Functional characterization of musculoskeletal genes by CRISPR/Cas9 mediated genome editing in zebrafish
Israel (FIMS) - Bar Ilan University, Safed
Genetics of Musculoskeletal Disease Lab
David Karasik, PhD
Chen Shochat, PhD
English, Hebrew, Portuguese
4 weeks
Cities/Months Jan Feb Mar Apr May Jun Jul Augt Sep Oct Nov Dec
No No No No No No Yes Yes No No No No
Type of Research Project
- Basic science
What is the background of the project?
Bone strength and muscle mass share genetic determinants; therefore, genes with potentially pleiotropic effects exist and have to be functionally validated. The aim of our studies is to develop an innovative evolution-based approach in teleost fish models to functionally characterize novel bone relevant genes that were earlier identified by us in a genome-wide association study (GWAS) for pleiotropic musculoskeletal (MSK) factors. Teleost fish are models for studying both evolutionary bone biology and to mimic human bone diseases. We use three fish species (incl. zebrafish), which are encompassing the spectrum of ray-finned fish evolution. The chosen fish models offer unique opportunities for gene modification by CRISPR/Cas9 technologies followed by gross morphological and live imaging of dynamic bone cell behavior. This will allow to identify key players involved in teleost bone homeostasis and also provide evidence for roles these genes play in human bones in norm and disease. By identifying commonalities as well as differences in the genetic control of bone formation in distant fish species, we will obtain important insight into the functional conservation or divergence of regulatory pathways.
What is the aim of the project?
Analysis of expression patterns of novel candidate genes identified by human musculoskeletal GWAS studies in bones of the zebrafish;Characterization of the activity of novel bone genes for bone maintenance and repair, by live observation, imaging and muscle histology
What techniques and methods are used?
Real Time quantitative polymerase chain reaction (RT-qPCR) for candidate gene's expression in bones of the embryonic & adult zebrafish;live imagingof embryos (they are transparent) and muscle histology in adults (staining for bones and muscles, respectively)
What is the role of the student?
- The tasks will be done under supervision
What are the tasks expected to be accomplished by the student?
Perform Real Time quantitative polymerase chain reaction (RT-qPCR) for expression of pre-specified genes in bones of the embryonic & adult zebrafish, incl. Housekeeping gene(s);perform quantitative analysis; participate in the live imaging of embryos(perform staining for bones with photographing) Perform muscle histology in adults (staining and microtome cuts; imaging and analyzing; perform quantitative analysis between genotype groups)
Will there be any theoretical teaching provided (preliminary readings, lectures, courses, seminars etc)
Yes Preliminary readings, journal club(s), seminars in the lab
What is expected from the student at the end of the research exchange? What will be the general outcome of the student?
- The student will prepare a poster
- The student will prepare a presentation
- The student will prepare a scientific report
- The student will prepare an abstract
- The student will have the opportunity to present the results together with the supervisor at a conference
What skills are required of the student? Is there any special knowledge or a certain level of studies needed?
Insights into molecular biology and animal modeling. Needed Bachelors in Biology or equivalent. Previous experience with: molecular biology, PCR.
Are there any legal limitations in the student’s involvement
Handling of animals needs an Animal Committee approval
Type of students accepted
This project accepts:
- Graduated students (less than 6 months)
- Students in biomedical fields
- Karasik D; Rivadeneira F; Johnson M. L. Genetics of Bone Mass and Susceptibility to Bone Diseases. Nat Rev Rheumat. 2016; 12: 323–334
- Khajuria; D. K.; Kumar; V.; Karasik; D*; Gedanken; A*. Fluorescent nanoparticles with tissue dependent affinity for live zebrafish imaging. ACS Applied Materials & Interfaces 2017; 9 (22); pp 18557–18565