Difference between revisions of "Part:BBa K5453004"
Line 19: Line 19:
 
We employed the CRISPRi technology to simultaneously inhibit the pfkA and zwf genes, introducing random mutations at the 7th and 8th nucleotide positions of their sgRNA, and constructed an R6K-dCas9-zwf-pfkA mutant library using Golden Gate technology.
 
We employed the CRISPRi technology to simultaneously inhibit the pfkA and zwf genes, introducing random mutations at the 7th and 8th nucleotide positions of their sgRNA, and constructed an R6K-dCas9-zwf-pfkA mutant library using Golden Gate technology.
  
−
 
+
<html>
−
<center><html>< img src ="https://static.igem.wiki/teams/5453/engineering-success/engineering-success-9.png" width = "50%"><br></html></center>
+
        <div class="col-lg" style="margin:auto;text-align:center;">
−
<center>Figure 1:PCR Results Figure. A: Gel electrophoresis image of PCR amplification for sgRNA-pfkA-M, sgRNA-zwf-M. B: Gel electrophoresis image of colony PCR performed on 10 randomly picked colonies from the plate. C: Gel electrophoresis image of plasmid digestion with KpnI and XhoI enzymes.D: Sequencing of the correct plasmids verified by colony PCR and enzyme digestion.</center>
+
                <img style="margin:20px auto 5px auto;" src="https://static.igem.wiki/teams/4630/wiki/parts/bba-k4630100-stgrna-barcode-cassette-1/1-part-stgrna-barcode-cassette1.svg" width="80%">
 +
                <p style="color:Gray; padding:0px 30px 10px;">Figure B1. DNA manipulating methods from the literatures</p>
 +
        </div>
 +
</html>

Revision as of 11:07, 29 September 2024


dcas9-sgRNA-pfkA-lac operator-lacI promoter-sgRNA-zwf-lac operator-lacI promoter

dcas9-sgRNA-pfkA-lac operator-lacI promoter-sgRNA1-zwf-lac operator-lacI promoter

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal NheI site found at 1096
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BamHI site found at 3375
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]


We employed the CRISPRi technology to simultaneously inhibit the pfkA and zwf genes, introducing random mutations at the 7th and 8th nucleotide positions of their sgRNA, and constructed an R6K-dCas9-zwf-pfkA mutant library using Golden Gate technology.

Figure B1. DNA manipulating methods from the literatures