Difference between revisions of "Team:UCL/Notebook"
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− | <div class="week" data-week=" | + | <div class="week" data-week="9" id="week9"> |
− | <h2>Week | + | <h2>Week 9(10<sup>th</sup> Aug – 16<sup>th</sup> Aug)</h2> |
<div class="day"> | <div class="day"> | ||
<h3>Monday 14<sup>th</sup></h3> | <h3>Monday 14<sup>th</sup></h3> | ||
− | + | <h4>Mind the Gut: Effectors</h4> | |
+ | <p>Repeated the digestion of CHAT and TPH1 and ran the gel for digested and undigested CHAT and TPH1. The DNA fingerprint showed only CHAT had been digested. </p> | ||
+ | <p> PCR cleanup of the amplified antiTNA 1 and 2 from last Tuesday was carried out. Nano drop showed nucleic acid concentrations from 20-35 ng/ul.</p> | ||
+ | <Purified antiTNA 1 and 2 along with pSBIC3 backbone were digested using restriction enzymes XbaI and Pst.1:5 and 1:3 molar ratio ligation of backbone to antiTNA was done and transformed to DH5α competent cells.</p> | ||
+ | |||
+ | <p>In the afternoon, we repeated the digestion of CHAT and TPH1 again but with single digestion. We ran the gel with linear pSB1C3, CHAT and TPH1. It confirmed ligation of CHAT however, linear plasmids for TPH1 were shorter than pSB1C3, hence unsuccessful for TPH1.</p> | ||
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<div class="day"> | <div class="day"> | ||
<h3>Tuesday 15<sup>th</sup></h3> | <h3>Tuesday 15<sup>th</sup></h3> | ||
− | + | <h4>Mind the Gut: Effectors</h4> | |
+ | <p> Started the day with mini preps of GADX4, KATX4,FldHC-TPH1X4,Pyear-TPH1X4 using thermoscientific kit. Nanodrop showed better concentrations of nucleic acids for FldHC-TPH1X4,Pyear-TPH1X4 ranging from 78.4-180 ng/ul whereas KAT and GAD only had 20-30 ng/ul.</p> | ||
+ | <p>We ran the digestion of effectors namely KAT,GAD and FldHC-TPH1/Pyear-TPH1. Promoter Ptac in the backbone PSBIC3 plasmid was also digested.</p> | ||
+ | <p> The double digestion when ran on gel yielded inconsistent results and needed repetition.We also thought best to do single digestion with this repeat.</p> | ||
+ | <p>We had also been looking for alternative restriction sites in CHAT and found couple of them. We used PstI and BglI. The gel confirmed RBS-CHAT biobrick.</p> | ||
</div> | </div> | ||
Revision as of 21:58, 27 August 2015