Difference between revisions of "Part:BBa K2842669"
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===Functional Parameters===
 
===Functional Parameters===
 
<partinfo>BBa_K2842669 parameters</partinfo>
 
<partinfo>BBa_K2842669 parameters</partinfo>
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<span class='h3bb'>Sequence and Features</span>
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<partinfo>BBa_K2842669 SequenceAndFeatures</partinfo>
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<html><!--- Please copy this table containing parameters for BBa_ at the end of the parametrs section ahead of the references. ---><style type="text/css">table#AutoAnnotator {border:1px solid black; width:100%; border-collapse:collapse;} th#AutoAnnotatorHeader { border:1px solid black; width:100%; background-color: rgb(221, 221, 221);} td.AutoAnnotator1col { width:100%; border:1px solid black; } span.AutoAnnotatorSequence { font-family:'Courier New', Arial; } td.AutoAnnotatorSeqNum { text-align:right; width:2%; } td.AutoAnnotatorSeqSeq { width:98% } td.AutoAnnotatorSeqFeat1 { width:3% } td.AutoAnnotatorSeqFeat2a { width:27% } td.AutoAnnotatorSeqFeat2b { width:97% } td.AutoAnnotatorSeqFeat3 { width:70% } table.AutoAnnotatorNoBorder { border:0px; width:100%; border-collapse:collapse; } table.AutoAnnotatorWithBorder { border:1px solid black; width:100%; border-collapse:collapse; } td.AutoAnnotatorOuterAmino { border:0px solid black; width:20% } td.AutoAnnotatorInnerAmino { border:1px solid black; width:50% } td.AutoAnnotatorAminoCountingOuter { border:1px solid black; width:40%;  } td.AutoAnnotatorBiochemParOuter { border:1px solid black; width:60%; } td.AutoAnnotatorAminoCountingInner1 { width: 7.5% } td.AutoAnnotatorAminoCountingInner2 { width:62.5% } td.AutoAnnotatorAminoCountingInner3 { width:30% } td.AutoAnnotatorBiochemParInner1 { width: 5% } td.AutoAnnotatorBiochemParInner2 { width:55% } td.AutoAnnotatorBiochemParInner3 { width:40% } td.AutoAnnotatorCodonUsage1 { width: 3% } td.AutoAnnotatorCodonUsage2 { width:14.2% } td.AutoAnnotatorCodonUsage3 { width:13.8% } td.AutoAnnotatorAlignment1 { width: 3% } td.AutoAnnotatorAlignment2 { width: 10% } td.AutoAnnotatorAlignment3 { width: 87% } td.AutoAnnotatorLocalizationOuter {border:1px solid black; width:40%} td.AutoAnnotatorGOOuter {border:1px solid black; width:60%} td.AutoAnnotatorLocalization1 { width: 7.5% } td.AutoAnnotatorLocalization2 { width: 22.5% } td.AutoAnnotatorLocalization3 { width: 70% } td.AutoAnnotatorGO1 { width: 5% } td.AutoAnnotatorGO2 { width: 35% } td.AutoAnnotatorGO3 { width: 60% } td.AutoAnnotatorPredFeat1 { width:3% } td.AutoAnnotatorPredFeat2a { width:27% } td.AutoAnnotatorPredFeat3 { width:70% } div.AutoAnnotator_trans { position:absolute; background:rgb(11,140,143); background-color:rgba(11,140,143, 0.8); height:5px; top:100px; } div.AutoAnnotator_sec_helix { position:absolute; background:rgb(102,0,102); background-color:rgba(102,0,102, 0.8); height:5px; top:110px; } div.AutoAnnotator_sec_strand { position:absolute; background:rgb(245,170,26); background-color:rgba(245,170,26, 1); height:5px; top:110px; } div.AutoAnnotator_acc_buried { position:absolute; background:rgb(89,168,15); background-color:rgba(89,168,15, 0.8); height:5px; top:120px; } div.AutoAnnotator_acc_exposed { position:absolute; background:rgb(0, 0, 255); background-color:rgba(0, 0, 255, 0.8); height:5px; top:120px; } div.AutoAnnotator_dis { position:absolute; text-align:center; font-family:Arial,Helvetica,sans-serif; background:rgb(255, 200, 0); background-color:rgba(255, 200, 0, 1); height:16px; width:16px; top:80px; border-radius:50%; } </style><div id='AutoAnnotator_container_1539812017099'><table id="AutoAnnotator"><tr><!-- Time stamp in ms since 1/1/1970 1539812017099 --><th id="AutoAnnotatorHeader" colspan="2">Protein data table for BioBrick <a href="https://parts.igem.org/wiki/index.php?title=Part:BBa_<!------------------------Enter BioBrick number here------------------------>">BBa_<!------------------------Enter BioBrick number here------------------------></a> automatically created by the <a href="http://2013.igem.org/Team:TU-Munich/Results/AutoAnnotator">BioBrick-AutoAnnotator</a> version 1.0</th></tr><tr><td class="AutoAnnotator1col" colspan="2"><strong>Nucleotide sequence</strong> in <strong>RFC 10</strong>: (underlined part encodes the protein)<br><span class="AutoAnnotatorSequence">&nbsp;GCTTCTACAAACGCGGCTTCTTCCAAAGAGACCTAATACGACTCACTATAGGGGTTGTGAGCGGATAACAACCGCAAATTTAAGGATTCTCAAAA<u>ATGTTCCGC&nbsp;...&nbsp;TTCGAAAAA</u><br>TGAGTGACAGTTGAAAAGCGAAAAAAAAACCCCGCCCCTGACAGGGCGGGGTTTTTTTTGGTCTCAACGGACGACGCCGGTTACTACATTGA</span><br>&nbsp;<strong>ORF</strong> from nucleotide position 96 to 1166 (excluding stop-codon)</td></tr><tr><td class="AutoAnnotator1col" colspan="2"><strong>Amino acid sequence:</strong> (RFC 25 scars in shown in bold, other sequence features underlined; both given below)<br><span class="AutoAnnotatorSequence"><table class="AutoAnnotatorNoBorder"><tr><td class="AutoAnnotatorSeqNum">1&nbsp;<br>101&nbsp;<br>201&nbsp;<br>301&nbsp;</td><td class="AutoAnnotatorSeqSeq">MFRTNTNNIKILSPNGFSNFNGIQKVERNLYQHIIFDDDTEIKTSIDHPFGKDKIQARDVKVGDYLNSKKVLYNELVNENIFLYDPINVEKESLYITNGV<br>VSHNCYNGGRASMVSKGEAVIKEFMRFKVHMEGSMNGHEFEIEGEGEGRPYEGTQTAKLKVTKGGPLPFSWDILSPQFMYGSRAFTKHPADIPDYYKQSF<br>PEGFKWERVMNFEDGGAVTVTQDTSLEDGTLIYKVKLRGTNFPPDGPVMQKKTMGWEASTERLYPEDGVLKGDIKMALRLKDGGRYLADFKTTYKAKKPV<br>QMPGAYNVDRKLDITSHNEDYTVVEQYERSEGRHS<b>TG</b>GMDELYKGSSSE<u>WSHPQFEK</u>*</td></tr></table></span></td></tr><tr><td class="AutoAnnotator1col" colspan="2"><strong>Sequence features:</strong> (with their position in the amino acid sequence, see the <a href="http://2013.igem.org/Team:TU-Munich/Results/Software/FeatureList">list of supported features</a>)<table class="AutoAnnotatorNoBorder"><tr><td class="AutoAnnotatorSeqFeat1"></td><td class="AutoAnnotatorSeqFeat2a">RFC25 scar (shown in bold):&nbsp;</td><td class="AutoAnnotatorSeqFeat3">336 to 337</td></tr><tr><td class="AutoAnnotatorSeqFeat1"></td><td class="AutoAnnotatorSeqFeat2a"><i>Strep</i>-tag II:&nbsp;</td><td class="AutoAnnotatorSeqFeat3">350 to 357</td></tr></table></td></tr><tr><td class="AutoAnnotator1col" colspan="2"><strong>Amino acid composition:</strong><table class="AutoAnnotatorNoBorder"><tr><td class="AutoAnnotatorOuterAmino"><table class="AutoAnnotatorWithBorder"><tr><td class="AutoAnnotatorInnerAmino">Ala (A)</td><td class="AutoAnnotatorInnerAmino">12 (3.4%)</td></tr><tr><td class="AutoAnnotatorInnerAmino">Arg (R)</td><td class="AutoAnnotatorInnerAmino">15 (4.2%)</td></tr><tr><td class="AutoAnnotatorInnerAmino">Asn (N)</td><td class="AutoAnnotatorInnerAmino">20 (5.6%)</td></tr><tr><td class="AutoAnnotatorInnerAmino">Asp (D)</td><td class="AutoAnnotatorInnerAmino">23 (6.4%)</td></tr></table></td><td class="AutoAnnotatorOuterAmino"><table class="AutoAnnotatorWithBorder"><tr><td class="AutoAnnotatorInnerAmino">Cys (C)</td><td class="AutoAnnotatorInnerAmino">1 (0.3%)</td></tr><tr><td class="AutoAnnotatorInnerAmino">Gln (Q)</td><td class="AutoAnnotatorInnerAmino">11 (3.1%)</td></tr><tr><td class="AutoAnnotatorInnerAmino">Glu (E)</td><td class="AutoAnnotatorInnerAmino">29 (8.1%)</td></tr><tr><td class="AutoAnnotatorInnerAmino">Gly (G)</td><td class="AutoAnnotatorInnerAmino">33 (9.2%)</td></tr></table></td><td class="AutoAnnotatorOuterAmino"><table class="AutoAnnotatorWithBorder"><tr><td class="AutoAnnotatorInnerAmino">His (H)</td><td class="AutoAnnotatorInnerAmino">9 (2.5%)</td></tr><tr><td class="AutoAnnotatorInnerAmino">Ile (I)</td><td class="AutoAnnotatorInnerAmino">18 (5.0%)</td></tr><tr><td class="AutoAnnotatorInnerAmino">Leu (L)</td><td class="AutoAnnotatorInnerAmino">20 (5.6%)</td></tr><tr><td class="AutoAnnotatorInnerAmino">Lys (K)</td><td class="AutoAnnotatorInnerAmino">32 (9.0%)</td></tr></table></td><td class="AutoAnnotatorOuterAmino"><table class="AutoAnnotatorWithBorder"><tr><td class="AutoAnnotatorInnerAmino">Met (M)</td><td class="AutoAnnotatorInnerAmino">12 (3.4%)</td></tr><tr><td class="AutoAnnotatorInnerAmino">Phe (F)</td><td class="AutoAnnotatorInnerAmino">18 (5.0%)</td></tr><tr><td class="AutoAnnotatorInnerAmino">Pro (P)</td><td class="AutoAnnotatorInnerAmino">17 (4.8%)</td></tr><tr><td class="AutoAnnotatorInnerAmino">Ser (S)</td><td class="AutoAnnotatorInnerAmino">22 (6.2%)</td></tr></table></td><td class="AutoAnnotatorOuterAmino"><table class="AutoAnnotatorWithBorder"><tr><td class="AutoAnnotatorInnerAmino">Thr (T)</td><td class="AutoAnnotatorInnerAmino">21 (5.9%)</td></tr><tr><td class="AutoAnnotatorInnerAmino">Trp (W)</td><td class="AutoAnnotatorInnerAmino">4 (1.1%)</td></tr><tr><td class="AutoAnnotatorInnerAmino">Tyr (Y)</td><td class="AutoAnnotatorInnerAmino">18 (5.0%)</td></tr><tr><td class="AutoAnnotatorInnerAmino">Val (V)</td><td class="AutoAnnotatorInnerAmino">22 (6.2%)</td></tr></table></td></tr></table></td></tr><tr><td class="AutoAnnotatorAminoCountingOuter"><strong>Amino acid counting</strong><table class="AutoAnnotatorNoBorder"><tr><td class="AutoAnnotatorAminoCountingInner1"></td><td class="AutoAnnotatorAminoCountingInner2">Total number:</td><td class="AutoAnnotatorAminoCountingInner3">357</td></tr><tr><td class="AutoAnnotatorAminoCountingInner1"></td><td class="AutoAnnotatorAminoCountingInner2">Positively charged (Arg+Lys):</td><td class="AutoAnnotatorAminoCountingInner3">47 (13.2%)</td></tr><tr><td class="AutoAnnotatorAminoCountingInner1"></td><td class="AutoAnnotatorAminoCountingInner2">Negatively charged (Asp+Glu):</td><td class="AutoAnnotatorAminoCountingInner3">52 (14.6%)</td></tr><tr><td class="AutoAnnotatorAminoCountingInner1"></td><td class="AutoAnnotatorAminoCountingInner2">Aromatic (Phe+His+Try+Tyr):</td><td class="AutoAnnotatorAminoCountingInner3">49 (13.7%)</td></tr></table></td><td class="AutoAnnotatorBiochemParOuter"><strong>Biochemical parameters</strong><table class="AutoAnnotatorNoBorder"><tr><td class="AutoAnnotatorBiochemParInner1"></td><td class="AutoAnnotatorBiochemParInner2">Atomic composition:</td><td class="AutoAnnotatorBiochemParInner3">C<sub>1814</sub>H<sub>2783</sub>N<sub>487</sub>O<sub>554</sub>S<sub>13</sub></td></tr><tr><td class="AutoAnnotatorBiochemParInner1"></td><td class="AutoAnnotatorBiochemParInner2">Molecular mass [Da]:</td><td class="AutoAnnotatorBiochemParInner3">40694.8</td></tr><tr><td class="AutoAnnotatorBiochemParInner1"></td><td class="AutoAnnotatorBiochemParInner2">Theoretical pI:</td><td class="AutoAnnotatorBiochemParInner3">6.05</td></tr><tr><td class="AutoAnnotatorBiochemParInner1"></td><td class="AutoAnnotatorBiochemParInner2">Extinction coefficient at 280 nm [M<sup>-1</sup> cm<sup>-1</sup>]:</td><td class="AutoAnnotatorBiochemParInner3">48820 / 48883 (all Cys red/ox)</td></tr></table></td></tr><tr><td class="AutoAnnotator1col" colspan="2"><strong>Plot for hydrophobicity, charge, predicted secondary structure, solvent accessability, transmembrane helices and disulfid bridges</strong>&nbsp;<input type='button' id='hydrophobicity_charge_button' onclick='show_or_hide_plot_1539812017099()' value='Show'><span id="hydrophobicity_charge_explanation"></span><div id="hydrophobicity_charge_container" style='display:none'><div id="hydrophobicity_charge_placeholder0" style="width:100%;height:150px"></div><div id="hydrophobicity_charge_placeholder1" style="width:100%;height:150px"></div></div></td></tr><tr><td class="AutoAnnotator1col" colspan="2"><strong>Codon usage</strong><table class="AutoAnnotatorNoBorder"><tr><td class="AutoAnnotatorCodonUsage1"></td><td class="AutoAnnotatorCodonUsage2">Organism:</td><td class="AutoAnnotatorCodonUsage3"><i>E. coli</i></td><td class="AutoAnnotatorCodonUsage3"><i>B. subtilis</i></td><td class="AutoAnnotatorCodonUsage3"><i>S. cerevisiae</i></td><td class="AutoAnnotatorCodonUsage3"><i>A. thaliana</i></td><td class="AutoAnnotatorCodonUsage3"><i>P. patens</i></td><td class="AutoAnnotatorCodonUsage3">Mammals</td></tr><tr><td class="AutoAnnotatorCodonUsage1"></td><td class="AutoAnnotatorCodonUsage2">Codon quality (<a href="http://en.wikipedia.org/wiki/Codon_Adaptation_Index">CAI</a>):</td><td class="AutoAnnotatorCodonUsage3">good (0.73)</td><td class="AutoAnnotatorCodonUsage3">good (0.68)</td><td class="AutoAnnotatorCodonUsage3">acceptable (0.56)</td><td class="AutoAnnotatorCodonUsage3">good (0.65)</td><td class="AutoAnnotatorCodonUsage3">excellent (0.85)</td><td class="AutoAnnotatorCodonUsage3">excellent (0.86)</td></tr></table></td></tr><tr><td class="AutoAnnotator1col" colspan="2"><strong>Alignments</strong> (obtained from <a href='http://predictprotein.org'>PredictProtein.org</a>)<br>&nbsp;&nbsp;&nbsp;There were no alignments for this protein in the data base. The BLAST search was initialized and should be ready in a few hours.</td></tr><tr><th id='AutoAnnotatorHeader' colspan="2"><strong>Predictions</strong> (obtained from <a href='http://predictprotein.org'>PredictProtein.org</a>)</th></tr><tr><td class="AutoAnnotator1col" colspan="2">&nbsp;&nbsp;&nbsp;There were no predictions for this protein in the data base. The prediction was initialized and should be ready in a few hours.</td><tr><td class="AutoAnnotator1col" colspan="2"> The BioBrick-AutoAnnotator was created by <a href="http://2013.igem.org/Team:TU-Munich">TU-Munich 2013</a> iGEM team. For more information please see the <a href="http://2013.igem.org/Team:TU-Munich/Results/Software">documentation</a>.<br>If you have any questions, comments or suggestions, please leave us a <a href="http://2013.igem.org/Team:TU-Munich/Results/AutoAnnotator">comment</a>.</td></tr></table></div><br><!-- IMPORTANT: DON'T REMOVE THIS LINE, OTHERWISE NOT SUPPORTED FOR IE BEFORE 9 --><!--[if lte IE 8]><script language="javascript" type="text/javascript" src="http://2013.igem.org/Team:TU-Munich/excanvas.js"></script><![endif]--><script type='text/javascript' src='http://code.jquery.com/jquery-1.10.0.min.js'></script><script type='text/javascript' src='http://2013.igem.org/Team:TU-Munich/Flot.js?action=raw&ctype=text/js'></script><script>var jqAutoAnnotator = jQuery.noConflict(true);function show_or_hide_plot_1539812017099(){hydrophobicity_datapoints = [[2.5,-0.80],[3.5,-1.32],[4.5,-2.58],[5.5,-2.38],[6.5,-1.34],[7.5,-1.42],[8.5,-0.38],[9.5,1.08],[10.5,1.62],[11.5,0.40],[12.5,0.48],[13.5,-0.50],[14.5,-0.70],[15.5,-0.70],[16.5,-1.08],[17.5,0.18],[18.5,-0.44],[19.5,-1.08],[20.5,-0.02],[21.5,-0.02],[22.5,-1.36],[23.5,0.18],[24.5,-0.44],[25.5,-2.24],[26.5,-2.24],[27.5,-0.70],[28.5,-1.80],[29.5,-1.80],[30.5,-1.54],[31.5,0.06],[32.5,0.20],[33.5,1.02],[34.5,1.02],[35.5,0.96],[36.5,-0.64],[37.5,-1.68],[38.5,-2.94],[39.5,-1.34],[40.5,-1.42],[41.5,-0.86],[42.5,-0.88],[43.5,0.72],[44.5,-0.88],[45.5,-0.74],[46.5,-0.92],[47.5,-0.20],[48.5,-1.18],[49.5,-1.26],[50.5,-1.32],[51.5,-1.78],[52.5,-1.44],[53.5,-2.06],[54.5,-0.92],[55.5,-1.12],[56.5,-1.04],[57.5,-1.10],[58.5,-1.18],[59.5,-0.70],[60.5,0.12],[61.5,0.12],[62.5,-0.98],[63.5,0.56],[64.5,-0.98],[65.5,-1.06],[66.5,-1.14],[67.5,-1.66],[68.5,-1.58],[69.5,-0.12],[70.5,-0.22],[71.5,-0.14],[72.5,-0.06],[73.5,-0.14],[74.5,-0.06],[75.5,-0.50],[76.5,-0.50],[77.5,-0.50],[78.5,-0.36],[79.5,-0.64],[80.5,0.82],[81.5,1.26],[82.5,1.26],[83.5,0.04],[84.5,0.38],[85.5,-1.08],[86.5,0.02],[87.5,0.02],[88.5,-0.44],[89.5,-2.04],[90.5,-1.50],[91.5,-1.58],[92.5,-1.14],[93.5,0.54],[94.5,1.10],[95.5,0.56],[96.5,-0.28],[97.5,0.82],[98.5,0.76],[99.5,0.74],[100.5,0.80],[101.5,0.18],[102.5,-0.16],[103.5,-1.26],[104.5,-1.80],[105.5,-1.24],[106.5,-0.62],[107.5,-2.02],[108.5,-1.40],[109.5,-0.86],[110.5,-0.40],[111.5,0.52],[112.5,1.26],[113.5,0.12],[114.5,0.20],[115.5,-0.88],[116.5,-1.36],[117.5,-0.36],[118.5,1.32],[119.5,0.62],[120.5,0.62],[121.5,0.82],[122.5,0.36],[123.5,-1.44],[124.5,-0.10],[125.5,-0.18],[126.5,0.10],[127.5,-0.92],[128.5,0.36],[129.5,-0.90],[130.5,-0.20],[131.5,-1.20],[132.5,-0.18],[133.5,-1.26],[134.5,-0.64],[135.5,-1.20],[136.5,-1.74],[137.5,-1.56],[138.5,-1.56],[139.5,-0.58],[140.5,-0.64],[141.5,-0.02],[142.5,-1.28],[143.5,-0.66],[144.5,-2.26],[145.5,-1.64],[146.5,-2.46],[147.5,-2.08],[148.5,-2.26],[149.5,-2.26],[150.5,-2.26],[151.5,-1.50],[152.5,-1.88],[153.5,-1.76],[154.5,-0.70],[155.5,-1.40],[156.5,-0.50],[157.5,-0.58],[158.5,0.40],[159.5,-0.10],[160.5,-0.10],[161.5,-0.94],[162.5,-0.24],[163.5,-1.40],[164.5,-0.50],[165.5,-0.04],[166.5,0.60],[167.5,0.52],[168.5,0.66],[169.5,-0.80],[170.5,0.42],[171.5,0.62],[172.5,0.62],[173.5,0.48],[174.5,0.48],[175.5,0.14],[176.5,-0.24],[177.5,-0.34],[178.5,-0.10],[179.5,0.44],[180.5,-1.02],[181.5,-1.04],[182.5,-0.22],[183.5,-0.28],[184.5,-0.90],[185.5,-0.64],[186.5,-1.32],[187.5,-1.52],[188.5,-2.08],[189.5,-0.40],[190.5,-0.08],[191.5,-0.46],[192.5,-1.08],[193.5,-0.64],[194.5,-2.32],[195.5,-2.70],[196.5,-2.16],[197.5,-1.34],[198.5,-1.40],[199.5,-1.32],[200.5,-0.70],[201.5,0.02],[202.5,-1.32],[203.5,-1.18],[204.5,-1.18],[205.5,-2.00],[206.5,-1.72],[207.5,-0.56],[208.5,-1.08],[209.5,0.18],[210.5,0.38],[211.5,-1.16],[212.5,-1.62],[213.5,-1.00],[214.5,-1.20],[215.5,0.34],[216.5,0.90],[217.5,1.82],[218.5,1.76],[219.5,0.70],[220.5,-0.84],[221.5,-0.84],[222.5,-1.84],[223.5,-0.94],[224.5,-0.94],[225.5,-0.94],[226.5,-0.88],[227.5,-0.86],[228.5,-0.86],[229.5,0.74],[230.5,1.18],[231.5,0.48],[232.5,1.46],[233.5,-0.08],[234.5,-0.22],[235.5,-0.86],[236.5,-0.16],[237.5,-1.14],[238.5,-1.06],[239.5,-1.26],[240.5,-0.68],[241.5,-0.92],[242.5,-1.48],[243.5,-0.86],[244.5,-1.74],[245.5,-0.58],[246.5,0.12],[247.5,0.12],[248.5,-0.58],[249.5,-1.04],[250.5,-2.02],[251.5,-2.02],[252.5,-1.40],[253.5,-0.80],[254.5,-0.72],[255.5,-0.22],[256.5,-0.76],[257.5,-0.82],[258.5,-1.34],[259.5,-1.54],[260.5,-1.14],[261.5,-1.24],[262.5,-1.42],[263.5,-1.42],[264.5,-1.22],[265.5,-2.06],[266.5,-0.96],[267.5,0.12],[268.5,0.04],[269.5,0.66],[270.5,0.04],[271.5,0.10],[272.5,-1.44],[273.5,-0.28],[274.5,0.16],[275.5,1.62],[276.5,-0.18],[277.5,1.36],[278.5,0.20],[279.5,-0.86],[280.5,-1.70],[281.5,-0.88],[282.5,-2.54],[283.5,-2.02],[284.5,-0.56],[285.5,-0.12],[286.5,-0.74],[287.5,0.72],[288.5,0.20],[289.5,-0.70],[290.5,-1.20],[291.5,-0.76],[292.5,-2.10],[293.5,-0.96],[294.5,-1.60],[295.5,-2.24],[296.5,-2.30],[297.5,-0.68],[298.5,-1.74],[299.5,-0.58],[300.5,-0.12],[301.5,0.12],[302.5,-0.36],[303.5,0.08],[304.5,-1.00],[305.5,0.16],[306.5,-0.46],[307.5,-1.72],[308.5,-2.24],[309.5,-0.78],[310.5,-2.32],[311.5,-0.72],[312.5,0.04],[313.5,0.66],[314.5,-0.74],[315.5,-0.74],[316.5,-2.34],[317.5,-2.90],[318.5,-3.00],[319.5,-2.50],[320.5,-0.96],[321.5,0.58],[322.5,0.58],[323.5,0.14],[324.5,0.02],[325.5,-1.52],[326.5,-3.26],[327.5,-2.72],[328.5,-2.72],[329.5,-2.54],[330.5,-2.74],[331.5,-2.48],[332.5,-2.48],[333.5,-1.92],[334.5,-1.92],[335.5,-1.10],[336.5,-0.08],[337.5,-0.62],[338.5,-1.18],[339.5,-0.34],[340.5,-0.52],[341.5,-1.68],[342.5,-1.06],[343.5,-0.52],[344.5,-1.44],[345.5,-1.34],[346.5,-1.26],[347.5,-1.36],[348.5,-1.36],[349.5,-1.84],[350.5,-2.00],[351.5,-2.00],[352.5,-1.26],[353.5,-1.80],[354.5,-1.94]];charge_datapoints = 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= undefined;trans_datapoints = undefined;sec_helix_datapoints = undefined;sec_strand_datapoints = undefined;acc_exposed_datapoints = undefined;acc_buried_datapoints = undefined;flot_plot_options = []; flot_plot_options[0] = {grid: {borderWidth: {top: 0,right: 0,bottom: 0,left: 0}},legend: {show: false},xaxes: [{show: true,min: 0,max: 200,ticks: [[0.5, '1'], [24.5, '25'], [49.5, '50'], [74.5, '75'], [99.5, '100'], [124.5, '125'], [149.5, '150'], [174.5, '175'], [199.5, '200']],tickLength: -5}],yaxes: [{show: true,ticks: [[0, '0'], [4.5,'hydro-<br>phobic&nbsp;&nbsp;'], [-4.5,'hydro-<br>philic&nbsp;&nbsp;']],min: -4.5,max: +4.5,font: {size: 12,lineHeight: 14,style: 'italic',weight: 'bold',family: 'sans-serif',variant: 'small-caps',color: 'rgba(100,149,237,1)'}},{show: true,ticks: [[0, ''], [1,'positive<br>&nbsp;charge'], [-1,'negative<br>&nbsp;charge']],position: 'right',min: -1,max: 1,font: {size: 12,lineHeight: 14,style: 'italic',weight: 'bold',family: 'sans-serif',variant: 'small-caps',color: 'rgba(255,99,71,1)'}}]};number_of_plots = 2;for ( plot_num = 1 ; plot_num < number_of_plots ; plot_num ++){flot_plot_options[plot_num] = jqAutoAnnotator.extend(true, {} ,flot_plot_options[0]);flot_plot_options[plot_num].xaxes = [{min: plot_num*200,max: (plot_num + 1)*200,ticks: [ [plot_num*200 +  0.5, (plot_num*200 +  1).toString()], [plot_num*200 +  24.5, (plot_num*200 +  25).toString()], [plot_num*200 +  49.5, (plot_num*200 +  50).toString()], [plot_num*200 +  74.5, (plot_num*200 +  75).toString()], [plot_num*200 +  99.5, (plot_num*200 + 100).toString()], [plot_num*200 + 124.5, (plot_num*200 + 125).toString()], [plot_num*200 + 149.5, (plot_num*200 + 150).toString()], [plot_num*200 + 174.5, (plot_num*200 + 175).toString()], [plot_num*200 + 199.5, (plot_num*200 + 200).toString()] ],tickLength: -5}];};try {if( jqAutoAnnotator('#AutoAnnotator_container_1539812017099 #hydrophobicity_charge_button').val() =='Show' ){jqAutoAnnotator('#AutoAnnotator_container_1539812017099 #hydrophobicity_charge_container').css('display','block');jqAutoAnnotator('#AutoAnnotator_container_1539812017099 #hydrophobicity_charge_button').val('Hide');var description_html = '<div id=\'AutoAnnotator_plot_selectors\'>';description_html = description_html + '<br>&nbsp;<input type=\'checkbox\' id=\'hydrophobicity_checkbox\' checked=\'checked\'>&nbsp;Moving average over 5 amino acids for hydrophobicity (<img src=\'https://static.igem.org/mediawiki/2013/e/e9/TUM13_hydrophobicity_icon.png\' alt=\'blue graph\' height=\'10\'></img>)';description_html = description_html + '<br>&nbsp;<input type=\'checkbox\' id=\'charge_checkbox\' checked=\'checked\'>&nbsp;Moving average over 5 amino acids for charge (<img src=\'https://static.igem.org/mediawiki/2013/3/3e/TUM13_charge_icon.png\' alt=\'red graph\' height=\'10\'></img>)';description_html = description_html + '<br>&nbsp;<input type=\'checkbox\' id=\'dis_checkbox\' checked=\'checked\'>&nbsp;Predicted disulfid bridges (<img src=\'https://static.igem.org/mediawiki/2013/2/28/TUM13_dis_icon.png\' alt=\'yellow circle\' height=\'10\'></img>) with the number of the bridge in the center';description_html = description_html + '<br>&nbsp;<input type=\'checkbox\' id=\'trans_checkbox\' checked=\'checked\'>&nbsp;Predicted transmembrane helices (<img src=\'https://static.igem.org/mediawiki/2013/7/78/TUM13_trans_icon.png\' alt=\'turquois bars\' height=\'10\'></img>)';description_html = description_html + '<br>&nbsp;<input type=\'checkbox\' id=\'sec_checkbox\' checked=\'checked\'>&nbsp;Predicted secondary structure: Helices (<img src=\'https://static.igem.org/mediawiki/2013/b/bf/TUM13_helix_icon.png\' alt=\'violet bars\' height=\'10\'></img>) and beta-strands (<img src=\'https://static.igem.org/mediawiki/2013/b/bf/TUM13_strand_icon.png\' alt=\'yellow bars\' height=\'10\'></img>)';description_html = description_html + '<br>&nbsp;<input type=\'checkbox\' id=\'acc_checkbox\' checked=\'checked\'>&nbsp;Predicted solvent accessability: Exposed (<img src=\'https://static.igem.org/mediawiki/2013/1/16/TUM13_exposed_icon.png\' alt=\'blue bars\' height=\'10\'></img>) and buried (<img src=\'https://static.igem.org/mediawiki/2013/0/0b/TUM13_buried_icon.png\' alt=\'green bars\' height=\'10\'></img>) residues';description_html = description_html + '<br></div>';jqAutoAnnotator('#AutoAnnotator_container_1539812017099 #hydrophobicity_charge_explanation').html(description_html);plot_according_to_selectors_1539812017099();jqAutoAnnotator('#AutoAnnotator_container_1539812017099 #AutoAnnotator_plot_selectors').find('input').click(plot_according_to_selectors_1539812017099);}else{jqAutoAnnotator('#AutoAnnotator_container_1539812017099 #hydrophobicity_charge_container').css('display','none');jqAutoAnnotator('#AutoAnnotator_container_1539812017099 #hydrophobicity_charge_button').val('Show');jqAutoAnnotator('#AutoAnnotator_container_1539812017099 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<html><!--- Please copy this table containing parameters for BBa_ at the end of the parametrs section ahead of the references. ---><style type="text/css">table#AutoAnnotator {border:1px solid black; width:100%; border-collapse:collapse;} th#AutoAnnotatorHeader { border:1px solid black; width:100%; background-color: rgb(221, 221, 221);} td.AutoAnnotator1col { width:100%; border:1px solid black; } span.AutoAnnotatorSequence { font-family:'Courier New', Arial; } td.AutoAnnotatorSeqNum { text-align:right; width:2%; } td.AutoAnnotatorSeqSeq { width:98% } td.AutoAnnotatorSeqFeat1 { width:3% } td.AutoAnnotatorSeqFeat2a { width:27% } td.AutoAnnotatorSeqFeat2b { width:97% } td.AutoAnnotatorSeqFeat3 { width:70% } table.AutoAnnotatorNoBorder { border:0px; width:100%; border-collapse:collapse; } table.AutoAnnotatorWithBorder { border:1px solid black; width:100%; border-collapse:collapse; } td.AutoAnnotatorOuterAmino { border:0px solid black; width:20% } td.AutoAnnotatorInnerAmino { border:1px solid black; width:50% } td.AutoAnnotatorAminoCountingOuter { border:1px solid black; width:40%;  } td.AutoAnnotatorBiochemParOuter { border:1px solid black; width:60%; } td.AutoAnnotatorAminoCountingInner1 { width: 7.5% } td.AutoAnnotatorAminoCountingInner2 { width:62.5% } td.AutoAnnotatorAminoCountingInner3 { width:30% } td.AutoAnnotatorBiochemParInner1 { width: 5% } td.AutoAnnotatorBiochemParInner2 { width:55% } td.AutoAnnotatorBiochemParInner3 { width:40% } td.AutoAnnotatorCodonUsage1 { width: 3% } td.AutoAnnotatorCodonUsage2 { width:14.2% } td.AutoAnnotatorCodonUsage3 { width:13.8% } td.AutoAnnotatorAlignment1 { width: 3% } td.AutoAnnotatorAlignment2 { width: 10% } td.AutoAnnotatorAlignment3 { width: 87% } td.AutoAnnotatorLocalizationOuter {border:1px solid black; width:40%} td.AutoAnnotatorGOOuter {border:1px solid black; width:60%} td.AutoAnnotatorLocalization1 { width: 7.5% } td.AutoAnnotatorLocalization2 { width: 22.5% } td.AutoAnnotatorLocalization3 { width: 70% } td.AutoAnnotatorGO1 { width: 5% } td.AutoAnnotatorGO2 { width: 35% } td.AutoAnnotatorGO3 { width: 60% } td.AutoAnnotatorPredFeat1 { width:3% } td.AutoAnnotatorPredFeat2a { width:27% } td.AutoAnnotatorPredFeat3 { width:70% } div.AutoAnnotator_trans { position:absolute; background:rgb(11,140,143); background-color:rgba(11,140,143, 0.8); height:5px; top:100px; } div.AutoAnnotator_sec_helix { position:absolute; background:rgb(102,0,102); background-color:rgba(102,0,102, 0.8); height:5px; top:110px; } div.AutoAnnotator_sec_strand { position:absolute; background:rgb(245,170,26); background-color:rgba(245,170,26, 1); height:5px; top:110px; } div.AutoAnnotator_acc_buried { position:absolute; background:rgb(89,168,15); background-color:rgba(89,168,15, 0.8); height:5px; top:120px; } div.AutoAnnotator_acc_exposed { position:absolute; background:rgb(0, 0, 255); background-color:rgba(0, 0, 255, 0.8); height:5px; top:120px; } div.AutoAnnotator_dis { position:absolute; text-align:center; font-family:Arial,Helvetica,sans-serif; background:rgb(255, 200, 0); background-color:rgba(255, 200, 0, 1); height:16px; width:16px; top:80px; border-radius:50%; } </style><div id='AutoAnnotator_container_1539812017099'><table id="AutoAnnotator"><tr><!-- Time stamp in ms since 1/1/1970 1539812017099 --><th id="AutoAnnotatorHeader" colspan="2">Protein data table for BioBrick <a href="https://parts.igem.org/wiki/index.php?title=Part:BBa_<!------------------------Enter BioBrick number here------------------------>">BBa_<!------------------------Enter BioBrick number here------------------------></a> automatically created by the <a href="http://2013.igem.org/Team:TU-Munich/Results/AutoAnnotator">BioBrick-AutoAnnotator</a> version 1.0</th></tr><tr><td class="AutoAnnotator1col" colspan="2"><strong>Nucleotide sequence</strong> in <strong>RFC 10</strong>: (underlined part encodes the protein)<br><span class="AutoAnnotatorSequence">&nbsp;GCTTCTACAAACGCGGCTTCTTCCAAAGAGACCTAATACGACTCACTATAGGGGTTGTGAGCGGATAACAACCGCAAATTTAAGGATTCTCAAAA<u>ATGTTCCGC&nbsp;...&nbsp;TTCGAAAAA</u><br>TGAGTGACAGTTGAAAAGCGAAAAAAAAACCCCGCCCCTGACAGGGCGGGGTTTTTTTTGGTCTCAACGGACGACGCCGGTTACTACATTGA</span><br>&nbsp;<strong>ORF</strong> from nucleotide position 96 to 1166 (excluding stop-codon)</td></tr><tr><td class="AutoAnnotator1col" colspan="2"><strong>Amino acid sequence:</strong> (RFC 25 scars in shown in bold, other sequence features underlined; both given below)<br><span class="AutoAnnotatorSequence"><table class="AutoAnnotatorNoBorder"><tr><td class="AutoAnnotatorSeqNum">1&nbsp;<br>101&nbsp;<br>201&nbsp;<br>301&nbsp;</td><td class="AutoAnnotatorSeqSeq">MFRTNTNNIKILSPNGFSNFNGIQKVERNLYQHIIFDDDTEIKTSIDHPFGKDKIQARDVKVGDYLNSKKVLYNELVNENIFLYDPINVEKESLYITNGV<br>VSHNCYNGGRASMVSKGEAVIKEFMRFKVHMEGSMNGHEFEIEGEGEGRPYEGTQTAKLKVTKGGPLPFSWDILSPQFMYGSRAFTKHPADIPDYYKQSF<br>PEGFKWERVMNFEDGGAVTVTQDTSLEDGTLIYKVKLRGTNFPPDGPVMQKKTMGWEASTERLYPEDGVLKGDIKMALRLKDGGRYLADFKTTYKAKKPV<br>QMPGAYNVDRKLDITSHNEDYTVVEQYERSEGRHS<b>TG</b>GMDELYKGSSSE<u>WSHPQFEK</u>*</td></tr></table></span></td></tr><tr><td class="AutoAnnotator1col" colspan="2"><strong>Sequence features:</strong> (with their position in the amino acid sequence, see the <a href="http://2013.igem.org/Team:TU-Munich/Results/Software/FeatureList">list of supported features</a>)<table class="AutoAnnotatorNoBorder"><tr><td class="AutoAnnotatorSeqFeat1"></td><td class="AutoAnnotatorSeqFeat2a">RFC25 scar (shown in bold):&nbsp;</td><td class="AutoAnnotatorSeqFeat3">336 to 337</td></tr><tr><td class="AutoAnnotatorSeqFeat1"></td><td class="AutoAnnotatorSeqFeat2a"><i>Strep</i>-tag II:&nbsp;</td><td class="AutoAnnotatorSeqFeat3">350 to 357</td></tr></table></td></tr><tr><td class="AutoAnnotator1col" colspan="2"><strong>Amino acid composition:</strong><table class="AutoAnnotatorNoBorder"><tr><td class="AutoAnnotatorOuterAmino"><table class="AutoAnnotatorWithBorder"><tr><td class="AutoAnnotatorInnerAmino">Ala (A)</td><td class="AutoAnnotatorInnerAmino">12 (3.4%)</td></tr><tr><td class="AutoAnnotatorInnerAmino">Arg (R)</td><td class="AutoAnnotatorInnerAmino">15 (4.2%)</td></tr><tr><td class="AutoAnnotatorInnerAmino">Asn (N)</td><td class="AutoAnnotatorInnerAmino">20 (5.6%)</td></tr><tr><td class="AutoAnnotatorInnerAmino">Asp (D)</td><td class="AutoAnnotatorInnerAmino">23 (6.4%)</td></tr></table></td><td class="AutoAnnotatorOuterAmino"><table class="AutoAnnotatorWithBorder"><tr><td class="AutoAnnotatorInnerAmino">Cys (C)</td><td class="AutoAnnotatorInnerAmino">1 (0.3%)</td></tr><tr><td class="AutoAnnotatorInnerAmino">Gln (Q)</td><td class="AutoAnnotatorInnerAmino">11 (3.1%)</td></tr><tr><td class="AutoAnnotatorInnerAmino">Glu (E)</td><td class="AutoAnnotatorInnerAmino">29 (8.1%)</td></tr><tr><td class="AutoAnnotatorInnerAmino">Gly (G)</td><td class="AutoAnnotatorInnerAmino">33 (9.2%)</td></tr></table></td><td class="AutoAnnotatorOuterAmino"><table class="AutoAnnotatorWithBorder"><tr><td class="AutoAnnotatorInnerAmino">His (H)</td><td class="AutoAnnotatorInnerAmino">9 (2.5%)</td></tr><tr><td class="AutoAnnotatorInnerAmino">Ile (I)</td><td class="AutoAnnotatorInnerAmino">18 (5.0%)</td></tr><tr><td class="AutoAnnotatorInnerAmino">Leu (L)</td><td class="AutoAnnotatorInnerAmino">20 (5.6%)</td></tr><tr><td class="AutoAnnotatorInnerAmino">Lys (K)</td><td class="AutoAnnotatorInnerAmino">32 (9.0%)</td></tr></table></td><td class="AutoAnnotatorOuterAmino"><table class="AutoAnnotatorWithBorder"><tr><td class="AutoAnnotatorInnerAmino">Met (M)</td><td class="AutoAnnotatorInnerAmino">12 (3.4%)</td></tr><tr><td class="AutoAnnotatorInnerAmino">Phe (F)</td><td class="AutoAnnotatorInnerAmino">18 (5.0%)</td></tr><tr><td class="AutoAnnotatorInnerAmino">Pro (P)</td><td class="AutoAnnotatorInnerAmino">17 (4.8%)</td></tr><tr><td class="AutoAnnotatorInnerAmino">Ser (S)</td><td class="AutoAnnotatorInnerAmino">22 (6.2%)</td></tr></table></td><td class="AutoAnnotatorOuterAmino"><table class="AutoAnnotatorWithBorder"><tr><td class="AutoAnnotatorInnerAmino">Thr (T)</td><td class="AutoAnnotatorInnerAmino">21 (5.9%)</td></tr><tr><td class="AutoAnnotatorInnerAmino">Trp (W)</td><td class="AutoAnnotatorInnerAmino">4 (1.1%)</td></tr><tr><td class="AutoAnnotatorInnerAmino">Tyr (Y)</td><td class="AutoAnnotatorInnerAmino">18 (5.0%)</td></tr><tr><td class="AutoAnnotatorInnerAmino">Val (V)</td><td class="AutoAnnotatorInnerAmino">22 (6.2%)</td></tr></table></td></tr></table></td></tr><tr><td class="AutoAnnotatorAminoCountingOuter"><strong>Amino acid counting</strong><table class="AutoAnnotatorNoBorder"><tr><td class="AutoAnnotatorAminoCountingInner1"></td><td class="AutoAnnotatorAminoCountingInner2">Total number:</td><td class="AutoAnnotatorAminoCountingInner3">357</td></tr><tr><td class="AutoAnnotatorAminoCountingInner1"></td><td class="AutoAnnotatorAminoCountingInner2">Positively charged (Arg+Lys):</td><td class="AutoAnnotatorAminoCountingInner3">47 (13.2%)</td></tr><tr><td class="AutoAnnotatorAminoCountingInner1"></td><td class="AutoAnnotatorAminoCountingInner2">Negatively charged (Asp+Glu):</td><td class="AutoAnnotatorAminoCountingInner3">52 (14.6%)</td></tr><tr><td class="AutoAnnotatorAminoCountingInner1"></td><td class="AutoAnnotatorAminoCountingInner2">Aromatic (Phe+His+Try+Tyr):</td><td class="AutoAnnotatorAminoCountingInner3">49 (13.7%)</td></tr></table></td><td class="AutoAnnotatorBiochemParOuter"><strong>Biochemical parameters</strong><table class="AutoAnnotatorNoBorder"><tr><td class="AutoAnnotatorBiochemParInner1"></td><td class="AutoAnnotatorBiochemParInner2">Atomic composition:</td><td class="AutoAnnotatorBiochemParInner3">C<sub>1814</sub>H<sub>2783</sub>N<sub>487</sub>O<sub>554</sub>S<sub>13</sub></td></tr><tr><td class="AutoAnnotatorBiochemParInner1"></td><td class="AutoAnnotatorBiochemParInner2">Molecular mass [Da]:</td><td class="AutoAnnotatorBiochemParInner3">40694.8</td></tr><tr><td class="AutoAnnotatorBiochemParInner1"></td><td class="AutoAnnotatorBiochemParInner2">Theoretical pI:</td><td class="AutoAnnotatorBiochemParInner3">6.05</td></tr><tr><td class="AutoAnnotatorBiochemParInner1"></td><td class="AutoAnnotatorBiochemParInner2">Extinction coefficient at 280 nm [M<sup>-1</sup> cm<sup>-1</sup>]:</td><td class="AutoAnnotatorBiochemParInner3">48820 / 48883 (all Cys red/ox)</td></tr></table></td></tr><tr><td class="AutoAnnotator1col" colspan="2"><strong>Plot for hydrophobicity, charge, predicted secondary structure, solvent accessability, transmembrane helices and disulfid bridges</strong>&nbsp;<input type='button' id='hydrophobicity_charge_button' onclick='show_or_hide_plot_1539812017099()' value='Show'><span id="hydrophobicity_charge_explanation"></span><div id="hydrophobicity_charge_container" style='display:none'><div id="hydrophobicity_charge_placeholder0" style="width:100%;height:150px"></div><div id="hydrophobicity_charge_placeholder1" style="width:100%;height:150px"></div></div></td></tr><tr><td class="AutoAnnotator1col" colspan="2"><strong>Codon usage</strong><table class="AutoAnnotatorNoBorder"><tr><td class="AutoAnnotatorCodonUsage1"></td><td class="AutoAnnotatorCodonUsage2">Organism:</td><td class="AutoAnnotatorCodonUsage3"><i>E. coli</i></td><td class="AutoAnnotatorCodonUsage3"><i>B. subtilis</i></td><td class="AutoAnnotatorCodonUsage3"><i>S. cerevisiae</i></td><td class="AutoAnnotatorCodonUsage3"><i>A. thaliana</i></td><td class="AutoAnnotatorCodonUsage3"><i>P. patens</i></td><td class="AutoAnnotatorCodonUsage3">Mammals</td></tr><tr><td class="AutoAnnotatorCodonUsage1"></td><td class="AutoAnnotatorCodonUsage2">Codon quality (<a href="http://en.wikipedia.org/wiki/Codon_Adaptation_Index">CAI</a>):</td><td class="AutoAnnotatorCodonUsage3">good (0.73)</td><td class="AutoAnnotatorCodonUsage3">good (0.68)</td><td class="AutoAnnotatorCodonUsage3">acceptable (0.56)</td><td class="AutoAnnotatorCodonUsage3">good (0.65)</td><td class="AutoAnnotatorCodonUsage3">excellent (0.85)</td><td class="AutoAnnotatorCodonUsage3">excellent (0.86)</td></tr></table></td></tr><tr><td class="AutoAnnotator1col" colspan="2"><strong>Alignments</strong> (obtained from <a href='http://predictprotein.org'>PredictProtein.org</a>)<br>&nbsp;&nbsp;&nbsp;There were no alignments for this protein in the data base. The BLAST search was initialized and should be ready in a few hours.</td></tr><tr><th id='AutoAnnotatorHeader' colspan="2"><strong>Predictions</strong> (obtained from <a href='http://predictprotein.org'>PredictProtein.org</a>)</th></tr><tr><td class="AutoAnnotator1col" colspan="2">&nbsp;&nbsp;&nbsp;There were no predictions for this protein in the data base. The prediction was initialized and should be ready in a few hours.</td><tr><td class="AutoAnnotator1col" colspan="2"> The BioBrick-AutoAnnotator was created by <a href="http://2013.igem.org/Team:TU-Munich">TU-Munich 2013</a> iGEM team. For more information please see the <a href="http://2013.igem.org/Team:TU-Munich/Results/Software">documentation</a>.<br>If you have any questions, comments or suggestions, please leave us a <a href="http://2013.igem.org/Team:TU-Munich/Results/AutoAnnotator">comment</a>.</td></tr></table></div><br><!-- IMPORTANT: DON'T REMOVE THIS LINE, OTHERWISE NOT SUPPORTED FOR IE BEFORE 9 --><!--[if lte IE 8]><script language="javascript" type="text/javascript" src="http://2013.igem.org/Team:TU-Munich/excanvas.js"></script><![endif]--><script type='text/javascript' src='http://code.jquery.com/jquery-1.10.0.min.js'></script><script type='text/javascript' src='http://2013.igem.org/Team:TU-Munich/Flot.js?action=raw&ctype=text/js'></script><script>var jqAutoAnnotator = jQuery.noConflict(true);function show_or_hide_plot_1539812017099(){hydrophobicity_datapoints = 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){jqAutoAnnotator('#AutoAnnotator_container_1539812017099 #hydrophobicity_charge_container').css('display','block');jqAutoAnnotator('#AutoAnnotator_container_1539812017099 #hydrophobicity_charge_button').val('Hide');var description_html = '<div id=\'AutoAnnotator_plot_selectors\'>';description_html = description_html + '<br>&nbsp;<input type=\'checkbox\' id=\'hydrophobicity_checkbox\' checked=\'checked\'>&nbsp;Moving average over 5 amino acids for hydrophobicity (<img src=\'https://static.igem.org/mediawiki/2013/e/e9/TUM13_hydrophobicity_icon.png\' alt=\'blue graph\' height=\'10\'></img>)';description_html = description_html + '<br>&nbsp;<input type=\'checkbox\' id=\'charge_checkbox\' checked=\'checked\'>&nbsp;Moving average over 5 amino acids for charge (<img src=\'https://static.igem.org/mediawiki/2013/3/3e/TUM13_charge_icon.png\' alt=\'red graph\' height=\'10\'></img>)';description_html = description_html + '<br>&nbsp;<input type=\'checkbox\' id=\'dis_checkbox\' checked=\'checked\'>&nbsp;Predicted disulfid bridges (<img src=\'https://static.igem.org/mediawiki/2013/2/28/TUM13_dis_icon.png\' alt=\'yellow circle\' height=\'10\'></img>) with the number of the bridge in the center';description_html = description_html + '<br>&nbsp;<input type=\'checkbox\' id=\'trans_checkbox\' checked=\'checked\'>&nbsp;Predicted transmembrane helices (<img src=\'https://static.igem.org/mediawiki/2013/7/78/TUM13_trans_icon.png\' alt=\'turquois bars\' height=\'10\'></img>)';description_html = description_html + '<br>&nbsp;<input type=\'checkbox\' id=\'sec_checkbox\' checked=\'checked\'>&nbsp;Predicted secondary structure: Helices (<img src=\'https://static.igem.org/mediawiki/2013/b/bf/TUM13_helix_icon.png\' alt=\'violet bars\' height=\'10\'></img>) and beta-strands (<img src=\'https://static.igem.org/mediawiki/2013/b/bf/TUM13_strand_icon.png\' alt=\'yellow bars\' height=\'10\'></img>)';description_html = description_html + '<br>&nbsp;<input type=\'checkbox\' 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===Usage and Biology===
 
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<partinfo>BBa_K2842669 SequenceAndFeatures</partinfo>
 

Revision as of 21:40, 17 October 2018


mScarlet reporter with TerL-C intein on the N terminus

mScarlet with TerL-C intein
Function Standardised blue-white screening
Use in E. coli cells
Chassis Tested DH5α cells
Abstraction Hierarchy Composite Device
Related Device BBa_K1362101
RFC standard RFC10,RFC21,RFC23
& RFC25 compatible
Backbone pSB1C3
Submitted by [http://2018.igem.org/Team:UCL UCL iGEM 2018]

This DNA construct encodes a C terminal segment of the AceL-TerL intein fused to the N terminus of a mScarlet reporter protein which contains a C terminal StrepTag for purification. The AceL-TerL intein acts as a protein ligase that forms a peptide bond between mScarlet and another protein bound to a complementary split intein. During this process the intein splices itself out of protein and is not present in the final fusion protein. This construct is a modular platform for the creation of split intein fusion proteins through the SapI restriction sites located immediately upstream and downstream of the mScarlet reporter.BBa_K2832680 was designed to be used in conjunction with this construct as it contains the corresponding N terminal intein segment to enable intein trans-splicing.

Figure 1: Comparisons of fluorescence/OD600 of BBa_K2842669 and BBa_K1362101

A plate reader was used to measure the fluorescence of whole cell cultures.Plate reader settings: Filters were set to 540/590 , shaking at 200 rpm for 20 h at 25oC. (1-Red) BBa_K2842669 in BL21*(DE3) cells induced with 400 μM IPTG
(2-Orange) BBa_K2842669 in BL21*(DE3) cells without IPTG induction
(3-Grey) BBa_K1362101 expressed in BL21*(DE3) cells />


Experimental Results

This construct (BBa_K2842669) is also an improvement of the part BBa_K1362101. Both parts were designed to allow for the modular design and assembly of intein fusion proteins, but our part has distinct advantages and improvements. We used mScarlet as our reporter instead of mRFP1 as it is known to be brighter than mRFP1 and a fast folder. Our reporter protein mScarlet has an 11-fold increase in fluorescence/OD600 compared to BBa_K1362101 when driven by the expression from the T7 promoter (Fig3). Our construct also has a dual function as both an assembly construct and reporter for building new inteins but as an intein fusion by itself. Due to this, it can be used for both to test intein functionality in other constructs and to build new ones.


We measured the expression of BBa_K2842669 with a BMG Fluostar plate reader by analysis of it's fluorescence at varying IPTG concentrations. We found that mScarlet is very well expressed when produced from an IPTG-inducible T7 promoter. Increasing IPTG concentrations from 400 uM to 800 uM had little effect on expression levels and cell growth rate (Fig1A, Fig1B), indicating that IPTG concentrations at 400 uM are sufficient to overproduce our protein. We also tested the modularity of the construct by replacing mScarlet with with two new sequences to create the biobricks BBa_K2842710 and BBa_K2842720. We have confirmed this assembly with Sanger sequencing.

Figure 2: Expression of IP at varying IPTG concentrations
A plate reader was used to measure the fluorescence of whole cell cultures after induction with various concentrations of ITPG. Plate reader settings: Filters 540/590 , shaking at 200 rpm for 20 h at 25oC. This graph shows the fold change in fluorescence of BBa_K2832669 at a range of IPTG induction concentrations relative to uninduced BBa_K2832669. </p>
Figure 3: SDS-PAGE of BBa_K2832669

We expressed BBa_K2832669 in 50 mL cultures and extracted the intein fusion protein through sonication. The clear lysate was purified on Strep-Tactin columns then SDS-PAGE was performed on the samples. 3A shows the SDS gel when stained with instant blue for 6 hours. 3B shows the fluorescence reading from the gel. (L) Protein standard numbers are in kDa (1) The clear lysate of BBa_K2832669 before purification (2) After purification on Streptactin columns





Functional Parameters

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal NotI site found at 941
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI site found at 1224
    Illegal BsaI.rc site found at 28
    Illegal SapI site found at 1130
    Illegal SapI.rc site found at 419


Protein data table for BioBrick BBa_ automatically created by the BioBrick-AutoAnnotator version 1.0
Nucleotide sequence in RFC 10: (underlined part encodes the protein)
 GCTTCTACAAACGCGGCTTCTTCCAAAGAGACCTAATACGACTCACTATAGGGGTTGTGAGCGGATAACAACCGCAAATTTAAGGATTCTCAAAAATGTTCCGC ... TTCGAAAAA
TGAGTGACAGTTGAAAAGCGAAAAAAAAACCCCGCCCCTGACAGGGCGGGGTTTTTTTTGGTCTCAACGGACGACGCCGGTTACTACATTGA
 ORF from nucleotide position 96 to 1166 (excluding stop-codon)
Amino acid sequence: (RFC 25 scars in shown in bold, other sequence features underlined; both given below)
1 
101 
201 
301 		MFRTNTNNIKILSPNGFSNFNGIQKVERNLYQHIIFDDDTEIKTSIDHPFGKDKIQARDVKVGDYLNSKKVLYNELVNENIFLYDPINVEKESLYITNGV
VSHNCYNGGRASMVSKGEAVIKEFMRFKVHMEGSMNGHEFEIEGEGEGRPYEGTQTAKLKVTKGGPLPFSWDILSPQFMYGSRAFTKHPADIPDYYKQSF
PEGFKWERVMNFEDGGAVTVTQDTSLEDGTLIYKVKLRGTNFPPDGPVMQKKTMGWEASTERLYPEDGVLKGDIKMALRLKDGGRYLADFKTTYKAKKPV
QMPGAYNVDRKLDITSHNEDYTVVEQYERSEGRHSTGGMDELYKGSSSEWSHPQFEK*
Sequence features: (with their position in the amino acid sequence, see the list of supported features)
RFC25 scar (shown in bold): 336 to 337
Strep-tag II: 350 to 357
Amino acid composition:
Ala (A)12 (3.4%)
Arg (R)15 (4.2%)
Asn (N)20 (5.6%)
Asp (D)23 (6.4%)
Cys (C)1 (0.3%)
Gln (Q)11 (3.1%)
Glu (E)29 (8.1%)
Gly (G)33 (9.2%)
His (H)9 (2.5%)
Ile (I)18 (5.0%)
Leu (L)20 (5.6%)
Lys (K)32 (9.0%)
Met (M)12 (3.4%)
Phe (F)18 (5.0%)
Pro (P)17 (4.8%)
Ser (S)22 (6.2%)
Thr (T)21 (5.9%)
Trp (W)4 (1.1%)
Tyr (Y)18 (5.0%)
Val (V)22 (6.2%)
Amino acid counting
Total number:357
Positively charged (Arg+Lys):47 (13.2%)
Negatively charged (Asp+Glu):52 (14.6%)
Aromatic (Phe+His+Try+Tyr):49 (13.7%)
Biochemical parameters
Atomic composition:C1814H2783N487O554S13
Molecular mass [Da]:40694.8
Theoretical pI:6.05
Extinction coefficient at 280 nm [M-1 cm-1]:48820 / 48883 (all Cys red/ox)
Plot for hydrophobicity, charge, predicted secondary structure, solvent accessability, transmembrane helices and disulfid bridges 
Codon usage
Organism:E. coliB. subtilisS. cerevisiaeA. thalianaP. patensMammals
Codon quality (CAI):good (0.73)good (0.68)acceptable (0.56)good (0.65)excellent (0.85)excellent (0.86)
Alignments (obtained from PredictProtein.org)
   There were no alignments for this protein in the data base. The BLAST search was initialized and should be ready in a few hours.
Predictions (obtained from PredictProtein.org)
   There were no predictions for this protein in the data base. The prediction was initialized and should be ready in a few hours.
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