User:Karmella Haynes/Notebook/BioBrick cloning/2010/04/12

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04/12/10

 * &#x2713; Minipreps: KAH146-153
 * &#x2713; Assemblies: KAH96, 142, 146, 147/ V0201; KAH154-159

Minipreps > Check with E/P digests

--> 37°C/ ~15 min.

Assemblies
 * 1) KAH96/V0201: (1) KAH96/(N/S)/2825 + V0201/(N/S dp)/5157 &#x2713;
 * 2) KAH142/V0201: (2) KAH142/(N/S)/4571 + "
 * 3) KAH146/V0201: (3) KAH146/(N/S)/6386 + "
 * 4) KAH147/V0201: (4) KAH147/(N/S)/8132 + "
 * 5) KAH154: (5) KAH148/(X/P)/967 + KAH3/(S/P)/~3951 &#x2713;
 * 6) KAH155: (6) KAH149/(X/P)/967 + "
 * 7) KAH156: (7) KAH150/(X/P)/967 + "
 * 8) KAH157: (8) KAH151/(X/P)/967 + "
 * 9) KAH158: (9) KAH152/(X/P)/1931 + "
 * 10) KAH159: (10) KAH153/(X/P)/1934 + "

> Digests (Fermentas FD) --> Triple digest KAH96 w/ N/S/ApaLI --> Do all digests for at least 1 hour to ensure complete digestion

> Measure conc.'s

--> Already cut:
 * 1) V0201 (N/S dp) = 25 ng/μL
 * 2) KAH3 (S/P) = 27.9 ng/μL

> Ligations

--> Ligations 1-5: R.T./~1 hr.; Add to 25 μL Z-DH5α --> Ligations 6-12: R.T./~10 min.; Add to 25 μL Z-DH5α


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