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Team:Technion/14 August 2012
From 2012.igem.org
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==Ilya== | ==Ilya== | ||
- Gradient PCR of I0462 from F2620 failed. I need to design a different primer.<br> | - Gradient PCR of I0462 from F2620 failed. I need to design a different primer.<br> | ||
| + | - Glycerol stock of pSB3C5. Freezer position is 4C.<br> | ||
- Minipreped the pSB3C5 and R0062+Fus1. Got very high concentrations of pSB3C5 in comparison to pSB1A2. This is not logical because pSB1A2 is a high copy plasmid and pSB3C5 is a low-medium copy plasmid. I wanted to change the backbone to a low copy plasmid Roee has in his lab, but Orna told me to continue with this backbone. I bet we will get bad results in the plate reader eventually again.<br> | - Minipreped the pSB3C5 and R0062+Fus1. Got very high concentrations of pSB3C5 in comparison to pSB1A2. This is not logical because pSB1A2 is a high copy plasmid and pSB3C5 is a low-medium copy plasmid. I wanted to change the backbone to a low copy plasmid Roee has in his lab, but Orna told me to continue with this backbone. I bet we will get bad results in the plate reader eventually again.<br> | ||
| - | - Set restriction digest with XbaI and PstI of: pSB1C3+Fus2 (to get both, backbone and insert), R0062+Fus1 in pSB1A2 (to get the insert) and pSB3C5 (to get the backbone). | + | - Set restriction digest with XbaI and PstI of: pSB1C3+Fus2 (to get both, backbone and insert), R0062+Fus1 in pSB1A2 (to get the insert) and pSB3C5 (to get the backbone). After I got home, I realized that I should have digested with EcoRI and PstI instead of XbaI and PstI. |
==Inbal== | ==Inbal== | ||
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==Evgeni== | ==Evgeni== | ||
| + | I had done digestion and purification of pPROLar again, in final step I had used mbH20 and not elution buffer provided with the kit. Also I had used a smaller volume of elution liquid in final step of purification - 25 ul and not 50 ul, like previous day. After purification I had got a slightly better concentration of the digested plasmid, about 13 ng/ul. I'll continue to work with it. | ||
==Shahar== | ==Shahar== | ||
Latest revision as of 14:21, 19 August 2012
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Ilya
- Gradient PCR of I0462 from F2620 failed. I need to design a different primer.
- Glycerol stock of pSB3C5. Freezer position is 4C.
- Minipreped the pSB3C5 and R0062+Fus1. Got very high concentrations of pSB3C5 in comparison to pSB1A2. This is not logical because pSB1A2 is a high copy plasmid and pSB3C5 is a low-medium copy plasmid. I wanted to change the backbone to a low copy plasmid Roee has in his lab, but Orna told me to continue with this backbone. I bet we will get bad results in the plate reader eventually again.
- Set restriction digest with XbaI and PstI of: pSB1C3+Fus2 (to get both, backbone and insert), R0062+Fus1 in pSB1A2 (to get the insert) and pSB3C5 (to get the backbone). After I got home, I realized that I should have digested with EcoRI and PstI instead of XbaI and PstI.
Inbal
Asaf
Hila
Lior
Noa
Cut xyIE insert with Xmal and XhoI ??
Evgeni
I had done digestion and purification of pPROLar again, in final step I had used mbH20 and not elution buffer provided with the kit. Also I had used a smaller volume of elution liquid in final step of purification - 25 ul and not 50 ul, like previous day. After purification I had got a slightly better concentration of the digested plasmid, about 13 ng/ul. I'll continue to work with it.
