User:Karmella Haynes/Notebook/BioBrick cloning/2010/01/16

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01/16/10

 * &#x2713; Minipreps: KAH117, 118/MV2 (re-transformation clones, 2 each)
 * &#x2713; Cultures: KAH137(3), 138(3)
 * &#x2713; Assemblies: KAH94, 95/ JDS47; KAH94, 95/ JDS48 (re-do)

Minipreps > Check with E/P digests

--> KAH117/MV2 worked. Pick 4 more colonies from KAH118/MV2 re-transformation plate

Assemblies --> Re-do (see 1/14), use higher conc. of insert --> Increase ligation time (1 hr.)
 * 1) KAH94/JDS47: (1) KAH94/(N, blunted)/5011 + JDS47/(BglII, blunted, dp)/7509 &#x2713;
 * 2) KAH95/JDS47: (2) KAH94/(N, blunted)/5083  + "
 * 3) KAH94/JDS48: KAH94/(N, blunted)/5011 + JDS48/(BglII, blunted, dp)/7519 &#x2713;
 * 4) KAH95/JDS48: KAH94/(N, blunted)/5083  + "

> Digests (Fermentas FD)

> Measure conc.'s of inserts

> Blunting (NEB Quick Blunting kit) --> Blunt all DNA (vectors and inserts)


 * 1) KAH94 (max. vol.) [final] = 13.7 ng/μL
 * 2) KAH95 (max. vol.) [final] = 13.7 ng/μL

> Ligations


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