Resource Information for Grant Applications

 

The Research Technology Support Facility (RTSF) at Michigan State University is supported and managed under the auspices of the Office of the Vice President for Research and Graduate Studies.

The RTSF Cores provide technical and analytical support for Biomedical and Agricultural research at Michigan State University in Flow Cytometry, Genomics, Mass Spectrometry, and Proteomics. The Flow Cytometry Core offers two multiple laser-based optical flow cytometers that analyze cell populations for multiple characteristics simultaneously. The Genomics Core provides single gene to genomic scale DNA sequencing services, fragment analysis genotyping, qPCR, and other nucleic acid QC services. The Proteomics Core offers high-throughput mass spectral protein identification and determination of protein expression patterns. The Mass Spectrometry Core uses an array of mass spectral techniques for small molecule identification as well as analyses and for Metabolomic profiling.

 

Flow Cytometry Core

This facility has available two flow cytometers – an Influx cell sorter and an LSR II for cell analysis only. The Influx is capable of sorting cells from bacteria to plant protoplasts in size from low to high speeds as dictated by the cells being sorted. Single to 6 way collection of sorted cells is available including collection to a variety of culture plates and tubes. The Influx is a BSL2+ capable sorter installed in a Baker hood with the Influx having additional aerosol collection safety equipment. The machine is equipped with 5 lasers. A 561 laser is used to excite PE and its conjugates (PE, PE-Texas Red, PE-Cy5 or Cy5.5, and PE-Cy7) as well as mCherry, mTomato, and others. A blue laser (488nm) is used to excite FITC, PI, PerCP-Cy5.5 with a filter set for GFP/YFP. A red laser (640 nm) is used to excite APC, Alexa 700 and APCCy7. A violet laser (405nm) is used to excite cyan proteins or Pacific Blue, and Pacific Orange. A uv laser (355nm) allows use of indo-1 calcium flux, Hoechst 342 red/blue stem cell side population and DAPI for cell cycle analysis. Data acquisition is with Sortware.

The LSR II is equipped with 3 lasers. The blue laser has 4 detectors capable of detecting FITC, PE, PE-Cy7 and one of the following (PE-TxRed, PE-Cy5, PerCpCy5.5). It has a red laser with three detectors for 660 nm, 700 nm and 780 nm fluorescence. It also has a violet laser (405) with two detectors which are useful for violet excited cyan protein or Pacific blue and Pacific orange. Data acquisition is with DIVA software. Both machines use FCS3.0 data file format for export to post acquisition analysis.

Software for data analysis is available for cell cycle analysis (Modfit), and phenotyping (WinList 6 and FlowJo). A large variety of methods for the study of plant and animal cells may be executed on these machines. Apoptotic cell death, cell proliferation, cell activation and signaling, cell integrity, cell and organelle function, and presence or absence of plasma membrane, cytoplasmic or nuclear markers for up to 9 or 15 markers can be done simultaneously, LSR II or Influx respectively. Studies of cellular DNA can be done to determine DNA content and presence or absence of aneuploidy and polyploidy.

 

Genomics Core

The MSU Genomics Core provides small scale to genome size high throughput sequencing services. The Core also provides instrumentation for SNP profiling, fragment analysis genotyping, Quantitative Real Time PCR and other nucleic acid QC services. Equipment within the Core includes an Illumina HiSeq 4000, an Illumina NextSeq 500 and two Illumina MiSeqs for next-generation sequencing. An Oxford Nanopore GridION is available for long-read sequencing, and an Oxford Nanopore PromethION is expected to be introduced to the Core in late November 2019. An ABI 3730xl 96-capillary DNA sequencer performs traditional Sanger sequencing, fragment analysis and SNP sequencing. A TakaraBio SmartChip RT-PCR System and an Applied Biosystems QuantStudio 7 Flex are used for qPCR analysis. Gene expression analysis may also be performed on a NanoString nCounter. A Biomek FX robot and a BioMek 3000 robot are available for repetitive liquid handling tasks. Size-specific shearing of nucleic acids is achieved with a Covaris M220 Sonicator. An Agilent 4200 TapeStation and an Agilent 2100 Bioanalyzer are used for RNA and DNA quality and size analysis. Fluorescence-based nucleic acid quantification is performed on either an Invitrogen Qubit or a FLUOstar OPTIMA plate reader.

 

Mass Spectrometry Core

The RTSF Mass Spectrometry and Metabolomics Core is an open access resource where MSU researchers and facility staff perform analyses of small molecules, intact macromolecules, peptides, lipids, and metabolomic analyses using 12 mass spectrometers. Analyses typically involve compound identification or quantitative analysis.  Students and other researchers are encouraged to become certified users of the instruments, and the facility staff offers frequent training in the theory and operation of facility instruments. Trained instrument users enjoy 24/7 open access to instruments; four staff experts perform analyses and provide consulting regarding method development and data interpretation. The facility houses seven LC/MS/MS instruments including Waters Xevo TQ-S, Quattro Premier XE, Acquity TQ-D, and Quattro Micro API UPLC/MS/MS systems, all of which are tandem quadrupole systems dedicated to targeted analysis of small molecules. The complement of LC/MS/MS instruments offers analyses using electrospray, atmospheric pressure chemical ionization (APCI), nanoelectrospray, and atmospheric pressure photoionization (APPI). In addition, two Waters Xevo G2-XS QTof UPLC/MS/MS systems and a Thermo Q-Exactive UPLC/MS/MS instrument provide accurate mass (high resolution) analyses of synthetic and natural products, non-targeted and targeted metabolomic, lipidomic, and glycomic profiling, and analyses of peptides and intact macromolecules including hydrogen/deuterium exchange (HDX). Four GC/MS instruments are also available including Agilent 5975C and Thermo DSQ-II quadrupole GC/MS systems, an Agilent 7010B tandem quadrupole GC/MS/MS system, and a Waters GCT Premier GC/TOF high resolution MS instrument. GC/MS and MS/MS capabilities include EI and CI, automated SPME injections, and thermal desorption analysis. MALDI analyses of materials ranging from small molecules, peptides, oligonucleotides, metabolites, and polymers can be performed on a Shimadzu AXIMA-cfr plus instrument equipped with MALDI imaging capability. Metabolomic and metabolite profiling analyses can be performed using a variety of instrument platforms, and are supported with data processing and statistical analysis software. Methods are currently running for high-throughput LC/MS/MS profiling of amino acids, phytohormones, signaling oxylipins, structural lipids, and organic acid metabolites. The Core performs analyses of approximately 40,000 samples/year from institutions from around the world.

 

Proteomics Core

The Proteomics Core has at its disposal two state of the art LC/MS/MS mass spectrometers; a Thermo Scientific Q-Exactive and a Thermo Scientific Linear Ion-Trap (LTQ).  The Q-Exactive is interfaced with a Thermo EASY nLC-1000 UPLC and Flex-Spray Ionization Source.  Together this system combines unique High Resolution/Accurate Mass measurement capabilities and high sensitivity Orbitrap detection with exceptional mass selection and spectral multiplexing over extended, stable LC gradients making it ideal for deep proteome profiling of complex samples and highly specific peptide quantitation.  The LTQ is connected to a Waters nano-Acquity UPLC and offers high sensitivity, rapid scanning speed and MSn capabilities along with exceptional chromatographic separation.  The facility offers data processing and quantitation using Mascot Distiller, Proteome Discoverer, MaxQuant and Skyline software suites.  Mascot, SEQUEST and Andromeda search algorithms are available for peptide-to-spectrum matching as well as Scaffold for data visualization and statistical validation.  Up front chromatographic fractionation of complex peptide samples is offered using a Waters Acquity H-class UPLC and an Agilent 3100 Offgel Fractionator.  Protein level separation can be performed using the Invitrogen ZOOM IEF fractionator or with 1D and 2D PAGE.