Part:BBa_K4099000

sgRNA

Usage and Biology

BBa_K4099000 is a piece of RNAs that function as guides for RNA- or DNA-targeting enzymes, which they form complexes with.

Figure1. Principle diagram of CRISPR-Cas9..

The sgRNA and HR are inserted in the pLCNICK vector. (Figure 3).

Proof of function

We electrotransformed the plasmid pIB165 as the foreign DNA to test the transformation efficiency of our modified L. casei, it took several days to culture and finally saw the comparison results. As showing above, we can see that the modified L. casei (KO) has higher transformation efficiency with remarkably more colonies than the wild (Wild).

1.Transformation test result

Figure 2. Comparison between the wild L. casei (left) and modified L. casei (right, LSEI-2094 knocked out) in transformatio..

Graph 3. Comparison between the wild L. casei and modified L. casei in transformation

Figure 3. Histogram comparison between wild strain and KO strain..

In addition, we measured OD600 of these strain groups which were pre-spread plates with different volumes of bacteria solutions so as to quantify the transformation results as showing above (Fig. 3). In conclusion, it indicates that our modified L. casei has much higher efficiency of the foreign plasmid transformation than the wild and the modified L. casei has great potential to be used as the recombinant carrier in various areas.

Sequence and Features

Assembly Compatibility:

• 10
  INCOMPATIBLE WITH RFC[10]

  Illegal XbaI site found at 107
  
• 12
  COMPATIBLE WITH RFC[12]
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal BglII site found at 104
  
• 23
  INCOMPATIBLE WITH RFC[23]

  Illegal XbaI site found at 107
  
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal XbaI site found at 107
  
• 1000
  COMPATIBLE WITH RFC[1000]