Nuclear Magnetic Resonance Spectroscopy is one of the most important techniques for obtaining high resolution protein structure and for studying protein interaction.
The service provided by Allele Biotech benefits from over 15 years of experience with some of the most advanced equipment Bruker Avance 500 MHz (with cryoprobe), Avance 600 MHz (with cryoprobe), or Avance 800 MHz NMR spectrometers. We provide modular service that starts from bacterial culture in special medium with isotope labeling, to protein purification and analysis, to performing NMR experimental data collection.
Available Services are as follows, depending on your research needs, you can choose any starting phase or combinations
Phase I: Subcloning into Expression Vector
Cat#
ABP-CS-NMR0001
Description:
Amplification/isolation of the gene of interest out of a customer-supplied vector and subcloning it into a T7 promoter based high-level expression vector. Verification of the subcloned gene by restriction digest and sequencing. Preparation of the recombinant vector DNA.
Required Materials:
5 µg of plasmid containing gene of interest plus complete vector sequence in electronic format.
Deliverables:
1) 20 µg of expression-ready recombinant pET plasmid containing the gene of interest 2) Final sequencing data 3) Detailed research report
Duration:
2 weeks; For constructs insert > 2kb, one week additional per 1kb
Price (USD):
$999; For constructs insert > 2kb, $299 additional per 1kb
Phase II: Generation and Identification of Expression Positive Clones
Cat#
ABP-CS-NMR0002
Description:
Transformation of recombinant constructs into a suitable E .coli strain. Small scale induction to over-express target protein. Test for recombinant protein by SDS-PAGE and western blot (customer must provide appropriate antibodies).
Required Materials:
5 ug of expression-ready recombinant pET plasmid containing the gene of interest plus complete vector sequence in electronic format.
Deliverables:
1) expression positive clone ( E. coli glycerol stock or plate) 2) SDS-PAGE or Western Blot data 3) Detailed research report
Duration:
1 week
Price (USD):
$299
Phase III: Large-scale Culture
Cat#
ABP-CS-NMR0003
Description:
One liter of bacterial culture will be grown in M9 medium with 15N NH4Cl or 13C glucose as sole N or C source, respectively. Recombinant protein expression is induced with an appropriate inducer (e.g., IPTG, IAA) and harvested for protein purification.
Required Materials:
Expression positive clone ( E. coli glycerol stock or plate) together with a verified induction procedure
Deliverables:
1) Cell pellets from 1 liter fermentation 2) SDS-PAGE or Western Blot data 3) Detailed research report
Duration:
2 days
Price (USD):
$399 + Cost of 15N NH4Cl or 13C glucose
Phase IV: Protein Extraction
Cat#
ABP-CS-NMR0004
Description:
Cell pellet harvested from 1 liter of bacterial culture
after induction will be lysed with our optimized reagents and protocol. SDS Page and Western blotting (antibody provided by customer) will be performed for quality testing.
Required Materials:
Cell pellet harvested from 1 liter of bacterial culture after induction
Deliverables:
1) Extracted protein in standard or customer-specified buffer 2) SDS-PAGE or Western Blot data 3) Detailed research report
Duration:
2 days
Price (USD):
$199
Phase V: Protein Purification (For tagged protein only)
Cat#
ABP-CS-NMR0005
Description:
Protein purification using appropriate affinity resin for 6xHis or GST-tagged recombinant proteins. Purification can be performed either under native or denatured conditions (as desired by the customer).
Required Materials:
Protein solution from bacteria expression and detailed information about the buffer.
Deliverables:
1) Purified protein in standard or customer-specified solution 2) SDS-PAGE or Western Blot data 3) Detailed research report
Duration:
1 week
Price (USD):
$999
Phase VI: NMR Analysis
Cat#
ABP-CS-NMR0006
Description:
Routine 1D, 2D, 3D NMR experimental data collection on Bruker Avance 500 MHz (with cryoprobe), Avance 600 MHz (with cryoprobe), or Avance 800 MHz NMR spectrometers for protein backbone and side chain (1H, 15N, 13C) resonance assignments or for protein structure determination.
Required Materials:
Protein with appropriate labeling and purity in ___solutions.
Deliverables:
1) Raw NMR FID data 2) Processed NMR spectra with NMRDraw software 3) Other
The quality of data is dependent on the sample, we will deliver the best possible set of data to customer with suggestions for improvement.
Duration:
N/A
Price (USD):
Spectrometer time charge rate: (hourly based) $150 per hour for 800 MHz $90 per hour for 600 MHz
$75 per hour for 500 MHz NMR Experiment Setup charge:1D: $50 2D: $100 3D: $250
