Microbial Culturing & Engineering Core

About

The Microbial Culturing & Engineering Core provides physicians and investigators services for culturing and genetic engineering of bacteria in concordance with other mechanistic studies related to the microbiome. The facility offers services such as the characterization and verification of bacterial isolates, culture-based biochemical assays involving isolates or defined consortia to screen for specific microbial functions through access to an expanding library of human derived bacterial isolates. The facility can help in anaerobic processing of complex samples, such as stool or mucus, and the generation of gene knock-out or knock-in mutants of bacterial isolates.




Contacts

Mohammed  Dwidar, PhD

Mohammed Dwidar, PhD
Core Director
Project Staff
dwidarm@ccf.org

Akeem  Santos

Akeem Santos
Technician
Location:NE1-221
Phone:(216) 444-0534
santosa4@ccf.org

Services

  • All equipment and workstations are available to researchers after appropriate training.
  • Free consultation: Investigators are highly encouraged to meet with facility staff to discuss their objectives & experimental design related to culturing, isolating or genetically engineering the bacteria prior to purchasing bacteria/reagents. The facility can provide specific protocols for the preparation of general or specialized media, as well as for the cultivation and engineering of bacteria.
  • Basic characterization and cryogenic storage of acquired isolates.
  • Providing specific isolates from our in-house culture collection for investigator-initiated studies.
  • Providing pre-made synthetic consortium of bacteria for mice transplantation studies or for biochemical screening.
  • Help in collecting and processing donor fecal or other samples for either future transplant studies or future preparation of bacterial isolates library.
  • Preparing activity-guided library of bacterial isolates.
    • Samples can come from various host-derived sources that include stool, urine, mucus, skin, oral cavity, etc. Media will be designed according to the need and the sample will be plated. Colonies will be picked and pure cultures will be archived at -80°Cif needed.
    • Function or taxonomy of selected isolates can be tested through growth assays, 16S rDNA analysis, mass spectrometry or PCR using gene specific primers.
  • Antibiotic susceptibility tests for the selected isolates.
  • Other in vitro assays including bacterial invasion / adhesion, metabolic analyses and kinetic assays can be pursued when needed.
  • Generating gene knock-out mutants in the selected strains.
  • Chromosomal expression of genes (induced expression, or constitutive overexpression) in the selected strains.
  • Induced or constitutive expression of genes in suitable replicating plasmids.
  • Color selected strains through plasmid-coded fluorescent proteins for biofilm studies or other experiments.
  • Future engineering work will involve transposon mutagenesis.

Notes

  • Many projects to be conducted with the facility will be long-term. As such, it is typically more cost effective for investigators to include facility staff as co-investigators on grants rather than using the facility as fee-for-service. However, it is completely up to the investigator to decide the best way to collaborate with the facility.
  • The facility will obtain the necessary approvals from the Institutional Biosafety Committee (IBC) to perform the required bacterial culturing and engineering work. However, handling bacteria to investigators as part of any research project will require approval from the IBC. As such, it is the responsibility of the investigator to obtain the appropriate IBC approvals prior to handling any bacteria.
  • Many of the bacteria and other reagents/tools at the facility were obtained through material transfer agreements with outside vendors. As such, any potential commercial use of products resulting from research with materials from the facility must comply with the agreements on hand.

Equipment

  • 5 Coy anaerobic chambers each with a capacity to hold >500 petri plates, for use in the isolation or cultivation of anaerobic bacteria, or the anaerobic preparation of stool/microbial samples for transplant.
  • Specialized gas mixtures that allow growth of fastidious bacteria that require defined levels of oxygen.
  • A library of multi-species bacterial isolates (in preparation).
  • Aerobic biosafety cabinet for the isolation or cultivation of aerobic bacteria.
  • Coy Co-Culture Chamber for the growth of anaerobic bacteria along with adherent mammalian cells.
  • CDC biofilm reactor for biofilm studies.

Grant Information

The Microbial Culturing and Engineering facility provides investigators with a centralized resource for studies that involve the isolation, identification, and culturing of host-associated bacteria and/or the genetic engineering of bacteria. The facility is equipped with five Coy anaerobic chambers, each with a 500 culture plate capacity. The facility has additional biosafety cabinets and incubators for work with aerobic bacteria. This system allows for the culturing and maintenance of diverse host-associated bacteria with chambers allocated for specialized gas mixtures, spore-forming bacteria, as well as for initial isolations. The facility maintains a collection of multi-species-derived bacteria either acquired from other culture collections or isolated from different host sites as needed by investigators. The services provided by the culturing and engineering facility enables investigators to quantify specific groups of bacteria through culture-based methods, isolate target bacteria based on taxonomy and or metabolic functions, and genetically engineer bacteria to knock-out or knock-in genes to perform mechanistic host-microbe studies.

Microbial Culturing & Engineering Core

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