Journal: Cancers

Article Title: Gene Editing with CRISPR/Cas Methodology and Thyroid Cancer: Where Are We?

doi: 10.3390/cancers14030844

Figure Lengend Snippet: The basic components CRISPR/Cas9 system. ( A ) The crRNA associated with tracrRNA enters Cas9 endonuclease and guides the protein throughout the foreign DNA in search of a full-complementarity region adjacent to a PAM sequence in order to produce the double-strand break. ( B ) Detailed view of interaction of crRNA with the target DNA sequence (complementary sequence) and PAM sequence in the non-complementary strand of DNA.

Article Snippet: The Zhang Lab [ ] constructed these plasmids, which are available in Addgene as PX459 or PX458, respectively, for Cas9 + puromycin resistance, or Cas9 + GFP.

Techniques: CRISPR, Sequencing

Journal: Cancers

Article Title: Gene Editing with CRISPR/Cas Methodology and Thyroid Cancer: Where Are We?

doi: 10.3390/cancers14030844

Figure Lengend Snippet: Cas9 and the mutated variants used for gene editing and gene modulation.

Article Snippet: The Zhang Lab [ ] constructed these plasmids, which are available in Addgene as PX459 or PX458, respectively, for Cas9 + puromycin resistance, or Cas9 + GFP.

Techniques: Imaging

Journal: Cancers

Article Title: Gene Editing with CRISPR/Cas Methodology and Thyroid Cancer: Where Are We?

doi: 10.3390/cancers14030844

Figure Lengend Snippet: Gene modulation with CRISPR/dCas9 system. Dead Cas9 (dCas9) as a tool to modulate gene expression without gene editing. ( A ) In CRISPRi (interference) system, dCas9 is fused to transcriptional inhibitors such as KRAB that blocks transcription of sgRNA targeted region; ( B ) In the CRISPRa (activation) system, dCas9 is fused to transcriptional activators such as VP64, RTA and P65 to induce the expression of sgRNA targeted region.

Article Snippet: The Zhang Lab [ ] constructed these plasmids, which are available in Addgene as PX459 or PX458, respectively, for Cas9 + puromycin resistance, or Cas9 + GFP.

Techniques: CRISPR, Gene Expression, Activation Assay, Expressing

Journal: Cancers

Article Title: Gene Editing with CRISPR/Cas Methodology and Thyroid Cancer: Where Are We?

doi: 10.3390/cancers14030844

Figure Lengend Snippet: Plasmidial system to express CRISPR/Cas9 system. ( A ) CRISPR/Cas9 plasmid contains a site for BbsI digestion that creates the cloning site for the 20-nt sequence sgRNA; ( B ) The sgRNA sequence is ligated into BbsI-digested plasmids PX458 (GFP) or PX459 (puromycin resistance) as annealed DNA oligonucleotides with cohesive ends as shown in the figure CACC in the 5′ and CAAA in the 3′.

Article Snippet: The Zhang Lab [ ] constructed these plasmids, which are available in Addgene as PX459 or PX458, respectively, for Cas9 + puromycin resistance, or Cas9 + GFP.

Techniques: CRISPR, Plasmid Preparation, Cloning, Sequencing

Journal: Cancers

Article Title: Gene Editing with CRISPR/Cas Methodology and Thyroid Cancer: Where Are We?

doi: 10.3390/cancers14030844

Figure Lengend Snippet: Editing protein-coding genes with CRISPR/Cas9. Targeting protein coding genes with CRISPR/Cas9-mediate gene editing. The objective is to disrupt the coding sequence using sgRNAs targeting the first exons of the gene, close to the start codon “ATG”. As a result, loss of protein expression is expected due to deletions or insertions that may disrupt the start codon, change protein frame or even insert a premature stop codon.

Article Snippet: The Zhang Lab [ ] constructed these plasmids, which are available in Addgene as PX459 or PX458, respectively, for Cas9 + puromycin resistance, or Cas9 + GFP.

Techniques: CRISPR, Sequencing, Expressing

Journal: Cancers

Article Title: Gene Editing with CRISPR/Cas Methodology and Thyroid Cancer: Where Are We?

doi: 10.3390/cancers14030844

Figure Lengend Snippet: Editing non-coding genes with CRISPR/Cas9. Targeting non-coding genes with CRISPR/Cas9. In the figure the non-coding miRNA gene is represented and the sgRNAs target structural regions of the primary miRNA structure that are necessary for miRNA processing by DROSHA and DICER endonucleases.

Article Snippet: The Zhang Lab [ ] constructed these plasmids, which are available in Addgene as PX459 or PX458, respectively, for Cas9 + puromycin resistance, or Cas9 + GFP.

Techniques: CRISPR

Journal: Cancers

Article Title: Gene Editing with CRISPR/Cas Methodology and Thyroid Cancer: Where Are We?

doi: 10.3390/cancers14030844

Figure Lengend Snippet: Applications of CRISPR/Cas9 gene editing in thyroid cancer.

Article Snippet: The Zhang Lab [ ] constructed these plasmids, which are available in Addgene as PX459 or PX458, respectively, for Cas9 + puromycin resistance, or Cas9 + GFP.

Techniques: CRISPR, Over Expression, Mutagenesis, Migration, Gene Expression, In Vitro, In Vivo, Cell Differentiation