http://2014.igem.org/wiki/index.php?title=Team:SJTU-BioX-Shanghai/Part3_TAL_Improvement&feed=atom&action=historyTeam:SJTU-BioX-Shanghai/Part3 TAL Improvement - Revision history2024-03-29T01:59:27ZRevision history for this page on the wikiMediaWiki 1.16.5http://2014.igem.org/wiki/index.php?title=Team:SJTU-BioX-Shanghai/Part3_TAL_Improvement&diff=358527&oldid=prevHoPe Hao at 21:53, 17 October 20142014-10-17T21:53:15Z<p></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <p></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <p></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> The recognition sequence of the TALE protein:</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> The recognition sequence of the TALE protein:</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div> <center><font size="5" color="red"><del class="diffchange diffchange-inline">TCGATATCAAGC</del></font></center></p></div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div> <center><font size="5" color="red"><ins class="diffchange diffchange-inline">TTCGATATCAAGCT</ins></font></center></p></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <p>All parts are under artificial synthesis process, so there is few results to present, which can prove our changes are effective. However, with the principle of complementary base pairing, our choice should be better than original version. And if you want our data or use our method to create your own best sticky ends, just contact us!</p></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <p>All parts are under artificial synthesis process, so there is few results to present, which can prove our changes are effective. However, with the principle of complementary base pairing, our choice should be better than original version. And if you want our data or use our method to create your own best sticky ends, just contact us!</p></div></td></tr>
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</table>HoPe Haohttp://2014.igem.org/wiki/index.php?title=Team:SJTU-BioX-Shanghai/Part3_TAL_Improvement&diff=358496&oldid=prevHoPe Hao at 21:52, 17 October 20142014-10-17T21:52:52Z<p></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> The recognition sequence of the TALE protein:</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> The recognition sequence of the TALE protein:</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <center><font size="5" color="red">TCGATATCAAGC</font></center></p></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <center><font size="5" color="red">TCGATATCAAGC</font></center></p></div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div> <p>All parts are under artificial synthesis process, so there is few results, which can prove our changes are <del class="diffchange diffchange-inline">useful</del>. However, with the principle of complementary base pairing, our choice should be better than original <del class="diffchange diffchange-inline">vision</del>. And if you want our data or use our method to create your own best sticky ends, just contact us!</p></div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div> <p>All parts are under artificial synthesis process, so there is few results <ins class="diffchange diffchange-inline">to present</ins>, which can prove our changes are <ins class="diffchange diffchange-inline">effective</ins>. However, with the principle of complementary base pairing, our choice should be better than original <ins class="diffchange diffchange-inline">version</ins>. And if you want our data or use our method to create your own best sticky ends, just contact us!</p></div></td></tr>
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</table>HoPe Haohttp://2014.igem.org/wiki/index.php?title=Team:SJTU-BioX-Shanghai/Part3_TAL_Improvement&diff=358367&oldid=prevHoPe Hao at 21:51, 17 October 20142014-10-17T21:51:19Z<p></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><center><small><strong>Figure 1.4.10 Position in TALE amino acids sequence </strong></small></center></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><center><small><strong>Figure 1.4.10 Position in TALE amino acids sequence </strong></small></center></div></td></tr>
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<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div><h2>Reconstruct DNA <del class="diffchange diffchange-inline">Sequence</del></h2></div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><h2>Reconstruct DNA <ins class="diffchange diffchange-inline">sequences</ins></h2></div></td></tr>
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<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div> <p>Two main factors to reconstruct DNA <del class="diffchange diffchange-inline">sequence</del>:</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div> <p>Two main factors to reconstruct DNA <ins class="diffchange diffchange-inline">sequences</ins>:<ins class="diffchange diffchange-inline"><br></ins></div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div> 1.Use the table of best combination and rearrange the sticky ends <del class="diffchange diffchange-inline">with </del>your demand.</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div> 1.Use the table of best combination and rearrange the sticky ends <ins class="diffchange diffchange-inline">according to </ins>your demand.<ins class="diffchange diffchange-inline"><br></ins></div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div> 2.<del class="diffchange diffchange-inline">No </del>BsmBI recognition sequence in the reconstruct DNA sequence.</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div> 2.<ins class="diffchange diffchange-inline">There are no </ins>BsmBI recognition sequence in the reconstruct DNA sequence.<ins class="diffchange diffchange-inline"><br></ins></div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div> Final DNA Sequence for TALE protein:</p></div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div> Final DNA Sequence for <ins class="diffchange diffchange-inline">NEW </ins>TALE protein:</p></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <pre></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <pre></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> 1 CTGACCCCGG AACAGGTGGT GGCCATTGCA AGCAACGGTG GTGGCAAGCA GGCCCTGGAG</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> 1 CTGACCCCGG AACAGGTGGT GGCCATTGCA AGCAACGGTG GTGGCAAGCA GGCCCTGGAG</div></td></tr>
</table>HoPe Haohttp://2014.igem.org/wiki/index.php?title=Team:SJTU-BioX-Shanghai/Part3_TAL_Improvement&diff=358142&oldid=prevHoPe Hao at 21:48, 17 October 20142014-10-17T21:48:41Z<p></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <h2>How to improve the Golden Gate sticky ends? A big Table!</h2></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <h2>How to improve the Golden Gate sticky ends? A big Table!</h2></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> </div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> </div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div> <p>Three <del class="diffchange diffchange-inline">basic </del>key questions need to be answered:</p></div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div> <p>Three key questions need to be answered:</p></div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div> <p>1. <del class="diffchange diffchange-inline">Whether it’s </del>possible to find perfect match pair?</p></div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div> <p>1. <ins class="diffchange diffchange-inline">Is it </ins>possible to find perfect match pair?</p></div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div> <p>2. <del class="diffchange diffchange-inline">Whether </del>we <del class="diffchange diffchange-inline">can </del>find a certain number of sticky ends with least possibility <del class="diffchange diffchange-inline">to be mismatched</del>?</p></div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div> <p>2. <ins class="diffchange diffchange-inline">Can </ins>we find a certain number of sticky ends with least <ins class="diffchange diffchange-inline">mismatch </ins>possibility?</p></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <p>3. How to make this sticky-end score table?</p></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <p>3. How to make this sticky-end score table?</p></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <h3>Key algorithms derived from BLAST algorithm</h3></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <h3>Key algorithms derived from BLAST algorithm</h3></div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div> <p>Loose rule: Match: 1; Mismatch:<del class="diffchange diffchange-inline">-1</del>; Gap: 0</p></div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div> <p>Loose rule: Match: 1; Mismatch: <ins class="diffchange diffchange-inline">0</ins>; Gap: 0</p></div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div> <p>Strict rule: Match: 1; Mismatch:0; Gap: <del class="diffchange diffchange-inline">-</del>1</p></div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div> <p>Strict rule: Match: 1; Mismatch: 0; Gap: 1</p></div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div> <p>The sticky end is composed of four bases, which means that we can design 256 types of sticky ends <del class="diffchange diffchange-inline">at most.</del></div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div> <p>The sticky end is composed of four bases, which means that we can design 256 types of sticky ends<ins class="diffchange diffchange-inline">, which are </ins>represented as a 256*256 table. </p></div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div><del class="diffchange diffchange-inline"> The forming pair is </del>represented as a 256*256 table. </p></div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <h3>Find target groups of sticky ends</h3></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <h3>Find target groups of sticky ends</h3></div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div> <p>To solve the TALE parts problem, we need find seven sticky ends, and the similarity score<del class="diffchange diffchange-inline">(hereafter referred to as Score) </del>of each pair <del class="diffchange diffchange-inline">of </del>them are less than or equal to 1.</p></div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div> <p>To solve the TALE parts problem, we need find seven sticky ends, and the similarity score of each pair <ins class="diffchange diffchange-inline">in </ins>them are less than or equal to 1.</p></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <center><img src="https://static.igem.org/mediawiki/2014/1/12/Choices_for_group.png" width=400px></img></center></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <center><img src="https://static.igem.org/mediawiki/2014/1/12/Choices_for_group.png" width=400px></img></center></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><center><small><strong>Figure 1.4.7 Sticky ends choices table </strong></small></center></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><center><small><strong>Figure 1.4.7 Sticky ends choices table </strong></small></center></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <p>When we select Strict Algorithm to find these ends, it is impossible to find seven sticky ends, that each pair of them has score no more than 1. So we have to select Loose Algorithm.</p></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <p>When we select Strict Algorithm to find these ends, it is impossible to find seven sticky ends, that each pair of them has score no more than 1. So we have to select Loose Algorithm.</p></div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div> <h3 ><del class="diffchange diffchange-inline">Four </del>basepair sticky ends <del class="diffchange diffchange-inline">convert </del>to amino acid <del class="diffchange diffchange-inline">pair</del></h3></div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div> <h3 > <ins class="diffchange diffchange-inline">Convert four-</ins>basepair sticky ends to amino acid <ins class="diffchange diffchange-inline">pairs</ins></h3></div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div> <p><del class="diffchange diffchange-inline">What we are caring </del>about <del class="diffchange diffchange-inline">is </del>whether two amino acids <del class="diffchange diffchange-inline">can be </del>located on <del class="diffchange diffchange-inline">my </del>target sequence, rather than the 4bp <del class="diffchange diffchange-inline">bases</del>. So we should convert the sticky ends <del class="diffchange diffchange-inline">information </del>to <del class="diffchange diffchange-inline">2 </del>amino <del class="diffchange diffchange-inline">acids</del>.</p></div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div> <p><ins class="diffchange diffchange-inline">We care </ins>about whether two amino acids located on <ins class="diffchange diffchange-inline">our </ins>target sequence, rather than the 4bp. So we should convert the sticky ends <ins class="diffchange diffchange-inline">sequence </ins>to amino <ins class="diffchange diffchange-inline">acid pairs</ins>.</p></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <center><img src="https://static.igem.org/mediawiki/2014/4/45/Amino_acid_table.png"width=600px id="dianweidian7"></img></center></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <center><img src="https://static.igem.org/mediawiki/2014/4/45/Amino_acid_table.png"width=600px id="dianweidian7"></img></center></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><center><small><strong>Figure 1.4.8 4bp sticky ends convert to Amino acid table </strong></small></center></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><center><small><strong>Figure 1.4.8 4bp sticky ends convert to Amino acid table </strong></small></center></div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div> <p>Based on the above table, we are able to calculate the total scores of each combination and find the <del class="diffchange diffchange-inline">least </del>one.</p></div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div> <p>Based on the above table, we are able to calculate the total scores of each combination and find the <ins class="diffchange diffchange-inline">best </ins>one.</p></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <h2>Best choice for seven sticky ends on TALE protein</h2></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <h2>Best choice for seven sticky ends on TALE protein</h2></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <p>Best combination:</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <p>Best combination:</div></td></tr>
</table>HoPe Haohttp://2014.igem.org/wiki/index.php?title=Team:SJTU-BioX-Shanghai/Part3_TAL_Improvement&diff=357099&oldid=prevHoPe Hao at 21:38, 17 October 20142014-10-17T21:38:13Z<p></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> The table shows that more than 30% of pairs’ score is equal to 3, which means that the possibility of mismatch cannot be neglected.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> The table shows that more than 30% of pairs’ score is equal to 3, which means that the possibility of mismatch cannot be neglected.</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div> Even if we employ the relatively loose rule to <del class="diffchange diffchange-inline">calculate </del>the similarity, we can still find that error rates cannot be neglected.</p></div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div> Even if we employ the relatively loose rule to <ins class="diffchange diffchange-inline">evaluate </ins>the similarity, we can still find that error rates cannot be neglected.</p></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <center><img src="https://static.igem.org/mediawiki/2014/9/9b/Tal_%E8%A1%A8%E6%A0%BC%E7%B2%98%E6%80%A7%E6%9C%AB%E7%AB%AF2.png" width=400px></img></center></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <center><img src="https://static.igem.org/mediawiki/2014/9/9b/Tal_%E8%A1%A8%E6%A0%BC%E7%B2%98%E6%80%A7%E6%9C%AB%E7%AB%AF2.png" width=400px></img></center></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><center><small><strong>Figure 1.4.6 Loose rules score table </strong></small></center></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><center><small><strong>Figure 1.4.6 Loose rules score table </strong></small></center></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <h2> Why not other sticky ends?</h2></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <h2> Why not other sticky ends?</h2></div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div> <h3>The <del class="diffchange diffchange-inline">Reason </del>why Freiburg used these sticky ends</h3></div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div> <h3>The <ins class="diffchange diffchange-inline">reason </ins>why Freiburg used these sticky ends</h3></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <p>Failed to contact the original designers of these sticky ends, what we can do is just to find feasible advantages of these combinations.</p></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <p>Failed to contact the original designers of these sticky ends, what we can do is just to find feasible advantages of these combinations.</p></div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div> <p><del class="diffchange diffchange-inline">Review </del>the TALE <del class="diffchange diffchange-inline">repeated </del>amino acids sequence:</p></div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div> <p><ins class="diffchange diffchange-inline">Let's look at </ins>the TALE <ins class="diffchange diffchange-inline">direpeat unit </ins>amino acids sequence:</p></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <p><center>LTPEQVVAIAS(XX)GGKQALETVQRLLPVLCQAHG(34aa)</center></p></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <p><center>LTPEQVVAIAS(XX)GGKQALETVQRLLPVLCQAHG(34aa)</center></p></div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div> <p id="dianweidian6">The first amino acid is Leu, which is essential for all connection process. There are six different <del class="diffchange diffchange-inline">types of base arrangement </del>for Leu<del class="diffchange diffchange-inline">, one of the most number of base arrangement</del>. </p></div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div> <p id="dianweidian6">The first amino acid is Leu, which is essential for all connection process. There are six different <ins class="diffchange diffchange-inline">codons </ins>for Leu. </p></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <p><center>UUA,UUG,CUU,CUC,CUA,CUG</center></p></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <p><center>UUA,UUG,CUU,CUC,CUA,CUG</center></p></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div> <p>The <del class="diffchange diffchange-inline">counterproductive </del>sticky ends:</p></div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div> <p>The <ins class="diffchange diffchange-inline">2012 Freiburg project's </ins>sticky ends:</p></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <p><center>(C)TGAC,GCTC,CTTG,GCTT,ACTG,CCTG,ACTC</center></p></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <p><center>(C)TGAC,GCTC,CTTG,GCTT,ACTG,CCTG,ACTC</center></p></div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div> <p>The <del class="diffchange diffchange-inline">useless </del>of Degeneracy has helped to design seven sticky ends. However, since the codons for identical amino acid are highly similar<del class="diffchange diffchange-inline">.</del></div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div> <p>The <ins class="diffchange diffchange-inline">feature </ins>of Degeneracy has helped to design seven sticky ends. However, since the codons for identical amino acid are highly similar<ins class="diffchange diffchange-inline">,</ins></div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div> <del class="diffchange diffchange-inline">This </del>feature, for experimental scientists, is a double-edged sword.</p></div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div> <ins class="diffchange diffchange-inline">this </ins>feature, for experimental scientists, is a double-edged sword.</p></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <h2>How to improve the Golden Gate sticky ends? A big Table!</h2></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <h2>How to improve the Golden Gate sticky ends? A big Table!</h2></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> </div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> </div></td></tr>
</table>HoPe Haohttp://2014.igem.org/wiki/index.php?title=Team:SJTU-BioX-Shanghai/Part3_TAL_Improvement&diff=356361&oldid=prevHoPe Hao at 21:30, 17 October 20142014-10-17T21:30:53Z<p></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><center><small><strong> Figure 1.4.4 DNA sequence of AA1 part</strong></small></center></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><center><small><strong> Figure 1.4.4 DNA sequence of AA1 part</strong></small></center></div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div> <p><small><i>The underlined <del class="diffchange diffchange-inline">parts are </del>recognized by BsmBI. Vertical bar(|) is the cutting position. As for this sample, TGAC is one sticky end which can combine with other <del class="diffchange diffchange-inline">seven </del>sticky ends.</i></small></p></div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div> <p><small><i>The underlined <ins class="diffchange diffchange-inline">sequence is </ins>recognized by BsmBI. Vertical bar(|) is the cutting position. As for this sample, TGAC is one sticky end which can combine with <ins class="diffchange diffchange-inline">six </ins>other sticky ends.</i></small></p></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <h3>Evaluate seven sticky ends designed by 2012 Freiburg</h3></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <h3>Evaluate seven sticky ends designed by 2012 Freiburg</h3></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <p>2012 Freiburg's parts have seven sticky ends:</p></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <p>2012 Freiburg's parts have seven sticky ends:</p></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <p><center>TGAC,GCTC,CTTG,GCTT,ACTG,CCTG,ACTC</center></p></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <p><center>TGAC,GCTC,CTTG,GCTT,ACTG,CCTG,ACTC</center></p></div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div> <p>We all know that certain two parts can combine together, under base<del class="diffchange diffchange-inline">-pair rule</del>. However, <del class="diffchange diffchange-inline">whether it </del>is possible that <del class="diffchange diffchange-inline">unpaired </del>sticky ends can bind together? <del class="diffchange diffchange-inline">In </del>fact, the more similar they are, the more possibility that can form new but <del class="diffchange diffchange-inline">error </del>base pairs.</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div> <p>We all know that certain two parts can combine together, under <ins class="diffchange diffchange-inline">the principle of complementary </ins>base <ins class="diffchange diffchange-inline">pairing</ins>. However, is <ins class="diffchange diffchange-inline">it </ins>possible that <ins class="diffchange diffchange-inline">not totally matched </ins>sticky ends can bind together? <ins class="diffchange diffchange-inline">We found, in </ins>fact, the more similar they are, the more possibility that <ins class="diffchange diffchange-inline">they </ins>can form new but <ins class="diffchange diffchange-inline">incorrect </ins>base pairs.</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div> <del class="diffchange diffchange-inline">Spired </del>by BLAST algorithm, we <del class="diffchange diffchange-inline">calculate </del>the similarity of <del class="diffchange diffchange-inline">each other </del>sticky ends. </p></div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div> <ins class="diffchange diffchange-inline">Inspired </ins>by BLAST algorithm, we <ins class="diffchange diffchange-inline">evaluated </ins>the similarity of <ins class="diffchange diffchange-inline">every pair of </ins>sticky ends. </p></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <center><img src="https://static.igem.org/mediawiki/2014/8/8e/TAL%E7%B2%98%E6%80%A7%E6%9C%AB%E7%AB%AF%E8%A1%A8%E6%A0%BC.png" width=400px></img></center></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <center><img src="https://static.igem.org/mediawiki/2014/8/8e/TAL%E7%B2%98%E6%80%A7%E6%9C%AB%E7%AB%AF%E8%A1%A8%E6%A0%BC.png" width=400px></img></center></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><center><small><strong> Figure 1.4.5 Strict rules score table </strong></small></center></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><center><small><strong> Figure 1.4.5 Strict rules score table </strong></small></center></div></td></tr>
</table>HoPe Haohttp://2014.igem.org/wiki/index.php?title=Team:SJTU-BioX-Shanghai/Part3_TAL_Improvement&diff=355658&oldid=prevHoPe Hao at 21:23, 17 October 20142014-10-17T21:23:11Z<p></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <h3>Previous Review: Freiburg’s way of connection</h3></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <h3>Previous Review: Freiburg’s way of connection</h3></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div> <p>The main <del class="diffchange diffchange-inline">principles </del>of connection is built upon the idea of Golden Gate Connection.( Sanjana, N. E. et al. A transcription activator-like effector toolbox for genome engineering. Nature Protocols 7, 171–192 (2012).)</p></div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div> <p>The main <ins class="diffchange diffchange-inline">principle </ins>of connection is built upon the idea of Golden Gate Connection.( Sanjana, N. E. et al. A transcription activator-like effector toolbox for genome engineering. Nature Protocols 7, 171–192 (2012).)</p></div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div> <p>The <del class="diffchange diffchange-inline">gust </del>of <del class="diffchange diffchange-inline">these </del>procedures is <del class="diffchange diffchange-inline">more related to one type of restriction enzyme, </del>type II Restriction Enzyme, <del class="diffchange diffchange-inline">especially </del>BsmBI enzyme.</p></div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div> <p>The <ins class="diffchange diffchange-inline">key point </ins>of <ins class="diffchange diffchange-inline">their </ins>procedures is <ins class="diffchange diffchange-inline">a </ins>type II Restriction Enzyme, BsmBI enzyme.</p></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <center><img src="https://static.igem.org/mediawiki/2014/5/5b/SJTU14_tal_improvment_2.png" ></img></center></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <center><img src="https://static.igem.org/mediawiki/2014/5/5b/SJTU14_tal_improvment_2.png" ></img></center></div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><center><small><strong> Figure 1.4.3 BsmBI recognition site</strong></small></center></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><center><small><strong> Figure 1.4.3 BsmBI recognition site</strong></small></center></div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div> <p>The main feature of this enzyme is the recognition sequence is on only one side of cleavage site. It provides the way which can be used to get certain <del class="diffchange diffchange-inline">incision without damaging </del>the whole sequence. The sticky end has 4bp <del class="diffchange diffchange-inline">base</del>, and it could be designed <del class="diffchange diffchange-inline">even </del>for combination of multiple sticky <del class="diffchange diffchange-inline">end</del>. That feature is <del class="diffchange diffchange-inline">fancy </del>at first, but we cannot <del class="diffchange diffchange-inline">regardless </del>its latent shortcomings. </p></div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div> <p>The main feature of this enzyme is <ins class="diffchange diffchange-inline">that </ins>the recognition sequence is on only one side of cleavage site. It provides the way which can be used to get certain <ins class="diffchange diffchange-inline">sticky ends with out breaking </ins>the whole sequence. The sticky end has 4bp, and it could be designed for combination of multiple sticky <ins class="diffchange diffchange-inline">ends</ins>. That feature is <ins class="diffchange diffchange-inline">excellent </ins>at first, but we cannot <ins class="diffchange diffchange-inline">ignore </ins>its latent shortcomings. </p></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <p>Let’s analyze the example (AA1) provided by Freiburg. </p></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <p>Let’s analyze the example (AA1) provided by Freiburg. </p></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <center><img src="https://static.igem.org/mediawiki/2014/b/bc/SJTU14_tal_improvment_3.png" width=700px></img></center></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <center><img src="https://static.igem.org/mediawiki/2014/b/bc/SJTU14_tal_improvment_3.png" width=700px></img></center></div></td></tr>
</table>HoPe Haohttp://2014.igem.org/wiki/index.php?title=Team:SJTU-BioX-Shanghai/Part3_TAL_Improvement&diff=355075&oldid=prevVisionve at 21:17, 17 October 20142014-10-17T21:17:01Z<p></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> The recognition sequence of the TALE protein:</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> The recognition sequence of the TALE protein:</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <center><font size="5" color="red">TCGATATCAAGC</font></center></p></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <center><font size="5" color="red">TCGATATCAAGC</font></center></p></div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div> <p>All parts are under artificial synthesis process, so there is few results, which can prove our changes are useful. However, with the principle of complementary base pairing, our <del class="diffchange diffchange-inline">chioce </del>should be better than original vision. And if you want our data or use our method to create your own best sticky ends, just contact us!</p></div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div> <p>All parts are under artificial synthesis process, so there is few results, which can prove our changes are useful. However, with the principle of complementary base pairing, our <ins class="diffchange diffchange-inline">choice </ins>should be better than original vision. And if you want our data or use our method to create your own best sticky ends, just contact us!</p></div></td></tr>
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</table>Visionvehttp://2014.igem.org/wiki/index.php?title=Team:SJTU-BioX-Shanghai/Part3_TAL_Improvement&diff=354880&oldid=prevHoPe Hao at 21:15, 17 October 20142014-10-17T21:15:07Z<p></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><center><small><strong> Figure 1.4.2 TALE amino acid sequence</strong></small></center></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><center><small><strong> Figure 1.4.2 TALE amino acid sequence</strong></small></center></div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div> <p>What XX means is that it <del class="diffchange diffchange-inline">determine </del>the certain kind of base. For one unit of repetition, other amino acids can be identical.</p></div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div> <p>What XX means is that it <ins class="diffchange diffchange-inline">determines </ins>the certain kind of base. For one unit of repetition, other amino acids can be identical.</p></div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div> <p>2. A fully functional TALE protein contains <del class="diffchange diffchange-inline">one </del>sequence<del class="diffchange diffchange-inline">, that does not have </del>repetitive units, recognizing base T<del class="diffchange diffchange-inline">, and </del>similar sequence but is only half length as <del class="diffchange diffchange-inline">its end. That is, one complete TALE protein is able to </del>recognize <del class="diffchange diffchange-inline">certain number of repetitive units and two bases</del>.</p></div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div> <p>2. A fully functional TALE protein contains <ins class="diffchange diffchange-inline">a segment of </ins>sequence <ins class="diffchange diffchange-inline">before </ins>repetitive units, recognizing <ins class="diffchange diffchange-inline">first </ins>base T<ins class="diffchange diffchange-inline">. It also contains a </ins>similar <ins class="diffchange diffchange-inline">segment </ins>sequence but <ins class="diffchange diffchange-inline">it </ins>is only half length as <ins class="diffchange diffchange-inline">the repetitive unit which can </ins>recognize <ins class="diffchange diffchange-inline">the last base T</ins>. </p></div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div> <p>3. The length that can be recognized is not strictly twelve or fourteen. According to the published results, the length <del class="diffchange diffchange-inline">and certain </del>sequence are dependent on number <del class="diffchange diffchange-inline">and type </del>of monomer.</p><br></div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div> <p>3. The length that can be recognized is not strictly twelve or fourteen. According to the published results, the length <ins class="diffchange diffchange-inline">of recognition </ins>sequence are dependent on <ins class="diffchange diffchange-inline">the </ins>number of monomer.</p><br></div></td></tr>
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<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div> <p>We can gather 96 bioparts based on Freiburg, and each part has its counterproductive base on certain location(1,2,3,4,5 or 6). By picking two bases on certain location, we are able to design one TALE protein sequence.</p></div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div> <p>We can gather 96 bioparts based on Freiburg, and each part has its counterproductive <ins class="diffchange diffchange-inline">sticky ends </ins>base on certain location(1,2,3,4,5 or 6). By picking <ins class="diffchange diffchange-inline">every </ins>two bases on certain location, we are able to design one TALE protein sequence.</p></div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <h3>Previous Review: Freiburg’s way of connection</h3></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <h3>Previous Review: Freiburg’s way of connection</h3></div></td></tr>
</table>HoPe Haohttp://2014.igem.org/wiki/index.php?title=Team:SJTU-BioX-Shanghai/Part3_TAL_Improvement&diff=353729&oldid=prevHoPe Hao at 21:01, 17 October 20142014-10-17T21:01:57Z<p></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <p>3. With the same ratio of plasmid and TALE direpeats, add the TALE direpeats one by one and connect them in 22 ℃, 30 minutes.</p></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <p>3. With the same ratio of plasmid and TALE direpeats, add the TALE direpeats one by one and connect them in 22 ℃, 30 minutes.</p></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <p>4. Every two parts connect at one time, and try to make three intermediates of 400bp. And then mix the plasmids to make the complete TALE.</p></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <p>4. Every two parts connect at one time, and try to make three intermediates of 400bp. And then mix the plasmids to make the complete TALE.</p></div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div> <p>5. The same ratio and connect following the program of 22℃ 2min, 40℃ <del class="diffchange diffchange-inline">30</del>,25 repeats.</p></div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div> <p>5. The same ratio and connect following the program of 22℃ 2min, 40℃ <ins class="diffchange diffchange-inline">30s</ins>, 25 repeats.</p></div></td></tr>
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<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div> <h3>The motivation to <del class="diffchange diffchange-inline">debug </del>2012 Freiburg’s parts</h3></div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div> <h3>The motivation to <ins class="diffchange diffchange-inline">improve </ins>2012 Freiburg’s parts</h3></div></td></tr>
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<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div> <p id="dianweidian4">Unfortunately, all of our attempts failed. We didn’t manage to make a complete TALE, or even make two of them together. However, what is important for us is that when we <del class="diffchange diffchange-inline">try </del>the 5th protocol, we <del class="diffchange diffchange-inline">notice </del>an unexpected result. When we <del class="diffchange diffchange-inline">analyze </del>the sequence result, we <del class="diffchange diffchange-inline">find </del>that our left adaptor, 1st part and right adaptor <del class="diffchange diffchange-inline">connect </del>together. Why <del class="diffchange diffchange-inline">do </del>we get this result? We <del class="diffchange diffchange-inline">notice </del>that their sticky <del class="diffchange diffchange-inline">end is </del>TGAC, GCTC, and ACTC. That is to say, GCTC and ACTC <del class="diffchange diffchange-inline">connect </del>with each other by mistake. In another word, if the sticky ends are very similar, they probably connect with each other. Although we failed again, the result <del class="diffchange diffchange-inline">gives </del>us confidence to improve 2012 Freiburg's parts. </p></div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div> <p id="dianweidian4">Unfortunately, all of our attempts failed. We didn’t manage to make a complete TALE, or even make two of them together. However, what is important for us is that when we <ins class="diffchange diffchange-inline">tried </ins>the 5th protocol, we <ins class="diffchange diffchange-inline">noticed </ins>an unexpected result. When we <ins class="diffchange diffchange-inline">analyzed </ins>the sequence result, we <ins class="diffchange diffchange-inline">found </ins>that our left adaptor, 1st part and right adaptor <ins class="diffchange diffchange-inline">connected </ins>together. Why <ins class="diffchange diffchange-inline">did </ins>we get this result? We <ins class="diffchange diffchange-inline">noticed </ins>that their sticky <ins class="diffchange diffchange-inline">ends are </ins>TGAC, GCTC, and ACTC. That is to say, GCTC and ACTC <ins class="diffchange diffchange-inline">might have connected </ins>with each other by mistake. In another word, if the sticky ends are very similar, they probably connect with each other. Although we failed again, the result <ins class="diffchange diffchange-inline">gave </ins>us confidence to improve 2012 Freiburg's parts. </p></div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <h2>How do we connect certain monomer? </h2></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div> <h2>How do we connect certain monomer? </h2></div></td></tr>
</table>HoPe Hao