E. coli Vector Panel Expression Test

Panel Test Overview

Discover the vector that gives the best protein expression levels for your gene. Choose from three pre-selected panels: promoters, solubility tags or secretion signals; or create your own custom panel. ATUM will clone and panel test your gene for expression levels to identify the best vector.


    Availability

    • Test your construct in any bacterial vector panel – Promoters, Solubility Tags, or Secretion Signals
    • Cost – Expression Panel Testing ($395/construct, 10% discount for testing an entire panel), in addition to regular gene synthesis and vector panel costs
    • Contact info@atum.bio for ordering information

    Case Study: Promoter Panel Results

    Different promoters work better for different proteins. Expression of three proteins, phi29 (~61 kD), cutinase (~22 kD) and DasherGFP (~26 kD), under control of different promoters, T5 and T7 (IPTG-inducible), rham (rhamnose-inducible), phoA (inducible by phosphate starvation), and ara (arabinose-inducible, only DasherGFP expression tested) is shown. Expression values shown on the y-axis are measurements of expressed protein band densities from a SDS-PAGE gel (shown below) using BSA as standard.
    *DasherGFP measured fluorescence is lower with T7 and rham expressed GFP, indicating that a fraction of total protein is incompletely folded protein.

    Bacterial promoters panel test gel


    Case Study: Solubility Tag Panel Results

    Expression of total and soluble protein obtained using different tags. Three proteins encoded by genes A (~48 kD), B (~49 kD) and C (~59 kD), having shown poor solubility, were expressed in IPTG-inducible T5 promoter E. coli expression vectors (pD441-XXX) with various N-terminal solubility tags: MBP, GST, PpiB, and Fh8; 6XHis was included as a negative control. Cultures were grown at room temperature (RT) and 37°C. Overall, higher soluble protein expression was observed at RT (data shown above and seen in gel below). Percent solubility is shown along the Y-axis and the size of the circles corresponds to protein yield shown in nmole/µl. Legend shows approximate yield of protein based on circle size.

    Solubility Tags Panel Test Gel


    Case Study: Secretion Signal Panel Results

    Different proteins prefer different secretion signals. Periplasmic expression of MBP, cutinase, and alkaline phosphatase from rhamnose vectors (pD861, high copy) with different secretion signals. Total cellular protein levels are represented as circle diameter. Soluble protein in the periplasm is shown on the y-axis. Protein was quantified by densitometry of stained acrylamide gels.


    Secretion Signals Panel Test gel

    Periplasmic expression of Cutinase from rhamnose vectors (pD881, low copy) with different secretion signals. Total protein samples and periplasmic fractions were prepared and loaded on a 4-12% NuPAGE Bis-Tris MES gel and Coomasie stained. As observed from the gels, there are clear differences in expression and processing of the cutinase protein with different secretion signals seen in the periplasmic fractions (shown by arrows).

    Deliverables

    • ATUM Expression Data Report
    • SDS-PAGE results for total and soluble protein fractions
    • Semi-quantitative Western* results (optional for genes with His or Flag tag)
    • Growth and induction protocols

    Terms & Conditions

    • Expression panel tests are available for bacterial vector panels with a volume discount. Cost for expression testing is $395.00 USD per construct per vector with a 10% discount off the total testing cost. Discount does not apply to regular gene synthesis or cloning fees.
    • Prices are per construct (gene) and are in addition to regular gene synthesis and cloning fees.
    • * There is an additional $50 USD fee per gene for Western Blot service.
    • Listed prices are for genes <3kb in length. Please consult with our Expression Specialists to determine specific pricing for genes longer than 3kb.
    • Expression Test services must be placed at time of order for each gene.
    • Expression Standard is >5µg/ml E. coli culture under standard conditions (5.0 OD600 culture in our expression conditions) in the ATUM laboratory (detectable via SDS-PAGE and/or Western).
    • ATUM will provide electronic gel image and protocols for expression to the customer.
    • ATUM does not guarantee protein solubility.
    • ATUM does not guarantee the ability of customers, researchers or external labs to produce similar expression levels.
    • ATUM reserves the right to decline Expression Test orders.

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