Recombinant Saccharomyces cerevisiae Uncharacterized protein SCY_1535 (SCY_1535)
Description
Recombinant Production Methods
SCY_1535 is produced recombinantly using S. cerevisiae or bacterial expression systems. Key methodologies include:
Expression Systems
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Yeast Systems: Leverages S. cerevisiae’s native post-translational modification capabilities. Strain AS2.489 is commonly used with vectors like pScIKP for multi-gene co-expression .
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E. coli: Utilized for cost-effective production. Example: Full-length SCY_1535 fused to an N-terminal His-tag, expressed in E. coli and purified via affinity chromatography .
Cloning and Transformation
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Genes are amplified using PCR with primers containing restriction sites (e.g., Apa I) .
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Electrotransformation or chemical methods introduce linearized vectors (e.g., pScIKP-amy-ga-ap) into yeast .
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Selection markers like G418 (200 µg/ml) ensure recombinant strain isolation .
Potential Applications and Research Context
While SCY_1535’s biological role remains unclear, its recombinant production aligns with broader applications of S. cerevisiae proteins:
Biotechnological Uses
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Model for Protein Expression: SCY_1535 serves as a test case for optimizing full-length protein production in yeast, addressing challenges like codon bias and proteolytic degradation .
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Synthetic Biology: Used in strain engineering to study minimal functional protein units or synthetic pathways .
Comparative Studies
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SCY_1535 shares taxonomic context with characterized proteins (e.g., ARI1 in inhibitor-tolerant yeast strains), aiding studies on stress response mechanisms .
Challenges in Characterization
Research on SCY_1535 faces hurdles typical of uncharacterized proteins:
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Functional Annotation: Absence of homologous domains limits functional predictions .
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Stability Issues: Small proteins like SCY_1535 are prone to aggregation, necessitating optimized buffers (e.g., Tris/PBS with 6% trehalose) .
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Low Yield: Native expression levels are undetectable, requiring strong promoters (e.g., GAPDH) for recombinant overexpression .
Future Directions
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Functional Genomics: CRISPR/Cas9-based knockout studies could elucidate SCY_1535’s role in yeast physiology.
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Structural Biology: Cryo-EM or NMR may resolve its tertiary structure, guiding hypothesis-driven research .
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Industrial Relevance: Integration into engineered yeast strains for biofuel or pharmaceutical production, leveraging S. cerevisiae’s industrial robustness .
Product Specs
Note: We will prioritize shipping the format currently in stock. However, if you have specific format requirements, please indicate them in your order. We will accommodate your needs to the best of our ability.
Note: All of our proteins are shipped with standard blue ice packs. If dry ice shipping is required, please inform us in advance, as additional fees may apply.
Generally, liquid forms have a shelf life of 6 months at -20°C/-80°C. Lyophilized forms have a shelf life of 12 months at -20°C/-80°C.
Target Background
Q&A
Basic Research Questions
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What is SCY_1535 and what are its basic characteristics?
SCY_1535 is an uncharacterized protein from Saccharomyces cerevisiae with a full length of 72 amino acids. Currently available as a recombinant protein with His-tag, it is typically expressed in E. coli expression systems . As an uncharacterized protein, SCY_1535 presents unique research opportunities for functional genomics studies. S. cerevisiae's genome has been completely sequenced with approximately 6000 genes identified, of which 5570 are predicted to be protein-encoding . Uncharacterized proteins like SCY_1535 represent knowledge gaps in our understanding of yeast biology that require further investigation.
Property Description Source Organism Saccharomyces cerevisiae Length 72 amino acids Expression System E. coli Tag His Current Function Status Uncharacterized -
What initial experimental approaches are recommended for characterizing SCY_1535?
Initial characterization should follow a systematic approach:
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Bioinformatic analysis: Begin with sequence homology comparisons, domain prediction, and structural modeling to generate functional hypotheses.
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Gene expression profiling: Determine conditions under which SCY_1535 is expressed using RNA-seq methods. For reliable results, at least six biological replicates should be used, increasing to 12+ replicates when identifying all differentially expressed genes regardless of fold change .
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Knockout/knockdown studies: Create SCY_1535 deletion strains to observe phenotypic changes. S. cerevisiae's amenability to genetic manipulation makes it ideal for this approach .
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Protein localization: Use GFP-tagging to determine subcellular localization, which can provide functional insights.
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Interactome analysis: Perform yeast two-hybrid or co-immunoprecipitation studies to identify protein interaction partners .
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How many biological replicates are necessary for RNA-seq experiments when studying SCY_1535 expression?
Based on comprehensive RNA-seq benchmarking studies in S. cerevisiae, the number of biological replicates significantly impacts statistical power:
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With three biological replicates, most analytical tools identify only 20%-40% of significantly differentially expressed genes compared to larger replicate numbers .
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For high fold-change genes (>4-fold), 3-6 replicates may achieve >85% identification rate .
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To achieve >85% identification rate for all significantly differentially expressed genes regardless of fold change, more than 20 biological replicates are recommended .
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For studies with fewer than 12 replicates, EdgeR and DESeq2 show superior combined performance of true positives and false positives .
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For higher replicate numbers, DESeq marginally outperforms other tools when minimizing false positives is important .
Therefore, at minimum six biological replicates are recommended for studying SCY_1535 expression, increasing to 12+ for comprehensive profiling.
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What culture conditions should be considered when studying SCY_1535 in S. cerevisiae?
When designing experiments involving SCY_1535, consider multiple culture conditions as protein function may be context-dependent:
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Standard vs. stress conditions: Test multiple conditions including nutrient limitation, temperature variation, oxidative stress, and osmotic stress, as uncharacterized proteins may have condition-specific functions.
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Growth phase dependence: Examine expression during different growth phases (lag, exponential, stationary) as S. cerevisiae metabolism changes significantly between phases .
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Carbon source variations: Vary carbon sources (glucose, galactose, glycerol) to explore function under respiratory vs. fermentative conditions. S. cerevisiae exhibits the Crabtree effect, producing ethanol even under aerobic conditions when glucose is present .
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Nitrogen source variations: Test different nitrogen sources to identify potential regulatory roles.
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pH variations: Test function across pH ranges as this may affect protein activity or expression.
Documenting all culture parameters thoroughly is critical for experimental reproducibility.
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