Difference between revisions of "Part:BBa K2353001:Experience"

 
 
(6 intermediate revisions by 2 users not shown)
Line 5: Line 5:
  
 
===Applications of BBa_K2353001===
 
===Applications of BBa_K2353001===
 +
When fully assembled, p-lambda-r LacI transcribes tsPurple and LacI represses pLac, preventing transcription of ClpXP-CI. Upon induction with IPTG, LacI binds to IPTG which prevents repression of pLac. Therefore, ClpXP-CI is transcribed; ClpXP should recognize the DAS deg tag and degrade the tsPurple protein. The ensuing levels of expression could then be compared to that of BBa_K2353000 (tsPurple without a deg tag) to see the relative level of degradation experienced.
  
 
===User Reviews===
 
===User Reviews===
 
<!-- DON'T DELETE --><partinfo>BBa_K2353001 StartReviews</partinfo>
 
<!-- DON'T DELETE --><partinfo>BBa_K2353001 StartReviews</partinfo>
−
<!-- Template for a user review
+
 
 
{|width='80%' style='border:1px solid gray'
 
{|width='80%' style='border:1px solid gray'
−
|-
+
|-<partinfo>BBa_K2353001 Review 1</partinfo>
−
|width='10%'|
+
<I>jleveille</I>
−
<partinfo>BBa_K2353001 AddReview number</partinfo>
+
−
<I>Username</I>
+
 
|width='60%' valign='top'|
 
|width='60%' valign='top'|
−
Enter the review inofrmation here.
+
tsPurpleDAS was ordered and hydrated from IDT and then digested and ligated into 1C3 so that it could be sent for sequencing and then submitted. Running the digest on a gel showed tsPurpleDAS to be the correct length, however when sequenced, we discovered that the DAS degradation tag was not present. It is hypothesized that at some point during the cloning workflow, tsPurple was confused with tsPurpleDAS.
−
|};
+
<br>
−
<!-- End of the user review template -->
+
<html>
 +
<img src="https://static.igem.org/mediawiki/2017/5/58/Tspurples_gel.jpeg" width="400px">
 +
</html>
 +
<br>
 +
<i style="font-size:12px;"> Gel results from a digest of all three tsPurple parts. From top to bottom: BBa_K2353000, BBa_K2353002, BBa_K2353001 </i>
 +
<br>
 +
<html>
 +
<br>
 +
Using Eurofins for sequencing, we realized that our tsPurpleDAS sequence was missing DAS deg tag. The results are shown below as our tsPurpleDAS construct aligned with the tsPurpleDAS sequencing results.
 +
<img src="https://static.igem.org/mediawiki/2017/4/47/Tspurpledas.seqimage.png" width="400px">
 +
</html>
 +
|}
 
<!-- DON'T DELETE --><partinfo>BBa_K2353001 EndReviews</partinfo>
 
<!-- DON'T DELETE --><partinfo>BBa_K2353001 EndReviews</partinfo>

Latest revision as of 16:21, 1 November 2017


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_K2353001

When fully assembled, p-lambda-r LacI transcribes tsPurple and LacI represses pLac, preventing transcription of ClpXP-CI. Upon induction with IPTG, LacI binds to IPTG which prevents repression of pLac. Therefore, ClpXP-CI is transcribed; ClpXP should recognize the DAS deg tag and degrade the tsPurple protein. The ensuing levels of expression could then be compared to that of BBa_K2353000 (tsPurple without a deg tag) to see the relative level of degradation experienced.

User Reviews

UNIQ93e533f904f37342-partinfo-00000000-QINU

jleveille

tsPurpleDAS was ordered and hydrated from IDT and then digested and ligated into 1C3 so that it could be sent for sequencing and then submitted. Running the digest on a gel showed tsPurpleDAS to be the correct length, however when sequenced, we discovered that the DAS degradation tag was not present. It is hypothesized that at some point during the cloning workflow, tsPurple was confused with tsPurpleDAS.

Gel results from a digest of all three tsPurple parts. From top to bottom: BBa_K2353000, BBa_K2353002, BBa_K2353001

Using Eurofins for sequencing, we realized that our tsPurpleDAS sequence was missing DAS deg tag. The results are shown below as our tsPurpleDAS construct aligned with the tsPurpleDAS sequencing results.

UNIQ93e533f904f37342-partinfo-00000004-QINU