Part:BBa_K3402058
Triple-gene editing cassette
This device is composed of up-doLeu (BBa_K3402048), sgLeu (BBa_K3402049), 50bp-upPXA1 (BBa_K3402037), hph (BBa_K3402012), 50bp-doPXA1 (BBa_K3402038), Ptef1 (BBa_K3402007), sgPXA1 (BBa_K3402039), sgGFP (BBa_K3402024), Tsyn7 (BBa_K3402001), upGFP (BBa_K3402025), doGFP (BBa_K3402026).
Usage and Biology
Based on Pxa1, GFP expression vector, designed the sgRNA that targeted at Leu site. We construct the Pxa1, GFP, Leu editing expression cassette below, and transform it into the recombinant strain.
The transformants were cultured on YPD plus hygromycin plates. Then the positive transformants were conducted green fluorescence observation. After that, the positive transformants without fluorescence were dotted on the Complete Medium (CM) and Minimal Medium (MM). Then the colony grown on the Complete Medium but did not grow on the Minimal Medium were the right transformants.
The triple gene-editing efficiency was 30%.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21INCOMPATIBLE WITH RFC[21]Illegal XhoI site found at 776
Illegal XhoI site found at 3223 - 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal NgoMIV site found at 174
Illegal NgoMIV site found at 203 - 1000INCOMPATIBLE WITH RFC[1000]Illegal SapI.rc site found at 3859
None |