Tag: spread plate

Another frustrating day in the lab Grew pNC0412B…

Deborah / / Uncategorized

Another frustrating day in the lab!!
Grew pNC0412B to an OD of 1 then performed a serial dilution (A,B,C,D -1to-5.5), I innoculated a fresh 10ml LB broth with 100 microlitres from dilution -4 and placed into the 37C shaker overnight. I then spread plated 100 microlitres from dilution -5 onto a fresh LB agar plate, I also spread plated 100 microlitres from dilution -5.5 onto a fresh LB agar plate, then placed these into the 37C incubator overnight. I would have also done this for pOG04 NJC but it refused to grow, after 6 hours in the 37C shaker only an OD of 0.01 was achieved, i abandoned this set of broths. The plates i hoped to grow for today containing the plasmids pALA1029, pOG4003 and pOG04 DRW also showed no growth. I have tried again today so hopefully tomorrow will be a better day. I seriously think C2110 is the work of the devil!

Hi all counted yesterdays plates and replica plated…

Deborah / / Uncategorized

Hi all,
counted yesterdays plates and replica plated pOG4 onto LB and LB containing amp100, then i counted and replica plated pnc0412B onto LB and LB containing chl20, then i incubated them overnight at 37C. My next task was to complete the second serial dilution and spread plated 10-5.5 and 10-6 onto LB agar plates with no antibiotics. Next i checked on pOG04NJC and pnc0412B – NO GROWTH!!! I am sure they know its monday! Having Checked with Ross williams he thinks it could be the plasmid at fault. Tomorrow i try again.

Today has been an almost perfect day in…

Deborah / / Uncategorized

Today has been an almost perfect day in the lab.
pOG4 and pNC0412B have grown quite nicely to an OD of 1 (0.8-1), I have performed a serial dilution on both and left the innoculated broths overnight in the shaker at 37C, the spread plates of 5 & 5.5 are in the 37C incubator. Also today i innoculated another flask of LB broth with C2110 and grew to an OD of 0.35. Then i carried out cell competancy then transformed them, using the plasmids pOG04 NJC and pNC0412B. I spread plated pOG04NJC onto an LB agar plate containing amp50 and spread plated pNC0412B onto an LB agar plated containing chl10. these were then placed in the 37C incubator overnight. Busy day tomorrow as agars, broths, velvets and NaCl need to be made-up/autoclaved, as well as tomorrows tasks. Early start i think!

Hey all Yesterday i took the overnight broths…

Deborah / / Uncategorized

Hey all,
Yesterday i took the overnight broths for the two plasmids and completed another dilution series and spread plated the ones from -5.5 and -6. I incubated these at 37C overnight. Then i took out the T0 plates and counted the colonies, the ones that were around 100 i replica plated them onto an LB amp100 agar plate and then olnto an LB agar plate. These were then incubated overnight at 37C.
Today started out with me making more LB agar and LBamp100 agar plates, and making up 3x50ml LB broth, ready for monday morning. Then i autoclaved my velvets ready for next week. Once this was completed the science could start. I counted Tend plates and replica plated the ones with around 100 colonies, same as yesterday i replica plated 1x LBamp100 and 1xLB agar. The last job of the day was to count the colonies from yesterdays replica plating and record them. I also re streaked my C2110 so that i will be working with a fresh culture on monday.