 (C) HeLa cells incubated with the FITC-(MTX + PEG)-CS-NPs. (D) HeLa cells blocked with excess of the free FA and then incubated with the FITC-(MTX + PEG)-CS-NPs. Incubation was carried out at 37°C for 6 h. The concentration of FITC was equivalent in all formulations. All images were taken using identical

instrumental conditions and presented at the same intensity scale. Figure 7 Cellular uptake of FITC-PEG-CS-NPs, FITC-(FA + PEG)-CS-NPs, and FITC-(MTX + PEG)-CS-NPs (equivalent FITC concentration) on HeLa cells by flow cytometry (mean ± SD, n  = 3). Statistical significance: *P <0.05. These quantitative results were consistent with those qualitative results, giving a further proof of high targeting efficacy of the (MTX + PEG)-CS-NPs to HeLa cells. The possible reason is that the integral binding avidity of the (MTX + PEG)-CS-NPs towards FA receptor presents a great advantage of targeting efficacy outperformed that of the (FA + PEG)-CS-NPs towards FA receptor. As mentioned above, MTX has a suboptimal affinity to FA receptor compared Tenoxicam with FA and may be less efficient to target to FA receptor than FA. Nevertheless, it was reported that multivalent binding avidity can be kinetically limited if the binding affinity of an individual receptor-ligand pair is too tight [44, 45]. Well consistent with the above theoretical analysis, our result further suggested that the targeting specificity of the nanoscaled drug delivery systems for a particular cell type can be enhanced by the weaker binding affinity of each individual receptor-ligand pair. Indeed, the integral binding avidity plays a predominant role in the targeting efficacy; the higher integral binding avidity increases the targeting efficacy.

After removal of RNA, 2 μg of cDNA was fragmented with DNase and

After removal of RNA, 2 μg of cDNA was fragmented with DNase and end-labeled (GeneChip®

WT Terminal Labeling Kit; Affymetrix). Size distribution of the fragmented and end-labeled cDNA, was assessed using an Agilent 2100 Bioanalyzer. 2 μg of end-labeled fragmented cDNA was used in a 200-μl hybridization cocktail containing added hybridization controls and hybridized on arrays for 16 hours at 48°C. Standard Caspase cleavage post hybridization wash and double-stain protocols (FS450_0001; Selleckchem CT99021 GeneChip HWS kit, Affymetrix) were used on an Affymetrix GeneChip Fluidics Station 450. Arrays were scanned on an Affymetrix GeneChip scanner 3000 7G. Microarray analysis Scanned arrays were first analyzed using Affymetrix Expression Console software to obtain Absent/Present

calls and assure that all quality parameters were in the recommended range. Subsequent analysis was carried out with DNA-Chip Analyzer 2008. First a digital mask was applied, leaving for analysis only the 8305 probe sets on the array representing Sinorhizobium meliloti transcripts. Then the 6 arrays were normalized to a baseline array with median CEL intensity by applying an Invariant Set Normalization Method [51]. Normalized CEL intensities of the arrays were used to obtain model-based gene expression indices based on a PM (Perfect Match)-only model [52]. Replicate data (triplicates) for each of the wild-type and tolC mutant strains were weighted gene-wise by using inverse squared standard error as weights.

Genes compared were considered to be differentially expressed if the 90% lower confidence bound of the fold change between experiment and baseline was PD0332991 nmr above 1.2, resulting in 3155 differentially expressed transcripts with a median False Discovery Rate (FDR) of 0.4%. The lower confidence bound criterion means that we can be 90% confident that the fold change is a value between the lower confidence bound and a variable upper confidence bound. Li and Wong [52] have shown that the lower confidence bound is a conservative estimate of the fold change and therefore more reliable as a ranking statistic for changes CYTH4 in gene expression. For a second analysis Partek Genomics Suite 6.4 was used. Here the 6 arrays were normalized and modeled using Robust Multichip Averaging (RMA). After RMA, probe sets analyzing expression of transcripts of Medicago truncatula and Medicago sativa, were filtered out. For the remaining S. meliloti probe sets differential expression was determined using 1-way Analysis of Variance (ANOVA). FDR analysis with a cut-off of 5% determined 2842 transcripts as differentially expressed, corresponding to an ANOVA p-value cut-off of <0.017. A set of 2067 differentially expressed transcripts was identified in the two independent analyses performed. All further analyses focused on this core set. Fold change values presented in Tables 1 and 2 and in the additional files 1 and 2 were obtained using Partek Genomics Suite 6.4.

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