Part:BBa_K2644122

GGG+crRNA EGFP（lbCas12a）65289;

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  COMPATIBLE WITH RFC[21]
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  COMPATIBLE WITH RFC[1000]

Usage and Biology

This biological part is specially designed to target a EGFP fragment. The principle behind this is that CRISPR-Cas12a (Cpf1) proteins are RNA-guided enzymes that bind and cut DNA as components of bacterial adaptive immune systems like CRISPR-Cas9. We designed the crRNA which is able to combine with the Cas12a protein and recognize the target DNA. According to this principle, we designed a crRNA that matched with the EGFP fragment., which brings Cas12a to its target, specifically cleaves the EGFP fragment. Through this method, we are theoretically capable of targeting any single base mutation on ctDNA which are dispersed in people’s blood.

Cleaving experiment

Figure 1.The FnCas12a trans-cleavage activity on ssDNA. Line M, marker. Line 1, dsDNA about 40 bps. Line 2, ssDNA about 40 nts. Line 3, cleavage according to: dsDNA : ssDNA = 1:20. Line 4, cleavage according to: dsDNA : ssDNA = 1:40. Line 5, cleavage according to: dsDNA : ssDNA = 1:80. Line 6, cleavage according to: dsDNA : ssDNA = 1:100. Line 7, cleavage according to: dsDNA : ssDNA = 1:125.