Difference between revisions of "Part:BBa K4585008"
 
Line 1: Line 1:
−
 
 
__NOTOC__
 
__NOTOC__
 
<partinfo>BBa_K4585008 short</partinfo>
 
<partinfo>BBa_K4585008 short</partinfo>
  
−
The objective was to synthesize the pGL4.35-3 HA-9 GAL4UAS-KRAB-NLS plasmid by homologous recombination with the KRAB homologous recombination fragment .  
+
The objective was to synthesize the pGL4.35-3×HA-9×GAL4UAS-KRAB-NLS plasmid by homologous recombination with the KRAB homologous recombination fragment.  
  
 
<html>
 
<html>
Line 20: Line 19:
 
             <div class="pageContent-main__textBox">
 
             <div class="pageContent-main__textBox">
 
                 <!--all the content must included in this div-->
 
                 <!--all the content must included in this div-->
−
                 <p>We based on the sequence of the pGL4.35 plasmid. Through homologous recombination experiments, the Gal 4 sequence, 3× HA sequence were ligated to the pGL4.35 plasmid, and nine UAS sequences were inserted simultaneously. Special primers were designed, and then the plasmid was cut into linear sequences of a length of about 4500 bp by PCR. The two ends of the linear sequence can be complementary to both ends of the KRAB homologous recombination fragment, and the purpose is to perform homologous recombination with the KRAB homologous recombination fragment to obtain the target product pGL4.35-3×HA-9×GAL4UAS-KRAB-NLS plasmid.
+
                 <p>We based on the sequence of the pGL4.35 plasmid. Through homologous recombination experiments, the GAL4 sequence, 3×HA sequence were ligated to the pGL4.35 plasmid, and nine UAS sequences were inserted simultaneously. Special primers were designed, and then the plasmid was cut into linear sequences of a length of about 4500 bp by PCR. The two ends of the linear sequence can be complementary to both ends of the KRAB homologous recombination fragment, and the purpose is to perform homologous recombination with the KRAB homologous recombination fragment to obtain the target product pGL4.35-3×HA-9×GAL4UAS-KRAB-NLS plasmid.
 
                 </p>
 
                 </p>
 
                 <!--put text here, <p>content</p>-->
 
                 <!--put text here, <p>content</p>-->
Line 26: Line 25:
 
             <h2 class="pageContent-main__title pageContent-main__subtitle">
 
             <h2 class="pageContent-main__title pageContent-main__subtitle">
 
                 <!--class="pageContent-main__title" means it is the sub title-->
 
                 <!--class="pageContent-main__title" means it is the sub title-->
−
                 1 Pattern Diagram
+
                 1 Diagrams
 
             </h2>
 
             </h2>
 
             <div class="pageContent-main__textBox">
 
             <div class="pageContent-main__textBox">
Line 34: Line 33:
 
                 <br />
 
                 <br />
 
                 <!--put image's url here-->
 
                 <!--put image's url here-->
−
                 <p style="width: 80%;text-align:center;font-size: .9rem; margin: -1rem auto 1rem auto; color: #888;">Fig.1 The agarose gel electrophoresis of pGL4.35-3×HA-9×GAL4UAS-KRAB-NLS linearized vector</p>
+
                 <p style="width: 80%;text-align:center;font-size: .9rem; margin: -1rem auto 1rem auto; color: #888;">Fig 1. The agarose gel electrophoresis of pGL4.35-3×HA-9×GAL4UAS-KRAB-NLS linearized vector</p>
−
            </div>
+
         
−
            <h2 class="pageContent-main__title pageContent-main__subtitle">
+
−
                <!--class="pageContent-main__title" means it is the sub title-->
+
−
2 Experiment
+
−
            </h2>
+
−
            <h2 class="pageContent-main__title pageContent-main__subtitle">
+
−
                <!--class="pageContent-main__title" means it is the sub title-->
+
−
                2.1 Method
+
−
            </h2>
+
−
            <div class="pageContent-main__textBox">
+
−
                <!--all the content must included in this div-->
+
−
                <p>Based on the sequence of the pGL4.35-3 plasmid, we designed the pGL4.35-3×HA-9×GAL4UAS-KRAB-NLS linearized vectror by using the GAL 4 homologous recombination fragment, 9×UAS homologous recombination fragment and homologous recombination experiments.
+
−
                </p>
+
−
                <!--put text here, <p>content</p>-->
+
−
            </div>
+
−
            <h2 class="pageContent-main__title pageContent-main__subtitle">
+
−
                <!--class="pageContent-main__title" means it is the sub title-->
+
−
                2.2 Results
+
−
            </h2>
+
−
            <div class="pageContent-main__textBox">
+
−
                <!--all the content must included in this div-->
+
−
                <p style="text-align: center;">
+
−
                    <img width="400px" src="https://static.igem.wiki/teams/4585/wiki/the-model-diagram-of-pgl4-35-3-ha-9-gal4uas-krab-nls.png"></p>
+
−
                <br />
+
−
                <!--put image's url here-->
+
−
                <p style="width: 80%;text-align:center;font-size: .9rem; margin: -1rem auto 1rem auto; color: #888;">Fig.1 The model diagram of pGL4.35-3×HA-9×GAL4UAS-KRAB-NLS</p>
+
 
             </div>
 
             </div>
 
             <h2 class="pageContent-main__title pageContent-main__subtitle">
 
             <h2 class="pageContent-main__title pageContent-main__subtitle">
 
                 <!--class="pageContent-main__title" means it is the sub title-->
 
                 <!--class="pageContent-main__title" means it is the sub title-->
−
                 3.Caution
+
                 2 Caution
 
             </h2>
 
             </h2>
 
             <div class="pageContent-main__textBox">
 
             <div class="pageContent-main__textBox">

Latest revision as of 03:47, 12 October 2023

pGL4.35-3xHA-9xGAL4UAS-KRAB-NLS linearized vector

The objective was to synthesize the pGL4.35-3×HA-9×GAL4UAS-KRAB-NLS plasmid by homologous recombination with the KRAB homologous recombination fragment.

pGL4.35-3×HA-9×GAL4UAS-KRAB-NLS linearized vector

We based on the sequence of the pGL4.35 plasmid. Through homologous recombination experiments, the GAL4 sequence, 3×HA sequence were ligated to the pGL4.35 plasmid, and nine UAS sequences were inserted simultaneously. Special primers were designed, and then the plasmid was cut into linear sequences of a length of about 4500 bp by PCR. The two ends of the linear sequence can be complementary to both ends of the KRAB homologous recombination fragment, and the purpose is to perform homologous recombination with the KRAB homologous recombination fragment to obtain the target product pGL4.35-3×HA-9×GAL4UAS-KRAB-NLS plasmid.

1 Diagrams


Fig 1. The agarose gel electrophoresis of pGL4.35-3×HA-9×GAL4UAS-KRAB-NLS linearized vector

2 Caution

The two ends of the pGL4.35-3×HA-9×GAL4UAS-KRAB-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 4423
    Illegal XbaI site found at 151
    Illegal XbaI site found at 1933
    Illegal XbaI site found at 4260
    Illegal XbaI site found at 4495
    Illegal SpeI site found at 4019
    Illegal PstI site found at 3108
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal EcoRI site found at 4423
    Illegal NheI site found at 4451
    Illegal SpeI site found at 4019
    Illegal PstI site found at 3108
    Illegal NotI site found at 3087
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal EcoRI site found at 4423
    Illegal BglII site found at 4498
    Illegal BamHI site found at 413
    Illegal XhoI site found at 4303
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal EcoRI site found at 4423
    Illegal XbaI site found at 151
    Illegal XbaI site found at 1933
    Illegal XbaI site found at 4260
    Illegal XbaI site found at 4495
    Illegal SpeI site found at 4019
    Illegal PstI site found at 3108
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal EcoRI site found at 4423
    Illegal XbaI site found at 151
    Illegal XbaI site found at 1933
    Illegal XbaI site found at 4260
    Illegal XbaI site found at 4495
    Illegal SpeI site found at 4019
    Illegal PstI site found at 3108
    Illegal NgoMIV site found at 55
    Illegal NgoMIV site found at 166
    Illegal NgoMIV site found at 1644
    Illegal NgoMIV site found at 1661
    Illegal NgoMIV site found at 1793
    Illegal NgoMIV site found at 1884
    Illegal NgoMIV site found at 2136
    Illegal AgeI site found at 1939
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal SapI site found at 434
    Illegal SapI site found at 2145