IGEM:Peking/2007/Count-Conjugation-Notebook/2007-8-5: Difference between revisions
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Qu Mingzhi (talk | contribs) (New page: ==Bacteria Mating== * By Yang Yifan * Donnor: E.coli strain C600 with plasmid R751 and pSC101, use cells with only R751 as negative control; <br> Recipient: E.coli strain DH5α with plasmi...) |
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* 500ul culture centrifuge and resuspension with LB for two times to wash the antibiotics. | * 500ul culture centrifuge and resuspension with LB for two times to wash the antibiotics. | ||
* Mix, and culture under 37°C for 90 min. Culture two more stationary-stationary mixture for 60min and 120min as a time control. | * Mix, and culture under 37°C for 90 min. Culture two more stationary-stationary mixture for 60min and 120min as a time control. | ||
==Amplification Culture of F-OriT_pSB1A2 <- J23066== | |||
*by Qu Mingzhi | |||
*Culture at Amp+ plate for 12 hours. | |||
*select Positive F-OriT_J23066_pSB1A2 Colonies from Plate, Culture in liquid LB,waiting for mini-prep. | |||
*F-OriT_pSB1A2 self-ligation nagetive-control received 10 clones. | |||
Revision as of 20:18, 5 August 2007
Bacteria Mating
- By Yang Yifan
- Donnor: E.coli strain C600 with plasmid R751 and pSC101, use cells with only R751 as negative control;
Recipient: E.coli strain DH5α with plasmid pSB1A2 confering ampicilin resistance.
- Recipient are grown to stationary phase, Donnor are grown to log phase and stationary phase in two independent experiments.
- OD600: recipient - 1.118AU; donnor(stationary) - 1.308AU; donnor(log-phase) - 0.524AU; donnor(control) - AU.
- 500ul culture centrifuge and resuspension with LB for two times to wash the antibiotics.
- Mix, and culture under 37°C for 90 min. Culture two more stationary-stationary mixture for 60min and 120min as a time control.
Amplification Culture of F-OriT_pSB1A2 <- J23066
- by Qu Mingzhi
- Culture at Amp+ plate for 12 hours.
- select Positive F-OriT_J23066_pSB1A2 Colonies from Plate, Culture in liquid LB,waiting for mini-prep.
- F-OriT_pSB1A2 self-ligation nagetive-control received 10 clones.
