Difference between revisions of "Part:BBa K1926012"
 
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<partinfo>BBa_K1926012 short</partinfo>
 
<partinfo>BBa_K1926012 short</partinfo>
  
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This part is the VIKA UNIT of cyclebow system including an sv40 nuclear location sequence (NLS), a recombinase gene named VIKA and an sv40 terminator flanked by homodromous recognition site of recombinase VIKA named Vox. The whole part can be cut off by VIKA. This part is the improvement of the [[BBa_K1641003]], which is a plain VIKA.  
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This part is the VIKA UNIT of cyclebow system including an sv40 nuclear location sequence (NLS), a recombinase gene named VIKA and an sv40 terminator flanked by homodromous recognition site of recombinase VIKA named Vox. The whole part can be cut off by VIKA. This part is the improvement of the [[Part:BBa_K1641003]], which is a plain VIKA.  
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--[https://parts.igem.org/Part:BBa_K1641003]
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The VIKA with tags and RTSs are constructed using PCR and oligonucleotide ligation by the following procedure. Our UNITs all contain XbaI and SpeI enzyme sites, so we can easily line up the UNITs using 3A assembly.   
 
The VIKA with tags and RTSs are constructed using PCR and oligonucleotide ligation by the following procedure. Our UNITs all contain XbaI and SpeI enzyme sites, so we can easily line up the UNITs using 3A assembly.   
  
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[[File:5-连接过程改.png|800px|thumb|left|'''Figure 1:''' Three steps to construct our UNITs using PCR and oligonucleotide ligation.]]
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[[File:5-连接过程改.png|600px|thumb|left|'''Figure 1:''' Three steps to construct our UNITs using PCR and oligonucleotide ligation.]]
 
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===Gel analysis after EcoR1 digestion===
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The length of our biobricks are confirmed by gel analysis after EcoR1 digestion. The biobrick BBa_R0040, which is the negative control in 2016’s interlab, was used as a control because it only have a 54 bp part.
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[[File: 2016--SYSU-CHINA--biobrick2.png|400px|thumb|left|'''Figure 2:''' AGE image of biobricks BBa_K1926001-BBa_K1926003. The samples were pre dyed with gelred]]
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Latest revision as of 07:29, 19 October 2016

The VIKA UNIT: recombinase VIKA flanked by Vox

This part is the VIKA UNIT of cyclebow system including an sv40 nuclear location sequence (NLS), a recombinase gene named VIKA and an sv40 terminator flanked by homodromous recognition site of recombinase VIKA named Vox. The whole part can be cut off by VIKA. This part is the improvement of the Part:BBa_K1641003, which is a plain VIKA.


Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]


Usage

You may use this part to:

1) Test the cut-off efficiency of recombinase VIKA by qRT-PCR;

2) Fast cut off this part by transient transfecting recombinase gene into nucleus;

3) Do lineage tracking.


Source

The recombinase gene VIKA and its RTS Vox were got through denovo synthesis. NLS and sv40 terminator were got from commercial plasmid: pentry nls GFP/pcdn3.1.


Construction Design

The VIKA with tags and RTSs are constructed using PCR and oligonucleotide ligation by the following procedure. Our UNITs all contain XbaI and SpeI enzyme sites, so we can easily line up the UNITs using 3A assembly.

Figure 1: Three steps to construct our UNITs using PCR and oligonucleotide ligation.



Gel analysis after EcoR1 digestion

The length of our biobricks are confirmed by gel analysis after EcoR1 digestion. The biobrick BBa_R0040, which is the negative control in 2016’s interlab, was used as a control because it only have a 54 bp part.

Figure 2: AGE image of biobricks BBa_K1926001-BBa_K1926003. The samples were pre dyed with gelred