The Macromolecular Synthesis Core at Penn State College of Medicine was established in 1987, with primary grant support from the National Science Foundation as well as College of Medicine funds. The facility subsequently received grant support from the NIH and the Commonwealth of Pennsylvania via the Ben Franklin Partnership, and matching funds from the College of Medicine.
The core is heavily used, and generates approximately 2,000 oligonucleotides, 100 peptides and 40 protein sequences per year. A wide range of faculty from the College of Medicine use this core lab, as do many outside contractors. The goal has been to offer fair market costs, with the charges for services used to offset chemical and maintenance costs. Funding from NIH, NSF, Tobacco Settlement Funds and Commonwealth of Pennsylvania grants has helped to provide instrumentation for important new research services.
Services offered include:
- Protein sequencing
- Peptide synthesis
- Oligonucleotide synthesis
- Image analysis and gel band quantitation (for which instrumentation purchased with Tobacco Settlement Funds)
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Instrumentation and Services
The newest instrument for peptide synthesis is the core’s Tribute Peptide Synthesizer from Protein Technology (Summer 2010).
A Milligen 9050 Fmoc peptide synthesizer has been the workhorse for peptide synthesis, and the core has also used a second synthesizer (Perseptive 9050 Plus) through a donation from the duPont Nemours Company in Wilmington, Del.
A standard synthesis on any of these machines delivers between 50 and 80 mg of peptide. Custom syntheses are also possible (e.g., phosphopeptides, biotinylated peptides), with an additional cost being determined by the reagents used.
The Polygen synthesizer is used to generate oligonucleotides. The cost depends on the scale of the synthesis needed. Many users choose to have their oligonucleotides purified, which can be done with Nensorb columns. Custom syntheses such as modified bases and phosphorothioate chemistries are also available, as are larger-scale syntheses.
For quantitation of exposed film (radioactive or chemiluminescent detection), a BioRad GS800 Calibrated Densitometer is available to digitize film images. The densitometric image obtained can then be analyzed using Quantity One software to analyze 1D gels or slot/dot blot images, or PDQuest software to analyze two-dimensional gel images.
For quantitation of differences in protein amount between different samples, equal amounts of each sample can be labeled with different fluors, usually a combination of Cy2, Cy3 and Cy5, then mixed together before 2D gel separation. Fluorescent imaging and quantitation of the resulting 2D gels can be done with the Typhoon Imager, and spot-cut lists can be exported for automated gel spot excision using the Ettan Spot Picker.
Much peptide/protein sequencing can now be done on the College’s many mass spectrometry instruments.
Investigators who have samples for which they would still like Edman sequencing done are asked to contact the core, as arrangements can be made to send the samples to a non-Penn State facility that still does Edman sequencing.
Procedures, Protocols and Forms
Full-scale synthesis (approx. 0.2 mmol yield) or half-scale synthesis (approx. 0.1 mmol yield) is available, with fees that cover the cost of the reagents.
Peptides will be prepared as a lyophilized powder cleaved from the resin in the presence of appropriate side group preservatives, extensively washed and extracted following cleavage with various solvents to remove the majority of the impurities (preservatives and byproducts of the cleavage procedure.
For an extra charge, molecular weight verification and synthesis purity is available using mass spectrometry.
To place an order, complete the Peptide Synthesis Request Form.
The core can provide 0.05 µM scale synthesis, 0.25 µM scale synthesis or 1.0µM scale synthesis.
Two different levels of Nensorb purification are available: one for oligos synthesized at 0.05 µM or 0.2 µM scale synthesis, and another larger column used for oligos synthesized at 1.0µM scale synthesis.
Pickup procedure: After synthesis, oligos may be picked up from the top shelf of the freezer in room C1732, with the data sheets attached to the side of the freezer. An approximate yield of micrograms of oligo will be included on the data sheet, but because this is calculated based on the A260 while the oligo is still in methanol before lyophilization, it is only approximate. Investigators must requantitate the yield and concentration of each oligo after it is resuspended in water or TE buffer in order to have an accurate measure of yield/concentration.
To place an order, complete the Oligosynthesis Request Form.
Investigators should cite Penn State College of Medicine Macromolecular Synthesis Core and list the names of the instruments that were utilized either in the Materials and Methods section or the Acknowledgements section of any article.
All samples must be shipped to this address:
Macromolecular Synthesis Core
Penn State College of Medicine
500 University Dr.
Hershey, PA 17033
Work With Macromolecular Synthesis
For pricing on any services, including non-standard amino acids and non-standard nucleotides, or other specialized preparation procedures that may be necessary for a particular research purpose, call the core at 717-531-6087 or email Anne Stanley at firstname.lastname@example.org or Suja Maddukuri at email@example.com.