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Pro Tips

1. Run at the larger scale
2. ALWAYS kill it with PRoteinase K
3. Transform 4 uL of the reaction
4. Use ALL the transformation tricks
5. Plate ALL of it
6. ALWAYS run a PUC19 control for transformations

Instructions for newbies:

1.5 uL of 10 fmol/uL Dest

1.5 uL of 5 fmol/uL promoter

1.5 uL of 5 fmol/uL gene

1.5 uL of H₂O

1.5 uL of LR clonase

Pipette up and down.  Incubate at room temperature overnight.

Next day:  Add 1.5 uL of proteinase K.  Incubate for 15 minutes at 37C, then transform.

Instructions:

USE 3x VOLUME OF EVERYTHING (at least for now) 

Use nanodrop to measure concentration of pEntry vectors, make 5 femtomolar working solution of each pEntry.

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Combine into 1 aliquot:

• 1uL of 5 fmol/uL of pENTR_L4_Promoter_R1
• 1uL of 5 fmol/uL pENTR_L1_Gene_L2
• 1uL of 10 fmol/uL pDEST_R4_R2

• 3uL Total

WARNING: KEEP ALIQUOTED LR CLONASE MIX AT -80 AT ALL TIMES!

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