IGEM:Harvard/2009/Notebook/Harvard iGEM 2010/2010/07/21: Difference between revisions
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==Team Allergy== | ==Team Allergy== | ||
Yesterday, we plated transformants for the allergen panel and introns. Today, we will purify the plasmids from the transformants. | |||
===Procedures=== | |||
#Culture Colonies | |||
#Miniprep of colonies | |||
#Diagnostic Digest to check for inserts | |||
===Results=== | |||
'''Concentrations''' of Allergen Parts and Introns: | '''Concentrations''' of Allergen Parts and Introns: | ||
*Allergen Parts: | *Allergen Parts: | ||
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[[Image:AllergenPanel2DiagHC.jpg|240px]] | [[Image:AllergenPanel2DiagHC.jpg|240px]] | ||
==Team Fence== | ==Team Fence== |
Revision as of 22:17, 25 July 2010
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Team AllergyYesterday, we plated transformants for the allergen panel and introns. Today, we will purify the plasmids from the transformants. Procedures
ResultsConcentrations of Allergen Parts and Introns:
Diagnostic Digest lader; ladder; GerS (1-3); Ger A (1-3); Bet2S (1-3); Bet2A (1-3); BetA (!-3); LTPS (1-3); LTPA (1-3); Pal (1-3); ladder; ladder (More contrasted image below): Team FenceNanodroping 5xGal promoterNanodrop of ligated annealed oligos: 5200.8 ng/μL, 1.32 260/280 E-gel of 5xGal promoterRan 2% E-gel of .5μL of the ligation reaction (2.6μg) in 15μL water, with 10μL of KB plus ladder on either side. Ladder ran screwy, so new ladder used,and a 2% agarose Gel was poured for gel extraction Gel Extraction of 5xGal promoterpoured 2% agarose Gel for gel extraction, ran 2μL of the ligated annealing reaction in 13μL EB buffer, with two KB plus 1 ladders, one on each side.
Nanodrop for cleaned-up gel purified 5xGalpt negative - ran a 2% E-gel to check. There appears to be something at the .16kb size. Team FlavorMiraculin N/Brazzein C StrepII & Stop
Gel Lanes: 1. 1kb plus ladder 2. Mira N1 XbaI/PstI 3. Mira N2 XbaI/PstI 4. Brazz C1 XbaI/PstI 5. Brazz C2 XbaI/PstI 6. 1kb plus ladder 7. Mira N1 undigested 8. Mira N2 undigested 9. Brazz C1 undigested 10. Brazz C2 undigested
ODs Mira N1: 5.4 ng/μL Mira N2: 3.1 ng/μL Brazz C1: 6.1 ng/μL Brazz C2: 10.1 ng/μL
Ligation/Transformation of Mira/Brazz+StrepII
NEB T4 Ligase was used for all reactions Digestion & Gel Purification of V24
1039.9 ng of DNA was digested in both reactions Digestion reactions were run on a 1% Agarose gel at 125V for 30 min OD's V24-1 N/S: 13.5 ng/μL V24-2 N/S: 17.9 ng/μL
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