Cell cycle analysis - Revision history

Torsten Waldminghaus: /* Determination of initiation age (ai) and C+D: */

Torsten Waldminghaus — Wed, 04 May 2011 18:20:03 GMT

Determination of initiation age (ai) and C+D:

←Older revision		Revision as of 18:20, 4 May 2011
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	This gives:		This gives:

-	'''<math>a_i=\tau-\frac{log(2-F)}{log2}*\tau</math>'''	+	'''<math>a_i=\tau-\frac{log(2-F)}{log2}*\tau</math>'''
		+
		+	which is the same as this (log2 is 1):
		+
		+	'''<math>a_i=\tau-log(2-F)*\tau</math>'''

	If you have for example a generation time τ=84 minutes and the portion of cells with 4 origins is 66% the formula gives:		If you have for example a generation time τ=84 minutes and the portion of cells with 4 origins is 66% the formula gives:


-	'''<math>a_i=84-~~\frac{~~log(2-0.66)~~}{log2}~~*84=48.5</math>'''	+	'''<math>a_i=84-log(2-0.66)*84=48.5</math>'''


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	[[Image:C+D_1.jpg\|left\|250px]]		[[Image:C+D_1.jpg\|left\|250px]]
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	===Determination of the C and D periods:===		===Determination of the C and D periods:===

Torsten Waldminghaus at 15:21, 4 May 2011

Torsten Waldminghaus — Wed, 04 May 2011 15:21:59 GMT

←Older revision		Revision as of 15:21, 4 May 2011
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	'''<math>\frac{oriC}{terC}=2^{\frac{C}{\tau}}</math>'''		'''<math>\frac{oriC}{terC}=2^{\frac{C}{\tau}}</math>'''
		+

	which gives:		which gives:

-	'''<math>\frac{oriC}{terC}~~=2^{\frac{C}~~{\tau}}</math>'''	+
		+	'''<math>C=log(\frac{oriC}{terC})*{\tau}</math>'''

Torsten Waldminghaus: /* Determination of the C and D periods: */

Torsten Waldminghaus — Wed, 04 May 2011 15:18:37 GMT

Determination of the C and D periods:

←Older revision		Revision as of 15:18, 4 May 2011
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	The C period is found from the ''oriC/terC'' ratio obtained by Southern blot or qPCR analysis ([[oriC/ter ratio determination]]) and the generation time (τ):		The C period is found from the ''oriC/terC'' ratio obtained by Southern blot or qPCR analysis ([[oriC/ter ratio determination]]) and the generation time (τ):
		+
		+	'''<math>\frac{oriC}{terC}=2^{\frac{C}{\tau}}</math>'''
		+
		+	which gives:

	'''<math>\frac{oriC}{terC}=2^{\frac{C}{\tau}}</math>'''		'''<math>\frac{oriC}{terC}=2^{\frac{C}{\tau}}</math>'''
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	[[Image:C+D_2.jpg\|left\|250px]]		[[Image:C+D_2.jpg\|left\|250px]]
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	===The theoretical exponential DNA histogram:===		===The theoretical exponential DNA histogram:===

Torsten Waldminghaus at 16:23, 26 February 2009

Torsten Waldminghaus — Thu, 26 Feb 2009 16:23:04 GMT

←Older revision		Revision as of 16:23, 26 February 2009
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	===Determination of the C and D periods:===		===Determination of the C and D periods:===

-	The C period is found from the ''oriC/terC'' ratio obtained by Southern blot or qPCR analysis and the generation time (τ):	+	The C period is found from the ''oriC/terC'' ratio obtained by Southern blot or qPCR analysis ([[oriC/ter ratio determination]]) and the generation time (τ):

	'''<math>\frac{oriC}{terC}=2^{\frac{C}{\tau}}</math>'''		'''<math>\frac{oriC}{terC}=2^{\frac{C}{\tau}}</math>'''
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	The average number of replication forks: (6 x 0.10) + (4 x 0.13) + (12 x 0.77) = 10.4 forks		The average number of replication forks: (6 x 0.10) + (4 x 0.13) + (12 x 0.77) = 10.4 forks
		+
		+
		+	[[Category:Protocol]][[Category:Escherichia coli]]

Torsten Waldminghaus at 20:43, 21 January 2009

Torsten Waldminghaus — Wed, 21 Jan 2009 20:43:41 GMT

←Older revision		Revision as of 20:43, 21 January 2009
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	[[Image:C+D_1.jpg\|left\|250px]]		[[Image:C+D_1.jpg\|left\|250px]]
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	===Determination of the C and D periods:===		===Determination of the C and D periods:===
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	[[Image:C+D_2.jpg\|left\|250px]]		[[Image:C+D_2.jpg\|left\|250px]]
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	===The theoretical exponential DNA histogram:===		===The theoretical exponential DNA histogram:===
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	[[Image:C+D_3.jpg\|left\|350px]]		[[Image:C+D_3.jpg\|left\|350px]]
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Torsten Waldminghaus: /* Determination of the C and D periods: */

Torsten Waldminghaus — Wed, 21 Jan 2009 20:40:30 GMT

Determination of the C and D periods:

←Older revision		Revision as of 20:40, 21 January 2009
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	===Determination of the C and D periods:===		===Determination of the C and D periods:===

-	The C period is found from the ''oriC/terC'' ratio obtained by Southern blot analysis and the generation time (τ):	+	The C period is found from the ''oriC/terC'' ratio obtained by Southern blot or qPCR analysis and the generation time (τ):

-	'''''oriC/terC''=2~~<sup>~~C/τ</~~sup~~>'''	+	'''<math>\frac{oriC}{terC}=2^{\frac{C}{\tau}}</math>'''


	The D period is found from the C+D and C period:		The D period is found from the C+D and C period:

-	'''D = (C+D) - C'''	+	'''<math>D = (C+D) - C</math>'''


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	[[Image:C+D_2.jpg\|left\|250px]]		[[Image:C+D_2.jpg\|left\|250px]]
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	===The theoretical exponential DNA histogram:===		===The theoretical exponential DNA histogram:===

Torsten Waldminghaus: /* Determination of initiation age (ai) and C+D: */

Torsten Waldminghaus — Wed, 21 Jan 2009 13:25:10 GMT

Determination of initiation age (ai) and C+D:

←Older revision		Revision as of 13:25, 21 January 2009
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	From flow cytometry analysis of cells treated with rifampicin and cephalexin (run-out histogram) the proportions of cells that had not initiated replication at the time of drug action (4-origin-cells, streaked) and cells that had initiated (8-origin-cells) can be estimated.The initiation age (a<sub>i</sub>) can be found from the theoretical age distribution described by this formula,		From flow cytometry analysis of cells treated with rifampicin and cephalexin (run-out histogram) the proportions of cells that had not initiated replication at the time of drug action (4-origin-cells, streaked) and cells that had initiated (8-origin-cells) can be estimated.The initiation age (a<sub>i</sub>) can be found from the theoretical age distribution described by this formula,

-	'''<math>F=2-2^\frac{\tau-a_i~~</sub>~~)~~/τ)</sup>'''~~	+	'''<math>F=2-2^{\frac{(\tau-a_i)}{\tau}}</math>'''
-		+
-	~~This gives:~~	+
-		+
-	~~'''<math>a_i=~~\tau~~-\frac{log(2-F)~~}~~{log2~~}*\tau</math>'''	+


	where F is the fraction of cells that had not initiated and τ is the generation time, or from the estimated graph of the theoretical age distribution (streaked portion).		where F is the fraction of cells that had not initiated and τ is the generation time, or from the estimated graph of the theoretical age distribution (streaked portion).
-		+
-		+	This gives:

		+	'''<math>a_i=\tau-\frac{log(2-F)}{log2}*\tau</math>'''

		+	If you have for example a generation time τ=84 minutes and the portion of cells with 4 origins is 66% the formula gives:


		+	'''<math>a_i=84-\frac{log(2-0.66)}{log2}*84=48.5</math>'''


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	[[Image:C+D_1.jpg\|left\|250px]]		[[Image:C+D_1.jpg\|left\|250px]]
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	===Determination of the C and D periods:===		===Determination of the C and D periods:===

Torsten Waldminghaus at 13:13, 21 January 2009

Torsten Waldminghaus — Wed, 21 Jan 2009 13:13:51 GMT

←Older revision		Revision as of 13:13, 21 January 2009
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-	~~'''1.~~Determination of initiation age (a<sub>i</sub>) and C+D~~'''~~:[[Image:Theoretical_age_distr.jpg\|left\|200px]]	+	===Determination of initiation age (a<sub>i</sub>) and C+D:===
		+	[[Image:Theoretical_age_distr.jpg\|left\|200px]]
	From flow cytometry analysis of cells treated with rifampicin and cephalexin (run-out histogram) the proportions of cells that had not initiated replication at the time of drug action (4-origin-cells, streaked) and cells that had initiated (8-origin-cells) can be estimated.The initiation age (a<sub>i</sub>) can be found from the theoretical age distribution described by this formula,		From flow cytometry analysis of cells treated with rifampicin and cephalexin (run-out histogram) the proportions of cells that had not initiated replication at the time of drug action (4-origin-cells, streaked) and cells that had initiated (8-origin-cells) can be estimated.The initiation age (a<sub>i</sub>) can be found from the theoretical age distribution described by this formula,

-	'''F = 2 - 2~~<sup>((thau~~-~~a<sub>i~~</sub>)/τ)</sup>'''	+	'''<math>F=2-2^\frac{\tau-a_i</sub>)/τ)</sup>'''

	This gives:		This gives:
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-	~~'''2.~~ Determination of the C and D periods:~~'''~~	+	===Determination of the C and D periods:===

	The C period is found from the ''oriC/terC'' ratio obtained by Southern blot analysis and the generation time (τ):		The C period is found from the ''oriC/terC'' ratio obtained by Southern blot analysis and the generation time (τ):
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-	~~'''3.~~ The theoretical exponential DNA histogram:~~'''~~	+	===The theoretical exponential DNA histogram:===

	A theoretical exponential DNA histogram can be drawn to check whether the obtained values fit with the experimental data. From the C+D period the DNA content of the cells at different time points in the cell cycle can be calculated.		A theoretical exponential DNA histogram can be drawn to check whether the obtained values fit with the experimental data. From the C+D period the DNA content of the cells at different time points in the cell cycle can be calculated.
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-	~~'''4.~~ Calculation of the average number of replication forks when D=τ:~~'''~~	+	===Calculation of the average number of replication forks when D=τ:===

	In the example given above, 23% of the cells contain 4 replication forks (4-origin peak in run-out histogram) and 77% contain 12 replication forks (8-origin peak), hence the average number of replication forks in the cell population will be:		In the example given above, 23% of the cells contain 4 replication forks (4-origin peak in run-out histogram) and 77% contain 12 replication forks (8-origin peak), hence the average number of replication forks in the cell population will be:
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-	~~'''5.~~ Calculation of the average number of replication forks when D≠τ:~~'''~~	+	===Calculation of the average number of replication forks when D≠τ:===

	Example:		Example:

Torsten Waldminghaus at 13:06, 21 January 2009

Torsten Waldminghaus — Wed, 21 Jan 2009 13:06:51 GMT

←Older revision		Revision as of 13:06, 21 January 2009
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	From flow cytometry analysis of cells treated with rifampicin and cephalexin (run-out histogram) the proportions of cells that had not initiated replication at the time of drug action (4-origin-cells, streaked) and cells that had initiated (8-origin-cells) can be estimated.The initiation age (a<sub>i</sub>) can be found from the theoretical age distribution described by this formula,		From flow cytometry analysis of cells treated with rifampicin and cephalexin (run-out histogram) the proportions of cells that had not initiated replication at the time of drug action (4-origin-cells, streaked) and cells that had initiated (8-origin-cells) can be estimated.The initiation age (a<sub>i</sub>) can be found from the theoretical age distribution described by this formula,

-	'''F = 2 - 2<sup>((τ-a<sub>i</sub>)/τ)</sup>'''	+	'''F = 2 - 2<sup>((thau-a<sub>i</sub>)/τ)</sup>'''
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		+	This gives:
		+
		+	'''<math>a_i=\tau-\frac{log(2-F)}{log2}*\tau</math>'''
		+

	where F is the fraction of cells that had not initiated and τ is the generation time, or from the estimated graph of the theoretical age distribution (streaked portion).		where F is the fraction of cells that had not initiated and τ is the generation time, or from the estimated graph of the theoretical age distribution (streaked portion).

Ingvild Odsbu at 08:51, 18 August 2008

Ingvild Odsbu — Mon, 18 Aug 2008 08:51:32 GMT

←Older revision		Revision as of 08:51, 18 August 2008
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	(4 x 0.23) + (12 x 0.77) = 10.2 forks		(4 x 0.23) + (12 x 0.77) = 10.2 forks
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	[[Image:C+D_3.jpg\|left\|350px]]		[[Image:C+D_3.jpg\|left\|350px]]
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		+	12 forks → 8-origin peak in run-out histogram = 77% of the cells
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		+	6 and 4 forks → 4-origin peak in run-out histogram = 23% of the cells
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		+	The fraction of cells containing 6 forks: F = 2 - 2<sup>((τ-a<sub>t</sub>)/τ)</sup> = 2 – 2<sup>((27-2)/27)</sup> = 0.10
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		+	The fraction of cells containing 4 forks: 0.23 – 0.10 = 0.13
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		+	The average number of replication forks: (6 x 0.10) + (4 x 0.13) + (12 x 0.77) = 10.4 forks