Antibiotic Usage

How to make LB liquid plus antibiotics: o Ampicillin The frozen stock solutions of ampicillin are at 50mg/ml and 100mg/ml in H2O, and are marked with a red sticker. The final concentration for LB liquid culture is 50ul/ml. To obtain this in 100ml (the amount in each LB bottle), add 100ul stock solution. o Kanamycin The frozen stock solutions of kanamycin are at 50mg/ml in H2O, and are marked with green. The final concentration for LB liquid culture for growing plasmids is 50ug/ml, and for cosmids is 20ug/ml. To obtain 50ng/ml in 100ml of LB, add 100ul stock solution, and to obtain 20ug/ml, add 40ul stock solution. o Tetracycline The frozen stock solutions of tetracycline are at 15mg/ml in methanol and are marked with black. The final concentration for LB liquid culture is 15ug/ml. To obtain this in 100ml of LB, add 100ul stock solution. o Chloramphenicol The frozen stock solutions of chloramphenicol are at 25mg/ml in 100% ethanol and are marked with purple. The final concentration for LB liquid culture is 25mg/ml. To obtain this in 100ml of LB, add 100ul stock solution. How to make LB plates plus antibiotics: o Follow the recipe card in box for making LB plates, being sure to add the agar. After autoclaving, and when the agar has cooled enough that its not too hot to touch (about 1 to 1.5hrs), add antibiotics as follows: Ampicillin add 1ml ampicillin (at 100mg/ml) per liter of agar to obtain a final concentration of 100ug/ml. Mark the plate with a single red line on the side. Kanamycin add 1ml kanamycin stock (at 50mg/ml) per liter of agar to obtain a final concentration of 50ug/ml. Mark the plates with a single green line on the side. Tetracycline add 1ml tetracycline stock (at 15mg/ml) per liter of agar to obtain a final concentration of 15ug/ml. Mark the plates with a single black line on the side. Chloramphenicol add 1ml chloramphenicol stock (at 25mg/ml) per liter of agar to obtain a final concentration of 100ug/ml. Mark the plates with a single purple line on the side. How to determine volumes to use to obtain a certain concentration: o Use the following formula to determine amount of stock solution to use for any amount. Make sure units match! C1V1 = C2V2 C = concentration; V = volume Example: You have a stock solution of A at 100mg/ml. You want 150 ml at 100ug/ml. Use the C1V1 for what you want, and the C2V2 for what you have. Just plug in the numbers: (100ug/ml)(150ml) = (100000ug/ml)(x) 15000 = 100000x x = 0.15ml, or 150ul Remember that you have to convert mg/ml to ug/ml, which you can do by multiplying by 1000.

Uso de antibiticos
Cmo hacer LB lquido ms antibiticos:

Ampicilina - Las soluciones madre congeladas de ampicilina estn a 50mg/ml y 100mg/ml en H 2 O y se marcan con una etiqueta roja. La concentracin final de cultivo lquido LB es 50ul/ml. Para obtener este en 100 ml (la cantidad en cada botella LB), aadir solucin de 100 ul.

Kanamicina - Las soluciones madre congeladas de kanamicina estn en 50mg/ml en H 2 O y se marcan con verde. La concentracin final de cultivo lquido LB de plsmidos que crecen es 50ug/ml, y csmidos es 20ug/ml. Para obtener 50ng/ml en 100 ml de LB, aadir solucin de 100 ul y obtener 20ug/ml, aadir solucin madre 40ul.

Tetraciclina - Las soluciones madre congeladas de tetraciclina son a 15mg/ml en metanol y se marcan con el negro. La concentracin final de cultivo lquido LB es 15ug/ml. Para obtener este en 100 ml de LB, aadir solucin de 100 ul.

El cloranfenicol - Las soluciones madre congeladas de cloranfenicol estn en 25mg/ml en etanol al 100% y se marcan con color prpura. La concentracin final de cultivo lquido LB es 25mg/ml. Para obtener este en 100 ml de LB, aadir solucin de 100 ul.

Cmo hacer placas de LB ms antibiticos:

Siga la tarjeta de la receta en la caja para hacer placas de LB, asegurndose de aadir el agar. Despus de autoclave, y cuando el agar se ha enfriado lo suficiente para que no est demasiado caliente al tacto (aproximadamente 1 a 1,5 horas), aadir los antibiticos de la siguiente manera:

Ampicilina - Aadir 1 ml de ampicilina (a 100mg/ml) por litro de agar para obtener una concentracin final de 100ug/ml. Marque la placa con una sola lnea roja en el lateral.

Kanamicina - aadir 1 ml de stock kanamicina (a 50mg/ml) por litro de agar para obtener una concentracin final de 50ug/ml. Marque las placas con una sola lnea verde en el lado.

Tetraciclina - aadir 1 ml de stock de tetraciclina (a 15mg/ml) por litro de agar para obtener una concentracin final de 15ug/ml. Marque las placas con una sola lnea de negro en el lateral.

Cloranfenicol - aadir 1 ml de stock cloranfenicol (a 25mg/ml) por litro de agar para obtener una concentracin final de 100ug/ml. Marque las placas con una sola lnea de color morado en el lateral.

Cmo determinar volmenes a utilizar para obtener una cierta concentracin:

Utilice la siguiente frmula para determinar la cantidad de solucin madre a utilizar para cualquier cantidad. Asegurarse que las unidades partido!

C1V1=C2V2 C = concentracin; V = volumen Ejemplo: Usted tiene una solucin madre de A a 100mg/ml. Quieres 150 ml a 100ug/ml. Utilice el C 1 V 1 por lo que quieres, y el C 2 V 2 por lo que tienes. Slo tiene que conectar en los telfonos:

(100ug/ml) (150 ml) = (100000ug/ml) (x) 15000 = 100000x x = 0,15 ml, o 150ul Recuerde que usted tiene que convertir mg / ml de ug / ml, que se puede hacer multiplicando por 1000.