Unveiling the anticancer potentiality of single cell oils produced by marine oleaginous Paradendryphiella sp. under optimized economic growth conditions.

Bioprospecting about new marine oleaginous fungi that produce advantageous bioproducts in a green sustainable process is the key of blue bioeconomy. Herein, the marine Paradendryphiella sp. was utilized for single cell oils (SCOs) production economically, via central composite design, the lipid content enhanced 2.2-fold by 5.5 g/L lipid yeild on seawater-based media supplemented with molasses concentration 50 g/L, yeast extract, 2.25 g/L at initial pH value (5.3) and 8 days of static incubation. Subsequently, the fatty acid methyl esters profiles of SCOs produced on optimized media under different abiotic conditions were determined; signifying qualitative and quantitative variations. Interestingly, the psychrophilic-prolonged incubation increased the unsaturation level of fatty acids to 59.34%, while ω-6 and ω-3 contents representing 23.53% and 0.67% respectively. Remarkably, it exhibited the highest EC100 dose by 677.03 µg/mL on normal human lung fibroblast Wi-38 cells. Meanwhile, it showed the highest inhibiting proliferation potential on cancer cell lines of A549, MDA-MB 231 and HepG-2 cells by 372.37, 417.48 and 365.00 µg/mL, respectively. Besides, it elevated the oxidative stress, the expression of key apoptotic genes and suppressed the expression of key oncogenes (NF-κB, BCL2 and cyclin D); implying its promising efficacy in cancer treatment as adjuvant drug. This study denoted the lipogenesis capacity of Paradendryphiella sp. under acidic/alkaline and psychrophilic/mesophilic conditions. Hereby attaining efficient and economic process under seasonal variation with different Egyptian marine sources to fill the gap of freshwater crisis and simultaneously preserve energy.

Risk assessment of secondary metabolites produced by fungi in the genus Stemphylium.

The fungal genus Stemphylium (phylum Ascomycota, teleomorph Pleospora) includes plant pathogenic, endophytic, and saprophytic species with worldwide distributions. Stemphylium spp. produce prodigious numbers of airborne spores, so are a human health concern as allergens. Some species also produce secondary metabolites, such as glucosides, ferric chelates, aromatic polyketides, and others, that function as toxins that damage plants and other fungal species. Some of these compounds also exhibit a low level of mammalian toxicity. The high production of airborne spores by this genus can result in a high incidence of human exposure. Concern about toxin production appears to be the reason that Stemphylium vesicarium, which is a pathogen of several vegetable crops, was classified in Canada as a potential risk of harm to humans for many years. A detailed assessment of the risk of exposure was provided to the relevant regulatory body, the Public Health Agency of Canada, which then determined that Stemphylium spp. in nature or under laboratory conditions posed little to no risk to humans or animals, and the species was re-assigned as a basic (level 1) risk agent.

Resistance risk assessment for fludioxonil in Sclerotinia homoeocarpa in China.

Sclerotinia homoeocarpa causes dollar spot disease on turfgrass and is a serious problem on many species worldwide. Fludioxonil, a phenylpyrrole fungicide, is not currently registered for dollar spot control in China. In this study, the baseline sensitivity to fludioxonil was established using an in vitro assay for 105 isolates of S. homoeocarpa collected from 10 locations in different regions of China. Results indicate that the frequency distribution of effective concentration for 50% inhibition of mycelial growth (EC50) values of the S. homoeocarpa isolates was unimodal (W = 0.9847, P = .2730). The mean EC50 value was 0.0020 ±â€¯0.0006 µg/ml with a range from 0.0003 to 0.0035 µg/ml. A total of 7 fludioxonil-resistant mutants were obtained in laboratory, the mutants were stable in fludioxonil sensitivity after the 10th transfer, with resistance factor (RF) ranging from 4.320 to >13,901.4. The mutants showed a positive cross-resistance between fludioxonil and the dicarboximide fungicide iprodione, but not propiconazole, fluazinam, and thiophanate-methyl. When mycelial growth rate, pathogenicity and osmotic sensitivity were assessed, the mutants decreased in the fitness compared with their parental isolates. Sequence alignment of the histidine kinase gene Shos1 revealed a 13-bp fragment deletion only in one mutant, no mutations were observed on Shos1 in the rest resistant mutants.

Biotechnological application of endophytic filamentous bipolaris and curvularia: a review on bioeconomy impact.

The filamentous Bipolaris and Curvularia genera consist of species known to cause severe diseases in plants and animals amounting to an estimated annual loss of USD $10 billion worldwide. Despite the harmful effect of Bipolaris and Curvularia species, scarce attention is paid on beneficial areas where the fungi are used in industrial processes to generate biotechnological products. Catalytic potential of Bipolaris and Curvularia species in the production of biodiesel, bioflucculant, biosorbent, and mycoherbicide are promising for the bioeconomy. It is herein demonstrated that knowledge-based application of some endophytic Bipolaris and Curvularia species are indispensable vectors of sustainable economic development. In the twenty-first century, India, China, and the USA have taken progress in the biotechnological application of these fungi to generate wealth. As such, some Bipolaris and Curvularia species significantly impact on global crop improvement, act as catalyst in batch-reactors for biosynthesis of industrial enzymes and medicines, bioengineer of green-nanoparticle, agent of biofertilizer, bioremediation and bio-hydrometallurgy. For the first time, this study discusses the current advances in biotechnological application of Bipolaris and Curvularia species and provide new insights into the prospects of optimizing their bioengineering potential for developing bioeconomy.

Pullulan gum production from low-quality fig syrup using Aureobasidium pullulans.

Pullulan is an important polysaccharide with several potential applications in food science, pharmaceutical and cosmetic industries, but high costs of pullulan production are the main limitation for commercial utilization. Therefore, a cost-effective process for pullulan production was developed using fig syrup as an exclusive nutrient source. In particular, the feasibility of using low quality fig syrup as a supplemental substrate for pullulan gum production by Aureobasidium pullulans was investigated. Fermentation was carried out over a range of fig syrup and sucrose degrees Brix (5-15%). Maximum pullulan gum production was observed after 96h using 12.5% fig syrup, yielding approximately14.06 g/L. This value of pullulan production (14.06 g/L) was higher than the amount of pullulan produced using sucrose as substrate (5.01 g/L). In conclusion, fig syrup was an effective substrate for pullulan production by Aureobasidium pullulans, and, therefore, this byproduct deserves attention for the cost-effective and environmentally friendly pullulan production.

Economic co-production of poly(malic acid) and pullulan from Jerusalem artichoke tuber by Aureobasidium pullulans HA-4D.

BACKGROUND: poly(L-malic acid) (PMA) is a water-soluble polyester with many attractive properties in medicine and food industries, but the high cost of PMA fermentation has restricted its further application for large-scale production. To overcome this problem, PMA production from Jerusalem artichoke tubers was successfully performed. Additionally, a valuable exopolysaccharide, pullulan, was co-produced with PMA by Aureobasidum pullulans HA-4D. RESULTS: The Jerusalem artichoke medium for PMA and pullulan co-production contained only 100 g/L hydrolysate sugar, 30 g/L CaCO3 and 1 g/L NaNO3. Compared with the glucose medium, the Jerusalem artichoke medium resulted in a higher PMA concentration (114.4 g/L) and a lower pullulan concentration (14.3 g/L) in a 5 L bioreactor. Meanwhile, the activity of pyruvate carboxylase and malate dehydrogenas was significantly increased, while the activity of α-phosphoglucose mutase, UDP-glucose pyrophosphorylase and glucosyltransferase was not affected. To assay the economic-feasibility, large-scale production in a 1 t fermentor was performed, yielding 117.5 g/L PMA and 15.2 g/L pullulan. CONCLUSIONS: In this study, an economical co-production system for PMA and pullulan from Jerusalem artichoke was developed. The medium for PMA and pullulan co-production was significantly simplified when Jerusalem artichoke tubers were used. With the simplified medium, PMA production was obviously stimulated, which would be associated with the improved activity of pyruvate carboxylase and malate dehydrogenas.

Antimicrobial activity of pomegranate peel extracts as affected by cultivar.

BACKGROUND: Some studies have reported that different parts of the pomegranate fruit, especially the peel, may act as potential antimicrobial agents and thus might be proposed as a safe natural alternative to synthetic antimicrobial agents. The high tannin content, especially punicalagin, found in pomegranate extracts, has been reported as the main compound responsible for such antimicrobial activity. Because the pomegranate peel chemical composition may vary with the type of cultivar (sweet, sour-sweet and sour), pomegranates may also differ with respect to their antimicrobial capacity. RESULTS: The extract from PTO8 pomegranate cultivar peel had the highest antimicrobial activity, as well as the highest punicalagins (α and ß) and ellagic acid concentrations. In the results obtained from both antibacterial and antifungal activity studies, the sour-sweet pomegranate cultivar PTO8 showed the best antimicrobial activity, and the highest ellagic acid concentrations. CONCLUSION: The results of the present study suggest that ellagic acid content has a significant influence on the antimicrobial activity of the pomegranate extracts investigated. The pomegranate peel of the PTO8 cultivar is a good source of antifungal and antibacterial compounds, and may represent an alternative to antimicrobial agents of synthetic origin. © 2016 Society of Chemical Industry.

Production of poly(malic acid) from sugarcane juice in fermentation by Aureobasidium pullulans: Kinetics and process economics.

Poly(ß-l-malic acid) (PMA) is a biodegradable polymer with many potential biomedical applications. PMA can be readily hydrolyzed to malic acid (MA), which is widely used as an acidulant in foods and pharmaceuticals. PMA production from sucrose and sugarcane juice by Aureobasidium pullulans ZX-10 was studied in shake-flasks and bioreactors, confirming that sugarcane juice can be used as an economical substrate without any pretreatment or nutrients supplementation. A high PMA titer of 116.3g/L and yield of 0.41g/g were achieved in fed-batch fermentation. A high productivity of 0.66g/L·h was achieved in repeated-batch fermentation with cell recycle. These results compared favorably with those obtained from glucose and other biomass feedstocks. A process economic analysis showed that PMA could be produced from sugarcane juice at a cost of $1.33/kg, offering a cost-competitive bio-based PMA for industrial applications.

Polymalic acid fermentation by Aureobasidium pullulans for malic acid production from soybean hull and soy molasses: Fermentation kinetics and economic analysis.

Polymalic acid (PMA) production by Aureobasidium pullulans ZX-10 from soybean hull hydrolysate supplemented with corn steep liquor (CSL) gave a malic acid yield of ∼0.4g/g at a productivity of ∼0.5g/L·h. ZX-10 can also ferment soy molasses, converting all carbohydrates including the raffinose family oligosaccharides to PMA, giving a high titer (71.9g/L) and yield (0.69g/g) at a productivity of 0.29g/L·h in fed-batch fermentation under nitrogen limitation. A higher productivity of 0.64g/L·h was obtained in repeated batch fermentation with cell recycle and CSL supplementation. Cost analysis for a 5000 MT plant shows that malic acid can be produced at $1.10/kg from soy molasses, $1.37/kg from corn, and $1.74/kg from soybean hull. At the market price of $1.75/kg, malic acid production from soy molasses via PMA fermentation offers an economically competitive process for industrial production of bio-based malic acid.

Comparative assessment of fungal augmentation treatments of a fine-textured and historically oil-contaminated soil.

The removal of aged hydrophobic contaminants from fine-textured soils is a challenging issue in remediation. The objective of this study was to compare the efficacy of augmentation treatments to that of biostimulation in terms of total aliphatic hydrocarbon (TAH) and toxicity removal from a historically contaminated clay soil and to assess their impact on the resident microbial community. To this aim, Pleurotus ostreatus, Botryosphaeria rhodina and a combination of both were used as the inoculants while the addition of a sterilized lignocellulose mixture to soil (1:5, w/w) was used as a biostimulation approach. As opposed to the non-amended control soil, where no changes in TAH concentration and residual toxicity were observed after 60days, the activation of specialized bacteria was found in the biostimulated microcosms resulting in significant TAH removal (79.8%). The bacterial community structure in B. rhodina-augmented microcosms did not differ from the biostimulated microcosms due to the inability of the fungus to be retained within the resident microbiota. Best TAH removals were observed in microcosms inoculated with P. ostreatus alone (Po) and in binary consortium with B. rhodina (BC) (86.8 and 88.2%, respectively). In these microcosms, contaminant degradation exceeded their bioavailability thresholds determined by sequential supercritical CO2 extraction. Illumina metabarcoding of 16S rRNA gene showed that the augmentation with Po and BC led to lower relative abundances of Gram(+) taxa, Actinobacteria in particular, than those in biostimulated microcosms. Best detoxification, with respect to the non-amended incubation control, was found in Po microcosms where a drop in collembola mortality (from 90 to 22%) occurred. At the end of incubation, in both Po and BC, the relative abundances of P. ostreatus sequences were higher than 60% thus showing the suitability of this fungus in bioaugmentation-based remediation applications.

Optimization of genome shuffling for high-yield production of the antitumor deacetylmycoepoxydiene in an endophytic fungus of mangrove plants.

As an accelerated evolutionary tool, genome shuffling is largely dependent on the high fusion frequency of different parental protoplasts. However, it was unclear how many types of parental protoplasts would afford the highest fusion frequency. Here, we applied the Monte Carlo method to simulate the simplified processes of protoplast fusion, to achieve maximal useful fusions in genome shuffling. The basic principle of this simulation is that valid fusions would take place when the minimum distance between two different types of parent protoplasts is smaller than that between two of the same types. Accordingly, simulations indicated that the highest fusion frequency would be achieved from eight to 12 different parental protoplasts. Based on the simulation results, eight parental protoplasts of the fungal endophyte Phomopsis sp. A123 were subjected to genome shuffling for yield improvement of deacetylmycoepoxydiene (DAM), an antitumor natural product with a novel chemical structure. After only two rounds of genome shuffling, four high-yield DAM-producing strains, namely G2-119, G2-448, G2-866, and G2-919, were obtained with the aid of activity screening and HPLC analysis. The results showed that the DAM yield in these four strains were 243-, 241-, 225-, and 275-fold, respectively, higher than that of the starting strain A123. This is the first time Monte Carlo simulation is introduced into the field of cell fusion and is also the first report on the optimization of genome shuffling focusing on the number of parental types in protoplast fusions.

In silico analysis of the core signaling proteome from the barley powdery mildew pathogen (Blumeria graminis f.sp. hordei).

BACKGROUND: Compared to other ascomycetes, the barley powdery mildew pathogen Blumeria graminis f.sp. hordei (Bgh) has a large genome (ca. 120 Mbp) that harbors a relatively small number of protein-coding genes (ca. 6500). This genomic assemblage is thought to be the result of numerous gene losses, which likely represent an evolutionary adaptation to a parasitic lifestyle in close association with its host plant, barley (Hordeum vulgare). Approximately 8% of the Bgh genes are predicted to encode virulence effectors that are secreted into host tissue and/or cells to promote pathogenesis; the remaining proteome is largely uncharacterized at present. RESULTS: We provide a comparative analysis of the conceptual Bgh proteome, with an emphasis on proteins with known roles in fungal development and pathogenicity, for example heterotrimeric G proteins and G protein coupled receptors; components of calcium and cAMP signaling; small monomeric GTPases; mitogen-activated protein cascades and transcription factors. The predicted Bgh proteome lacks a number of proteins that are otherwise conserved in filamentous fungi, including two proteins that are required for the formation of anastomoses (somatic hyphal connections). By contrast, apart from minor modifications, all major canonical signaling pathways are retained in Bgh. A family of kinases that preferentially occur in pathogenic species of the fungal clade Leotiomyceta is unusually expanded in Bgh and its close relative, Blumeria graminis f.sp. tritici. CONCLUSIONS: Our analysis reveals characteristic features of the proteome of a fungal phytopathogen that occupies an extreme habitat: the living plant cell.

Efficient production of pullulan using rice hull hydrolysate by adaptive laboratory evolution of Aureobasidium pullulans.

Pullulan production by Aureobasidium pullulans CCTCC M 2012259 using rice hull hydrolysate as the carbon source was conducted. The acetic acid in the hydrolysate was demonstrated to exert a negative effect on pullulan biosynthesis. Instead of employing expensive methods to remove acetic acid from the hydrolysate, a mutant A. pullulans ARH-1 was isolated following 20 cycles of adaptive laboratory evolution of the parental strain on medium containing acetic acid. The maximum pullulan production achieved by the adapted mutant at 48 h using the hydrolysate of untreated rice hull was 22.2 g L(-1), while that obtained by the parental strain at 60 h was 15.6 g L(-1). The assay of key enzymes associated with pullulan biosynthesis revealed that acetic acid inhibited enzyme activity rather than suppressing enzyme synthesis. These results demonstrated that adaptive evolution highly improved the efficiency of pullulan production by A. pullulans using the hydrolysate of untreated rice hull.

Critical assessment of proteome-wide label-free absolute abundance estimation strategies.

There is a great interest in reliable ways to obtain absolute protein abundances at a proteome-wide scale. To this end, label-free LC-MS/MS quantification methods have been proposed where all identified proteins are assigned an estimated abundance. Several variants of this quantification approach have been presented, based on either the number of spectral counts per protein or MS1 peak intensities. Equipped with several datasets representing real biological environments, containing a high number of accurately quantified reference proteins, we evaluate five popular low-cost and easily implemented quantification methods (Absolute Protein Expression, Exponentially Modified Protein Abundance Index, Intensity-Based Absolute Quantification Index, Top3, and MeanInt). Our results demonstrate considerably improved abundance estimates upon implementing accurately quantified reference proteins; that is, using spiked in stable isotope labeled standard peptides or a standard protein mix, to generate a properly calibrated quantification model. We show that only the Top3 method is directly proportional to protein abundance over the full quantification range and is the preferred method in the absence of reference protein measurements. Additionally, we demonstrate that spectral count based quantification methods are associated with higher errors than MS1 peak intensity based methods. Furthermore, we investigate the impact of miscleaved, modified, and shared peptides as well as protein size and the number of employed reference proteins on quantification accuracy.

Assessment of the cytochrome B substitution G143A in the Algerian population of Mycosphaerella graminicola.

Mycosphoerella graminicola (anamorph: Zymoseptoria tritici), causal agent of Septoria tritici blotch, is currently one of the most damaging diseases on both bread and durum wheat crops worldwide. Since wheat resistance against this pathogen is always partial at various extents in most cultivars, disease control relies mainly on the use of fungicides. However, management of fungicide applications is necessary in order to avoid the emergence and widespread of fungicide resistant genotypes within populations of the pathogen. In the present study, we investigated for the first time the resistance of M. graminicola toward strobilurin fungicides in Algeria. This was performed by identifying the G143A substitution of the cytochrome b encoding sequence (which confers resistance to strobilurins) in a collection of 120 single-conidial isolates. These isolates have been sampled during the 2012 growing season from five distinct geographical locations (Guelma, Annaba, Constantine, Skikda and Oran). We used a PCR-based mismatch mutation assay allowing the amplification of either G143 (sensitive) or A143 (resistant) allele in each isolate. This study should give valuable information regarding the management of strobilurin use in order to control in a durable manner M. graminicola epidemics in Algeria.

13C pulse-labeling assessment of the community structure of active fungi in the rhizosphere of a genetically starch-modified potato (Solanum tuberosum) cultivar and its parental isoline.

• The aim of this study was to gain understanding of the carbon flow from the roots of a genetically modified (GM) amylopectin-accumulating potato (Solanum tuberosum) cultivar and its parental isoline to the soil fungal community using stable isotope probing (SIP). • The microbes receiving (13)C from the plant were assessed through RNA/phospholipid fatty acid analysis with stable isotope probing (PLFA-SIP) at three time-points (1, 5 and 12 d after the start of labeling). The communities of Ascomycota, Basidiomycota and Glomeromycota were analysed separately with RT-qPCR and terminal restriction fragment length polymorphism (T-RFLP). • Ascomycetes and glomeromycetes received carbon from the plant as early as 1 and 5 d after labeling, while basidiomycetes were slower in accumulating the labeled carbon. The rate of carbon allocation in the GM variety differed from that in its parental variety, thereby affecting soil fungal communities. • We conclude that both saprotrophic and mycorrhizal fungi rapidly metabolize organic substrates flowing from the root into the rhizosphere, that there are large differences in utilization of root-derived compounds at a lower phylogenetic level within investigated fungal phyla, and that active communities in the rhizosphere differ between the GM plant and its parental cultivar through effects of differential carbon flow from the plant.

Assessment of the stereoselective fungal biotransformation of albendazole and its analysis by HPLC in polar organic mode.

A high-performance liquid chromatographic method using polar organic mode was developed to analyze albendazole (ABZ), albendazole sulfone (ABZSO(2)) and the chiral and active metabolite albendazole sulfoxide (ABZSOX, ricobendazole) that was further applied in stereoselective fungal biotransformation studies. The chromatographic separation was performed on a Chiralpak AS column using acetonitrile:ethanol (97:3, v/v) plus 0.2% triethylamine and 0.2% acetic acid as the mobile phase at a flow rate of 0.5 mL min(-1). The present study employed hollow fiber liquid-phase microextraction as sample preparation. The method showed to be linear over the concentration range of 25-5000 ng mL(-1) for each ABZSOX enantiomer, 200-10,000 ng mL(-1) for ABZ and 50-1000 ng mL(-1) for ABZSO(2) metabolite with correlation coefficient (r)>0.9934. The mean recoveries for ABZ, rac-ABZSOX and ABZSO(2) were, respectively, 9%, 33% and 20% with relative standard deviation below 10%. Within-day and between-day precision and accuracy assays for these analytes were studied at three concentration levels and were lower than 15%. This study opens the door regarding the possibility of using fungi in obtaining of the active metabolite ricobendazole. Nigrospora sphaerica (Sacc.) E. W. Mason (SS67), Pestalotiopsis foedans (VR8), Papulaspora immersa Hotson (SS13) and Mucor rouxii were able to stereoselectively metabolize ABZ into its chiral metabolite. Among them, the fungus Mucor rouxii was the most efficient in the production of (+)-ABZSOX.

A new ß-glucosidase producing yeast for lower-cost cellulosic ethanol production from xylose-extracted corncob residues by simultaneous saccharification and fermentation.

This study reports a new yeast strain of Clavispora NRRL Y-50464 that is able to utilize cellobiose as sole source of carbon and produce sufficient native ß-glucosidase enzyme activity for cellulosic ethanol production using SSF. In addition, this yeast is tolerant to the major inhibitors derived from lignocellulosic biomass pre-treatment such as 2-furaldehyde (furfural) and 5-(hydroxymethyl)-2-furaldehyde (HMF), and converted furfural into furan methanol in less than 12h and HMF into furan-2,5-dimethanol within 24h in the presence of 15 mM each of furfural and HMF. Using xylose-extracted corncob residue as cellulosic feedstock, an ethanol production of 23 g/l was obtained using 25% solids loading at 37 °C by SSF without addition of exogenous ß-glucosidase. Development of this yeast aids renewable biofuels development efforts for economic consolidated SSF bio-processing.

Production of multi-fiber modifying enzyme from Mamillisphaeria sp. for refining of recycled paper pulp.

Enzymatic modification of pulp is receiving increasing interest for energy reduction at the refining step of the paper-making process. In this study, the production of a multi-fiber modifying enzyme from Mamillisphaeria sp. BCC8893 was optimized in submerged fermentation using a response-surface methodology. Maximal production was obtained in a complex medium comprising wheat bran, soybean, and rice bran supplemented with yeast extract at pH 6.0 and a harvest time of 7 d, resulting in 9.2 IU/mL of carboxymethyl cellulase (CMCase), 14.9 IU/mL of filter paper activity (FPase), and 242.7 IU/mL of xylanase. Treatment of old corrugated container pulp at 0.2-0.3 IU of CMCase/g of pulp led to reductions in refining energy of 8.5-14.8%. The major physical properties were retained, including tensile and compression strength. Proteomic analysis showed that the enzyme was a complex composite of endo-glucanases, cellobiohydrolases, beta-1,4-xylanases, and beta-glucanases belonging to various glycosyl hydrolase families, suggestive of cooperative enzyme action in fiber modification, providing the basis for refining efficiency.

Getting the most out of your fungal microarray data: two cost- and time-effective methods.

Advances in genome sequencing technologies have facilitated production of a wealth of fungal data; within the last 5 years, experimental costs and labor have diminished, shifting the production bottleneck from genomic data generation to data analysis. Genome sequences and microarrays now exist for many fungi, and transcriptional profiling has been shown to be an efficient way to examine how the entire genome changes in response to many different environments or treatments. Multiple platforms, programs, and protocols exist for analyzing such data, making this task daunting for the bench-based scientist. Furthermore, many existing programs are expensive and require license renewals on a yearly basis for each user in the laboratory. Costs may be prohibitively high for bench-based scientists in academia. Our combined experiences with this kind of analysis have favored two programs, depending upon whether the scientist is working with single- or dual-channel hybridization data. Our protocols are aimed toward helping the bench-based PI get the most possible information from their data, without the need for expensive software or an experienced bioinformaticist.