Proteomics and Genomics

The Proteomics and Genomics Facility CIB Margarita Salas is a support service of scientific and technological research for both CIB investigators, and researchers from other centers and institutions. The service  is a part of the laboratory network of the Community of Madrid (Lab 317). Since 2012 the laboratory is certified by AENOR according to UNE-EN ISO9001:2015 in “The molecular weight determination by mass spectrometry.”

IQNetES-0286-2009_2023-12-26. 

SERVICES OFFERED:

PROTEOMICS:

In the Service it can be performed separation, quantification, identification and characterization of proteins from prokaryotic and eukaryotic organisms by mono- and two-dimensional electrophoresis, MALDI-TOF/TOF and multidimensional liquid chromatography Q-Exactive mass spectrometries.

• Proteins separation by mono (SDS-PAGE) or two-dimensional electrophoresis in 7 cm gels and subsequent colloidal Coomassie Blue staining.
• Molecular mass determination of peptides and proteins (MALDI-TOF).
• Protein identification by their peptide fingerprint and peptide fragmentation (MALDI-TOF/TOF).

• Identification of proteins in complex mixtures (“shotgun proteomics”).

• Identification and relative quantification of protein expression level (with label isobaric or label free).

• Selective monitoring of parental ions and their fragments (SIM and PRM, respectively) in targeted proteomics experiments.

• Phosphopeptides identification from protein complex mixtures

GENOMICS:

The Facility has the necessary tools for basic studies of functional genomics and expression.

• Biomolecules quantification (NanoDrop spectrophotometer).
• Chip electrophoresis of:
  • RNA, for integrity evaluation prior to microarray or rt-PCR experiments
  • DNA
• Quantitative and qualitative analysis of nucleic acids.

• Real-time PCR.

• The Facility has the Roche Universal Probe Library collection of human genome probes.

SCIENTIFIC EQUIPMENT:

PROTEOMICS

• 2 Protean IEF cell (BIO-RAD), for first dimension of 2D gels in strips of 7 cm and different nonlinear pH 3–10 , 3-10 linear and 4-7 linear gradients.
• 1 Multigel Miniprotean Dodecacell (BIO-RAD), for up to 12 simultaneous electrophoresis gels of 8 x 7 cm. This equipment allows the optimization of the electrophoretic separation conditions with a smaller amount of sample and a low cost.
• MALDI-TOF-TOF Autoflex III (Bruker) Mass Spectrometer, for molecular mass determination and protein identification through peptide fingerprinting and fragmentation of peptides.

• Mass Spectrometer Q-Exactive (Thermo Scientific) equipped with an Easy Spray nano ionization source connected on-line to an Easy-LC 1000 nano-UPLC.

• High-Resolution mass spectrometer Orbitrap Exploris 240 connected on-line to a Vanquish Neo nano-UHPLC via the NanoSpray Flex NG ion source (Thermo Scientific)

       Both instruments combine the advantages of the quadrupole for mass filtering and those of the Orbitrap for mass resolution and mass accuracy. The Orbitrap Exploris 240 offers higher resolution, mass accuracy and sensitivity, allowing a higher number of identifications in a reduced time interval and with a lower sample concentration.

       Among the applications of these systems are: the identification of proteins in complex proteomes and in samples of low and medium complexity; the differential quantitative analysis of proteomes (with isobaric or label free); the analysis of post-translational modifications, such as phosphorylations, and the selective quantification of proteins by means of targeted proteomics methods such as Selected Reaction Monitoring (SRM) and Multiple Reaction Monitoring (MRM).

GENOMICS

• The Facility has the necessary tools for basic studies of functional genomics and expression.
• Nanodrop ND-1000. UV-visible spectrophotometer which allows fast quantification of DNA, RNA, proteins, labeled nucleic acids and proteins, etc., consuming only 1-2 μl of sample.
• Tape Station 4250 Agilent: The 4150 TapeStation is an automated electrophoresis platform for quality control of nucleic acid samples. It enables fast, easy, and reliable analysis of DNA and RNA samples from up to 16 samples per run. This system is the ideal solution for quality control of RNA and DNA samples in next-generation sequencing (NGS), biobanks, or vaccine development workflows.
• 2 Thermocyclers for real-time PCR (rt-PCR): iQ5 (BIO-RAD) and LightCycler 96 (Roche). Detection by fluorescence (SYBR Green or specific probes) allowing the multiplex analysis of up to 5 fluorophores. 96 well plates are used. Its applications include relative quantification of gene expression, allelic discrimination (SNPs), absolute quantification, viral load determination, endpoint analysis, etc.