Magni:Protocols/tecan - Revision history

Lorenzo Pasotti: /* List of filters */

Lorenzo Pasotti — Thu, 28 Mar 2013 18:00:32 GMT

List of filters

←Older revision		Revision as of 18:00, 28 March 2013
Line 26:		Line 26:
	**540/25		**540/25
	**535/25		**535/25
-	**485/20 (we have three of this)	+	**485/20 - we have three of this
	**430/35		**430/35

Lorenzo Pasotti: /* List of filters */

Lorenzo Pasotti — Thu, 28 Mar 2013 18:00:14 GMT

List of filters

←Older revision		Revision as of 18:00, 28 March 2013
Line 26:		Line 26:
	**540/25		**540/25
	**535/25		**535/25
-	**485/20 X3	+	**485/20 (we have three of this)
	**430/35		**430/35

Lorenzo Pasotti: /* Infinite F200 readings */

Lorenzo Pasotti — Thu, 28 Mar 2013 17:59:11 GMT

Infinite F200 readings

←Older revision		Revision as of 17:59, 28 March 2013
Line 7:		Line 7:
	=== Static readings ===		=== Static readings ===
	* Grow bacteria under the desired conditions.		* Grow bacteria under the desired conditions.
-	* Transfer 200 ul of ~~bacteria~~ in a well of a 96-well microplate.	+	* Transfer 200 ul of culture in a well of a 96-well microplate.
	* Measure with or without lid.		* Measure with or without lid.
	* Always measure a "blank" that can be subtracted to the other raw measurements (blank for absorbance is sterile media; blank for absorbance is non-fluorescent culture grown to an OD600 comparable to the fluorescent cultures).		* Always measure a "blank" that can be subtracted to the other raw measurements (blank for absorbance is sterile media; blank for absorbance is non-fluorescent culture grown to an OD600 comparable to the fluorescent cultures).
Line 13:		Line 13:
	=== Dynamic readings ===		=== Dynamic readings ===
	* Grow bacteria under the desired conditions.		* Grow bacteria under the desired conditions.
-	* Transfer 200 ul of ~~bacteria~~ in a well of a 96-well microplate, excluding the 36 wells of the "frame".	+	* Transfer 200 ul of culture in a well of a 96-well microplate, excluding the 36 wells of the "frame".
	* Dispense deionized water in all the unused wells.		* Dispense deionized water in all the unused wells.
	* Add inducers to the cultures when required.		* Add inducers to the cultures when required.
-	* Start a kinetic cycle of 18 hours with the i-control software (Tecan): 37degC during all the experiment, linear shaking (15 s, 3 mm amplitude), 5 s wait time (also 10 s is ok), absorbance reading (default parameters), fluorescence readings (gain=50 for medium and high copy plasmids, gain=50 or 80 for low copy plasmids, gain=80 or 100 for single-copy integrants), sampling time=5 min. The specified fluorescence gain factors are only guidelines and can be modified according to promoter/RBS strength, strain, growth medium and cell density.	+	* Start a kinetic cycle of 18 hours with the i-control software (Tecan): 37degC during all the experiment, linear shaking (15 s, 3 mm amplitude), 5 s wait time (also 10 s is ok), absorbance reading (default parameters), fluorescence readings (gain=50 for medium and high copy plasmids, gain=50 or 80 for low copy plasmids, gain=80 or 100 for single-copy integrants), sampling time=5 min. The specified fluorescence gain factors are only guidelines and can be modified according to promoter/RBS strength, strain, growth medium and cell density. RFP filters are 535/620nm, GFP filters are 485/540nm.
	* Always include a "blank" for absorbance and fluorescence, as described before.		* Always include a "blank" for absorbance and fluorescence, as described before.
		+	* The microplate should be covered with lid for dynamic readings >5-6 hours.
		+
		+	=== List of filters ===
		+	* for fluorescence (nm/bandwidth):
		+	**620/20
		+	**560/20
		+	**540/25
		+	**535/25
		+	**485/20 X3
		+	**430/35
		+
		+	* for absorbance (nm):
		+	**600
		+	**340
		+
		+	*Commonly used filter pairs for fluorescent proteins (EX=excitation filter, EM=emission filter):
		+	**CFP EX:430 EM:485
		+	**GFP EX:485 EM:540
		+	**YFP EX:485 EM:560
		+	**RFP EX:535 EM:620

Lorenzo Pasotti: /* Dynamic readings */

Lorenzo Pasotti — Sat, 02 Feb 2013 17:29:19 GMT

Dynamic readings

←Older revision		Revision as of 17:29, 2 February 2013
Line 16:		Line 16:
	* Dispense deionized water in all the unused wells.		* Dispense deionized water in all the unused wells.
	* Add inducers to the cultures when required.		* Add inducers to the cultures when required.
-	* Start a kinetic cycle of 18 hours with the i-control software (Tecan): 37degC during all the experiment, linear shaking (15 s, 3 mm amplitude), 5 s wait time (also 10 s is ok), absorbance reading (default parameters), fluorescence readings (gain=50 for medium and high copy plasmids, gain=80 for low copy plasmids, gain=80 or 100 for single-copy integrants), sampling time=5 min. The specified fluorescence gain factors are only guidelines and can be modified according to promoter/RBS strength, strain and cell density.	+	* Start a kinetic cycle of 18 hours with the i-control software (Tecan): 37degC during all the experiment, linear shaking (15 s, 3 mm amplitude), 5 s wait time (also 10 s is ok), absorbance reading (default parameters), fluorescence readings (gain=50 for medium and high copy plasmids, gain=50 or 80 for low copy plasmids, gain=80 or 100 for single-copy integrants), sampling time=5 min. The specified fluorescence gain factors are only guidelines and can be modified according to promoter/RBS strength, strain, growth medium and cell density.
	* Always include a "blank" for absorbance and fluorescence, as described before.		* Always include a "blank" for absorbance and fluorescence, as described before.

Lorenzo Pasotti: New page: {{Magni}}
= Infinite F200 readings = The microplate reader can be used for static or dynamic readings of bacteria cul...

Lorenzo Pasotti — Sat, 22 Sep 2012 18:29:13 GMT

New page: {{Magni}} <div style="padding: 10px; width: 720px; border: 5px solid #000000;"> = Infinite F200 readings = The microplate reader can be used for static or dynamic readings of bacteria cul...

New page

{{Magni}}

<div style="padding: 10px; width: 720px; border: 5px solid #000000;">
= Infinite F200 readings =
The microplate reader can be used for static or dynamic readings of bacteria cultures growth and fluorescent proteins.

=== Static readings ===
* Grow bacteria under the desired conditions.
* Transfer 200 ul of bacteria in a well of a 96-well microplate.
* Measure with or without lid.
* Always measure a "blank" that can be subtracted to the other raw measurements (blank for absorbance is sterile media; blank for absorbance is non-fluorescent culture grown to an OD600 comparable to the fluorescent cultures).

=== Dynamic readings ===
* Grow bacteria under the desired conditions.
* Transfer 200 ul of bacteria in a well of a 96-well microplate, excluding the 36 wells of the "frame".
* Dispense deionized water in all the unused wells.
* Add inducers to the cultures when required.
* Start a kinetic cycle of 18 hours with the i-control software (Tecan): 37degC during all the experiment, linear shaking (15 s, 3 mm amplitude), 5 s wait time (also 10 s is ok), absorbance reading (default parameters), fluorescence readings (gain=50 for medium and high copy plasmids, gain=80 for low copy plasmids, gain=80 or 100 for single-copy integrants), sampling time=5 min. The specified fluorescence gain factors are only guidelines and can be modified according to promoter/RBS strength, strain and cell density.
* Always include a "blank" for absorbance and fluorescence, as described before.

Magni:Protocols/tecan - Revision history

Lorenzo Pasotti: /* List of filters */

Lorenzo Pasotti: /* List of filters */

Lorenzo Pasotti: /* Infinite F200 readings */

Lorenzo Pasotti: /* Dynamic readings */

Lorenzo Pasotti: New page: {{Magni}} = Infinite F200 readings = The microplate reader can be used for static or dynamic readings of bacteria cul...

Lorenzo Pasotti: New page: {{Magni}}
= Infinite F200 readings = The microplate reader can be used for static or dynamic readings of bacteria cul...