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COURSE MATERIALS
Unboxing CRISPR
for transdisciplinary learning
Materials: Labwork
Inside a CRISPR Lab
(6’37’’)Video on the work of researchers at UC Berkeley using CRISPR. The video allows you to take a peek behind the scenes to see what goes into a human genome editing experiment and follow the process
General precautions when handling RNA
A Text with illustration explaining: a) general concepts and function of RNA, DNA, RNAase; degradation of RNA; b) the major sources of RNase contamination in a typical laboratory; c) precautions to avoid RNase contamination.
Cutting DNA
This is a basic protocol to cut DNA into smaller fragments. The cut is nonspecific, being an excellent example of contrast with the specificity of CRISPR/cas9.
Electrophoresis
This is a protocol to do an agarose gel electrophoresis, a standard lab procedure used to separate macromolecules based on size. The technique applies a negative charge so proteins move towards a positive charge. Electrophoresis is used extensively in DNA, RNA, and protein analysis.
In silico lab work: CRISPR cas9 gene editing, Fábio Ferreira
(28’47’’) Fábio Júnio Ferreira, i3S researcher, introduces how to design a sgRNA, particularly to edit Streptomycetes and zebrafish.
Guide for the synthesis of single-guide RNA (sgRNA)
Document with information, tools, and parameters to design sgRNA.
Synthesis of single guide RNA
(120’) This is a lab protocol of sgRNAs synthesis (guide RNA) using T7 polymerase for Higher Education students (used under the supervision of someone with experience in molecular biology lab techniques). This is a protocol to do the synthesis of a sgRNA with the EnGen sgRNA Synthesis kit (in vitro) and confirm the RNA synthesis by running sgRNA in a gel.
Purification of the sgRNAs using a commercial purification kit
This is a Lab protocol to purify RNA using a commercial purification kit and run the purified sgRNA to confirm the recovery of RNA and quantification. This is a Lab protocol used in CRISPR–Cas9 genome-editing technology and can be applied as a practical lab exercise for STEAM students.
Agarose Gel Electrophoresis
(75’)This is a protocol to do an agarose gel electrophoresis, a standard lab procedure that can be used to resolve linear RNA/DNA fragments on the basis of their molecular weight (length in base pairs) for visualization.
In vitro cleavage
This protocol guides students /teachers in the process of in vitro cleavage of target DNA while editing a genome with CRISPR-Cas9. The tool can be applied as a practical lab exercise for STEAM students.