Difference between revisions of "Part:BBa K4268000:Experience"

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===Applications of BBa_K4268000===
 
===Applications of BBa_K4268000===
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With the generosity of IDT, SUNY_Oneonta iGEM synthesized the Head-Tail Connector part. This part is RCF1000 compatible and contains SapI sites at either end of the sequence. The part was resuspended, restriction digested, and ligated into PSB1C00. The complex was then transformed into DH5&a cells and was left to grow overnight. After white colony growth was discovered in the transformed cells, five white colonies underwent a Colony PCR to determine if the desired insert was absorbed. Additionally, the selected colonies were restreaked.
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The part was received, resuspended, and cloned into the level 0 Golden Gate vector, PSB1C00. The ligation was then transformed into DH5&alpha cells. After overnight growth, white colonies (indicative of a replacement of the RFP reporter cassette) were subjected to screening for the correct insert using colony PCR with the VF/VR primer pair.
  
  
 
Insert gel figure
 
Insert gel figure
  
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Caption: Colony PCR of five colonies suspected of coating the insert and one colony containing an RFP. The target length is 1573bp.
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Caption: Colony PCR of five colonies (clones) suspected of coating the insert Head-tail connector insert. The insert length is 1573bp and the predicted size of the PCR product using VF/VR primers is 1739 bp.
  
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The gel indicates that all five colonies are likely to contain the correct insert and thus, the Head-Tail Connector was successfully cloned into a Level O Golden Gate Assembly basic part.
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The gel indicates that all five colonies are likely to contain the insert and thus, the Head-Tail Connector was successfully cloned into a Level O Golden Gate Assembly basic part. The confirmed colonies underwent further purification to prepare them for Level 1 Golden Gate Assembly.
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===User Reviews===
 
===User Reviews===

Revision as of 04:06, 1 October 2022


This experience page is provided so that any user may enter their experience using this part.
Please enter how you used this part and how it worked out.

Applications of BBa_K4268000

The part was received, resuspended, and cloned into the level 0 Golden Gate vector, PSB1C00. The ligation was then transformed into DH5&alpha cells. After overnight growth, white colonies (indicative of a replacement of the RFP reporter cassette) were subjected to screening for the correct insert using colony PCR with the VF/VR primer pair.


Insert gel figure

Caption: Colony PCR of five colonies (clones) suspected of coating the insert Head-tail connector insert. The insert length is 1573bp and the predicted size of the PCR product using VF/VR primers is 1739 bp.

The gel indicates that all five colonies are likely to contain the correct insert and thus, the Head-Tail Connector was successfully cloned into a Level O Golden Gate Assembly basic part.

User Reviews

UNIQb0d022fcb1a99670-partinfo-00000000-QINU UNIQb0d022fcb1a99670-partinfo-00000001-QINU