Team:UNICAMP-Brazil/Notebooks/September 18

ColiGuard

PY Promoter - Mini-prep and PCR confirmation

• Today we performed mini-preps (Protocol 2) to extract the plasmids from the selected colonies inoculated yesterday.

• After the extraction we performed PCR reactions with 3 pairs of primers to confirm the ligation:

- Ppy-F and Ppy-R

- Ppy-F and VR

- VF and VR

• Unfortunately, the sizes of the bands in the agarose gel were not compatible with the expected size of the bands. =(

Fabi and Léo

CeaB, CeiB and BBa_B0015 restriction, purification and quantification

• We made the DNA purification in order to clean all buffer PCR salts. Immediately, we cut CeaB and CeiB purified with SpeI and XbaI restriction enzymes. We made that, because we want to join the CeaB DNA with a terminator. The terminator chosen was BBa_B0015. Separately, we cut the terminator plasmid with XbaI enzyme restriction and we made DNA quantification. Then, we made DNA purification with kit Pure-link PCR.

Luige

YeastGuard

New biobricks - screening

• We tried to find the colonies containing the biobricks by colony PCR (using the forward primer of the insert and the reverse of the plasmid). We did 10 reactions of each biobrick. Unfortunately we didn´t find the correct bands.

GEL

Taís

• We decided to make a bigger screening of our colonies to find the ones containing the biobricks (pJen1, pDLD, Lysozyme, JEN1 orf). We grew 24 colonies from each biobrick O.N. in liquid medium.

Raíssa