Davidson Missouri W/Web tool - Revision history

Synthetic Students: Web tool moved to Davidson Missouri W/Web tool

2007-07-11T19:51:42Z

Web tool moved to Davidson Missouri W/Web tool

Synthetic Students at 19:22, 11 July 2007

2007-07-11T19:22:33Z

Synthetic Students: /* How the Primers Work */

2007-07-11T19:19:37Z

‎How the Primers Work

Synthetic Students: /* Tutorial */ add section how primers work

2007-07-11T19:11:00Z

‎Tutorial: add section how primers work

Synthetic Students: /* The Output */ image caption

2007-07-11T19:09:34Z

‎The Output: image caption

WikiSysop: /* Future Features */

2007-07-08T01:05:17Z

‎Future Features

WikiSysop: /* Tutorial */ Melting point algorithm

2007-07-08T00:31:59Z

‎Tutorial: Melting point algorithm

WikiSysop: italicize HixC

2007-07-08T00:24:35Z

italicize HixC

WikiSysop: /* The Output */

2007-07-01T17:23:59Z

‎The Output

WikiSysop: /* The Output */ Picture of primers

2007-07-01T17:11:12Z

‎The Output: Picture of primers

← Older revision		Revision as of 19:22, 11 July 2007
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		+	<center>[[Davidson/Missouri Western iGEM 2007\| <span style="color:red">Home</span>]] \| [[Background Information\| <span style="color:red">Background Information</span>]] \| [[Solving the HPP in vivo\| <span style="color:red">Current Project: Solving the Hamiltonian Path Problem ''in vivo''</span>]] \| [[Probability and Statistics\| <span style="color:red">Probability and Statistics</span>]] \| [[Gene splitting\| <span style="color:red"> Gene Splitting </span>]] \| [[Controlling Expression\| <span style="color:red"> Controlling Expression </span>]] \| [[Traveling Salesperson Problem\| <span style="color:red">Traveling Salesperson Problem</span> ]] \| [[Backwards promotion and read-through transcription\| <span style="color:red">Backwards promotion and Read-Through Transcription</span>]] \| [[Resources and Citations\|<span style="color:red">Resources and Citations</span>]]</center>
		+
		+	<hr>
		+
	The gene-flipping mechanism that powers our computer requires the positioning of ''HixC'' sites within reporter genes' coding sequences. Through the use of PCR, a gene can be split into two pieces with BioBrick ends attached at the extremities. This allows for the ligation with a ''HixC'' site in the middle. Our team has developed a web-based tool to automate the generation of these PCR primers. It also predicts the resultant DNA sequences for the two intermediary parts, the final part, and the resultant polypeptide. By using this program we hope to make the gene-splitting process faster and to remove the potential for human error. The tool can be accessed at [http://gcat.davidson.edu/iGEM07/genesplitter.html http://gcat.davidson.edu/iGEM07/genesplitter.html].		The gene-flipping mechanism that powers our computer requires the positioning of ''HixC'' sites within reporter genes' coding sequences. Through the use of PCR, a gene can be split into two pieces with BioBrick ends attached at the extremities. This allows for the ligation with a ''HixC'' site in the middle. Our team has developed a web-based tool to automate the generation of these PCR primers. It also predicts the resultant DNA sequences for the two intermediary parts, the final part, and the resultant polypeptide. By using this program we hope to make the gene-splitting process faster and to remove the potential for human error. The tool can be accessed at [http://gcat.davidson.edu/iGEM07/genesplitter.html http://gcat.davidson.edu/iGEM07/genesplitter.html].

← Older revision		Revision as of 19:19, 11 July 2007
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	==How the Primers Work==		==How the Primers Work==
		+	We use these DNA sequences as PCR primers to split our genes. There are a total of 4 primers to use: there are two gene portions, before and after the split, and each portion needs a forward and a reverse primer. These primers are denoted A, B, C, and D.
		+
		+	* Primer A is the forward primer for the first portion
		+	* Primer B is the reverse primer for the first portion
		+	* Primer C is the forward primer for the second portion
		+	* Primer D is the reverse primer for the second portion
		+
		+	If the user wants an insertion in a DNA sequence:
		+
		+	[[Image:Insertionpoint.jpg]]
		+
		+	Then the primers and inserted sequences will be as follows:
		+
		+	[[Image:Primers.jpg]]

	==Melting Point Calculations==		==Melting Point Calculations==

← Older revision		Revision as of 19:11, 11 July 2007
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	Once the primers have been displayed, the tool provides some additional useful information. The two intermediate parts, with BioBrick ends, are shown. This is useful if it is necessary to verify the PCR results by sequencing. The DNA sequences for the scars and the ''HixC'' site are provided, as well as which amino acids these will produce (along with the user-inputted 39th base). The final product is then displayed, with the inserted ''HixC'' site and BioBrick ends at the extremities. Also shown are what becomes translated (the final product without the BioBrick ends), and the translation itself.		Once the primers have been displayed, the tool provides some additional useful information. The two intermediate parts, with BioBrick ends, are shown. This is useful if it is necessary to verify the PCR results by sequencing. The DNA sequences for the scars and the ''HixC'' site are provided, as well as which amino acids these will produce (along with the user-inputted 39th base). The final product is then displayed, with the inserted ''HixC'' site and BioBrick ends at the extremities. Also shown are what becomes translated (the final product without the BioBrick ends), and the translation itself.
		+
		+	==How the Primers Work==

	==Melting Point Calculations==		==Melting Point Calculations==

@@ Line 19: / Line 19: @@
 The first calculations are the 4 primers for the two PCR reactions.  The display is color-coded so that the restriction enzyme sites in the BioBrick ends are easily distinguishable.  After the BioBrick ends, between 20 and 25 bases should follow.  These bases should function as PCR primers to the given gene.  Their length is optimized to match melting temperatures as much as possible.  This is done using Proligo's formula from their own web page.  Note that they use two formulas: a simple one for sequences of size less than 20, and a more complicated one for larger sequences.  Below the primer sequences are their computed melting temperatures and GC content, as well as their lengths (excluding BioBrick ends).
-[[Image:color_coded_primers.png|thumb|500px|center|]]
+[[Image:color_coded_primers.png|thumb|500px|center|Sample PCR primers generated by the tool.]]
 Once the primers have been displayed, the tool provides some additional useful information.  The two intermediate parts, with BioBrick ends, are shown.  This is useful if it is necessary to verify the PCR results by sequencing.  The DNA sequences for the scars and the ''HixC'' site are provided, as well as which amino acids these will produce (along with the user-inputted 39th base).  The final product is then displayed, with the inserted ''HixC'' site and BioBrick ends at the extremities.  Also shown are what becomes translated (the final product without the BioBrick ends), and the translation itself.

@@ Line 35: / Line 35: @@
 =Future Features=
 ==Displaying the Split in 3-D==
 ==Finding More Genes to Split==

← Older revision	Revision as of 19:51, 11 July 2007
(No difference)

← Older revision		Revision as of 00:31, 8 July 2007
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	Once the primers have been displayed, the tool provides some additional useful information. The two intermediate parts, with BioBrick ends, are shown. This is useful if it is necessary to verify the PCR results by sequencing. The DNA sequences for the scars and the ''HixC'' site are provided, as well as which amino acids these will produce (along with the user-inputted 39th base). The final product is then displayed, with the inserted ''HixC'' site and BioBrick ends at the extremities. Also shown are what becomes translated (the final product without the BioBrick ends), and the translation itself.		Once the primers have been displayed, the tool provides some additional useful information. The two intermediate parts, with BioBrick ends, are shown. This is useful if it is necessary to verify the PCR results by sequencing. The DNA sequences for the scars and the ''HixC'' site are provided, as well as which amino acids these will produce (along with the user-inputted 39th base). The final product is then displayed, with the inserted ''HixC'' site and BioBrick ends at the extremities. Also shown are what becomes translated (the final product without the BioBrick ends), and the translation itself.
		+
		+	==Melting Point Calculations==
		+	Melting-point temperatures for DNA strands are calculated using the following algorithm:
		+
		+	If the length of the strand is 20 or less:
		+
		+	Return 4(# Gs + # Cs) + 2(# As + # Ts).
		+
		+	Otherwise:
		+
		+	Compute sigma_dH and sigma_dS based on enthalpy and entropy values for base-pairs. Return (-1000 * sigma_dH / (-10.8 - sigma_dS + 1.987 * -23.5) - 273.15 + 16.6 * -1.3).

	=Future Features=		=Future Features=

@@ Line 1: / Line 1: @@
-The gene-flipping mechanism that powers our computer requires the positioning of HixC sites within reporter genes' coding sequences.  Through the use of PCR, a gene can be split into two pieces with BioBrick ends attached at the extremities.  This allows for the ligation with a HixC site in the middle.  Our team has developed a web-based tool to automate the generation of these PCR primers.  It also predicts the resultant DNA sequences for the two intermediary parts, the final part, and the resultant polypeptide.  By using this program we hope to make the gene-splitting process faster and to remove the potential for human error.  The tool can be accessed at [http://gcat.davidson.edu/iGEM07/genesplitter.html http://gcat.davidson.edu/iGEM07/genesplitter.html].
+The gene-flipping mechanism that powers our computer requires the positioning of ''HixC'' sites within reporter genes' coding sequences.  Through the use of PCR, a gene can be split into two pieces with BioBrick ends attached at the extremities.  This allows for the ligation with a ''HixC'' site in the middle.  Our team has developed a web-based tool to automate the generation of these PCR primers.  It also predicts the resultant DNA sequences for the two intermediary parts, the final part, and the resultant polypeptide.  By using this program we hope to make the gene-splitting process faster and to remove the potential for human error.  The tool can be accessed at [http://gcat.davidson.edu/iGEM07/genesplitter.html http://gcat.davidson.edu/iGEM07/genesplitter.html].
 =Tutorial=
@@ Line 9: / Line 9: @@
 # The DNA sequence for the gene to split.
-# The '''amino acid''' number where the insertion is desired.  The HixC site will be inserted immediately after the given amino acid.
+# The '''amino acid''' number where the insertion is desired.  The ''HixC'' site will be inserted immediately after the given amino acid.
-# The choice for an extra nucleotide.  Inserting a HixC site amounts to an addition of 38 bases.  Unless a 39th base is added, a frameshift will occur since 38 is not a multiple of 3.  The nucleotide choice results in either a glutamate or aspartate.
+# The choice for an extra nucleotide.  Inserting a ''HixC'' site amounts to an addition of 38 bases.  Unless a 39th base is added, a frameshift will occur since 38 is not a multiple of 3.  The nucleotide choice results in either a glutamate or aspartate.
 Once the user has entered this information, he or she can then click the 'submit' button to initiate the calculations.
@@ Line 21: / Line 21: @@
 [[Image:color_coded_primers.png|thumb|500px|center|]]
-Once the primers have been displayed, the tool provides some additional useful information.  The two intermediate parts, with BioBrick ends, are shown.  This is useful if it is necessary to verify the PCR results by sequencing.  The DNA sequences for the scars and the HixC site are provided, as well as which amino acids these will produce (along with the user-inputted 39th base).  The final product is then displayed, with the inserted HixC site and BioBrick ends at the extremities.  Also shown are what becomes translated (the final product without the BioBrick ends), and the translation itself.
+Once the primers have been displayed, the tool provides some additional useful information.  The two intermediate parts, with BioBrick ends, are shown.  This is useful if it is necessary to verify the PCR results by sequencing.  The DNA sequences for the scars and the ''HixC'' site are provided, as well as which amino acids these will produce (along with the user-inputted 39th base).  The final product is then displayed, with the inserted ''HixC'' site and BioBrick ends at the extremities.  Also shown are what becomes translated (the final product without the BioBrick ends), and the translation itself.
 =Future Features=

@@ Line 15: / Line 15: @@
 ==The Output==
-Once the user has submitted the information a new page opens.  Before any calculations are done, some information is provided to allow the user to verify that the tool is analyzing the correct sequence.  The first output is the original sequence.  This should be identical to the sequence provided by the user.  The next two sequences show the DNA sequences that lie before and after the insertion point.  The tool then restates the insertion point, by amino acid number.
+Once the user has submitted the information a new page opens.  Before any calculations are done, some information is provided to allow the user to verify that the tool is analyzing the correct sequence.  The first output is the original sequence.  This should be identical to the sequence provided by the user.  The next two sequences show the DNA sequences that lie before and after the insertion point.  The tool then restates the insertion point and the extra base.
 The first calculations are the 4 primers for the two PCR reactions.  The display is color-coded so that the restriction enzyme sites in the BioBrick ends are easily distinguishable.  After the BioBrick ends, between 20 and 25 bases should follow.  These bases should function as PCR primers to the given gene.  Their length is optimized to match melting temperatures as much as possible.  This is done using Proligo's formula from their own web page.  Note that they use two formulas: a simple one for sequences of size less than 20, and a more complicated one for larger sequences.  Below the primer sequences are their computed melting temperatures and GC content, as well as their lengths (excluding BioBrick ends).

← Older revision		Revision as of 17:11, 1 July 2007
Line 18:		Line 18:

	The first calculations are the 4 primers for the two PCR reactions. The display is color-coded so that the restriction enzyme sites in the BioBrick ends are easily distinguishable. After the BioBrick ends, between 20 and 25 bases should follow. These bases should function as PCR primers to the given gene. Their length is optimized to match melting temperatures as much as possible. This is done using Proligo's formula from their own web page. Note that they use two formulas: a simple one for sequences of size less than 20, and a more complicated one for larger sequences. Below the primer sequences are their computed melting temperatures and GC content, as well as their lengths (excluding BioBrick ends).		The first calculations are the 4 primers for the two PCR reactions. The display is color-coded so that the restriction enzyme sites in the BioBrick ends are easily distinguishable. After the BioBrick ends, between 20 and 25 bases should follow. These bases should function as PCR primers to the given gene. Their length is optimized to match melting temperatures as much as possible. This is done using Proligo's formula from their own web page. Note that they use two formulas: a simple one for sequences of size less than 20, and a more complicated one for larger sequences. Below the primer sequences are their computed melting temperatures and GC content, as well as their lengths (excluding BioBrick ends).
		+
		+	[[Image:color_coded_primers.png\|thumb\|500px\|center\|]]

	Once the primers have been displayed, the tool provides some additional useful information. The two intermediate parts, with BioBrick ends, are shown. This is useful if it is necessary to verify the PCR results by sequencing. The DNA sequences for the scars and the HixC site are provided, as well as which amino acids these will produce (along with the user-inputted 39th base). The final product is then displayed, with the inserted HixC site and BioBrick ends at the extremities. Also shown are what becomes translated (the final product without the BioBrick ends), and the translation itself.		Once the primers have been displayed, the tool provides some additional useful information. The two intermediate parts, with BioBrick ends, are shown. This is useful if it is necessary to verify the PCR results by sequencing. The DNA sequences for the scars and the HixC site are provided, as well as which amino acids these will produce (along with the user-inputted 39th base). The final product is then displayed, with the inserted HixC site and BioBrick ends at the extremities. Also shown are what becomes translated (the final product without the BioBrick ends), and the translation itself.