Difference between revisions of "Part:BBa K323214:Experience"

Latest revision as of 13:44, 26 September 2012

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Applications of BBa_K323214

User Reviews

[http://2012.igem.org/Team:Slovenia 2012 iGEM team Slovenia ] found out that NicTAL part deposited in the Registry by the Slovenian iGEM2010 team was missing subdomain in DNA-binding domain, so our team added this missing part on N terminal sequence. Apart from adding N terminal sequence, we added HIS tag on N terminal end and NLS on terminal C end of NicTAL12 (Figure 1). This construct was later used for designing TAL-based activator and repressor by adding VP16 and KRAB domain.

Figure 1:Sequence alignment of NicTAL10 and NicTAL12, showing differences between constructs.

Figure 2: Results of testing NicTAL:KRAB repressor demonstrated better binding ability of NicTAL12. HEK293T cells were cotransfected with NicTAL12:KRAB repressor under the control of a CMV promoter, and with a firefly luciferase reporter plasmid (BBa_K782023) containing 12 DNA-binding sites for NicTAL repressor upstream the CMV promoter. Check detailed results on [http://2012.igem.org/Team:Slovenia/TheSwitchDesignedTALregulators iGEM 2012 team Slovenia wiki].

UNIQc38a56bc48893b40-partinfo-00000000-QINU UNIQc38a56bc48893b40-partinfo-00000001-QINU

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 ===User Reviews===
-[http://2012.igem.org/Team:Slovenia 2012 iGEM team Slovenia ] discovered that [https://parts.igem.org/Part:BBa_K323214 NicTAL ] part deposited in the Registry by the Slovenian iGEM2010 team was missing subdomain in DNA-binding domain, so our team added this missing part on N terminal sequence. Apart from adding N terminal sequence, we added HIS tag on N terminal end and NLS on terminal C end of NicTAL12 (Figure 1).
+[http://2012.igem.org/Team:Slovenia 2012 iGEM team Slovenia ] found out that [https://parts.igem.org/Part:BBa_K323214 NicTAL ] part deposited in the Registry by the Slovenian iGEM2010 team was missing subdomain in DNA-binding domain, so our team added this missing part on N terminal sequence. Apart from adding N terminal sequence, we added HIS tag on N terminal end and NLS on terminal C end of NicTAL12 (Figure 1).  This construct was later used for designing TAL-based activator and repressor by adding [https://parts.igem.org/wiki/index.php?title=Part:BBa_K782066 VP16]  and [https://parts.igem.org/wiki/index.php?title=Part:BBa_K782011 KRAB]  domain.
 [[Image:Svn_12_NicTAL10vs12.PNG | 600 px]]
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-Results obtained by testing [https://parts.igem.org/Part:BBa_K782011#Characterization NicTAL12:KRAB] repressor proved that NicTAL12 binds to its binding sites better than NicTAL10.
 [[Image:Svn_12_NicTAL_graf.png]]
-'''Figure 2:''' Results of testing NicTAL12:KRAB repressor shows, that NicTAL12 binds to DNA better than NicTAL10.
+'''Figure 2:''' Results of testing  [https://parts.igem.org/wiki/index.php?title=Part:BBa_K782011 NicTAL:KRAB] repressor demonstrated better binding ability of NicTAL12. HEK293T cells were cotransfected with [https://parts.igem.org/Part:BBa_K782011 NicTAL12:KRAB] repressor under the control of a CMV promoter, and with a firefly luciferase reporter plasmid [https://parts.igem.org/Part:BBa_K782023 (BBa_K782023)] containing 12 DNA-binding sites for NicTAL repressor upstream the CMV promoter. Check detailed results on [http://2012.igem.org/Team:Slovenia/TheSwitchDesignedTALregulators iGEM 2012 team Slovenia wiki].