Difference between revisions of "Part:BBa K4585009"
Line 73: Line 73:
 
</body>
 
</body>
 
</html>
 
</html>
 +
 +
<!-- -->
 +
<span class='h3bb'>Sequence and Features</span>
 +
<partinfo>BBa_K4585009 SequenceAndFeatures</partinfo>

Revision as of 11:41, 9 October 2023


pcDNA3.1(+)-3xHA-Gal4-VP64-Degron-NLS linearized vector

The objective is to perform homologous recombination with the Degron homologous recombination fragment , The synthetic pcDNA3.1 (+) -3 HA-Gal 4-VP64-Degron-NLS plasmid was used for subsequent experiments.

pcDNA3.1(+)-3×HA-Gal4-VP64-Degron-NLS linearized vector

We based on the sequence of the pcDNA3.1(+)-3×HA-Gal4-VP64-NLS plasmid. Special primers were designed to cut the plasmid into a linear sequence of length of about 6237bp by PCR. The two ends of this linear sequence can be complementary to the Degron homologous recombination fragment of the homologous Degron fragment for homologous recombination to obtain the target product pcDNA3.1(+)-3×HA-Gal4-VP64-Degron-NLS plasmid.

1 Pattern Diagram


Fig.1 The agarose gel electrophoresis of pcDNA3.1(+)-3×HA-Gal4-VP64-Degron-NLS linearized vector

2 Experiment

2.1 Method

Based on the designed pcDNA3.1 (+) -3 HA-Gal 4-VP64-NLS plasmid , Design of the new pcDNA3.1 (+) -3 HA-Gal 4-VP64-Degron-NLS linearized vectror

2.2 Results


Fig.1 The model diagram of pcDNA3.1(+)-3×HA-GAL4-VP64-Degron-NLS

3.Caution

The two ends of the pcDNA3.1(+)-3×HA-Gal4-VP64-Degron-NLS linearized vectror need to be complementary to the two ends of the corresponding recombinant fragments to ensure the success of homologous recombination ligation.

Sequence and Features


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal EcoRI site found at 25
    Illegal XbaI site found at 64
    Illegal XbaI site found at 6170
    Illegal SpeI site found at 4828
    Illegal PstI site found at 30
    Illegal PstI site found at 1459
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal EcoRI site found at 25
    Illegal SpeI site found at 4828
    Illegal PstI site found at 30
    Illegal PstI site found at 1459
    Illegal NotI site found at 51
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal EcoRI site found at 25
    Illegal BglII site found at 4591
    Illegal BamHI site found at 6027
    Illegal BamHI site found at 6216
    Illegal XhoI site found at 58
    Illegal XhoI site found at 5797
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal EcoRI site found at 25
    Illegal XbaI site found at 64
    Illegal XbaI site found at 6170
    Illegal SpeI site found at 4828
    Illegal PstI site found at 30
    Illegal PstI site found at 1459
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal EcoRI site found at 25
    Illegal XbaI site found at 64
    Illegal XbaI site found at 6170
    Illegal SpeI site found at 4828
    Illegal PstI site found at 30
    Illegal PstI site found at 1459
    Illegal NgoMIV site found at 569
    Illegal NgoMIV site found at 1910
    Illegal NgoMIV site found at 2195
    Illegal AgeI site found at 122
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI site found at 5716
    Illegal BsaI.rc site found at 3723
    Illegal BsaI.rc site found at 5461
    Illegal SapI site found at 2640
    Illegal SapI.rc site found at 1759
    Illegal SapI.rc site found at 1969