http://openwetware.org/index.php?title=Reverse_Transcriptase&feed=atom&action=history 2013-05-23T20:59:25Z Revision history for this page on the wiki MediaWiki 1.13.2 http://openwetware.org/index.php?title=Reverse_Transcriptase&diff=118315&oldid=prev 2007-05-24T14:23:19Z

<p></p> <table style="background-color: white; color:black;"> <col class='diff-marker' /> <col class='diff-content' /> <col class='diff-marker' /> <col class='diff-content' /> <tr valign='top'> <td colspan='2' style="background-color: white; color:black;">←Older revision</td> <td colspan='2' style="background-color: white; color:black;">Revision as of 14:23, 24 May 2007</td> </tr> <tr><td colspan="2" class="diff-lineno">Line 58:</td> <td colspan="2" class="diff-lineno">Line 58:</td></tr> <tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>#Requires sequence specific primer to insure stable binding at elevated temperatures.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>#Requires sequence specific primer to insure stable binding at elevated temperatures.</div></td></tr> <tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>#Only capable of producing short products (~ 1-2 kb)</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>#Only capable of producing short products (~ 1-2 kb)</div></td></tr> <tr><td colspan="2">&nbsp;</td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins style="color: red; font-weight: bold; text-decoration: none;"></ins></div></td></tr> <tr><td colspan="2">&nbsp;</td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins style="color: red; font-weight: bold; text-decoration: none;">[[Category:Material]] [[Category:Enzyme]] [[Category:Polymerase]] [[Category:DNA]]</ins></div></td></tr> <!-- diff generator: internal 2013-05-23 20:59:25 --> </table>

Reshma P. Shetty http://openwetware.org/index.php?title=Reverse_Transcriptase&diff=11631&oldid=prev 2005-06-02T18:33:31Z

<p></p> <p><b>New page</b></p><div>==Characteristics of RT==<br /> #RNA dependent, DNA polymerase<br /> #Requires a suitable primer<br /> #*Sequence specific<br /> #*Random hexamers<br /> #*Poly-U (in Eukaryotes)<br /> #Two functional domains:<br /> #*DNA polymerase<br /> #*RNase H<br /> #Naturally occuring in RNA viruses:<br /> #*AMV = Avian Myeloblastosis Virus<br /> #*Mo-MLV = Moloney strain Murine Leukemia Virus<br /> #*HIV<br /> #*Most commercial RTs are either AMV or Mo-MLV derivatives<br /> <br /> ==AMV vs. Mo-MLV==<br /> ::{| border=&quot;1&quot; cellpadding=&quot;2&quot; cellspacing=&quot;0&quot;<br /> |-<br /> ! Mo-MLV || AMV<br /> |-<br /> | Single polypeptide chain || 2 polypeptide chains <br /> |-<br /> | 2 independently functioning, non-overlapping domains <br /> Amino terminal (450 aa) = DNA pol<br /> <br /> Carboxyl terminal (220 aa) = RNase H <br /> || RNase H and DNA polmerase activity associated with both chains<br /> |-<br /> | 37-42°C (up to 50°C) || 37-45°C (up to 55-60°C)<br /> |-<br /> | pH .6 || pH 8.3<br /> |-<br /> | Cloned in 1985 || Cloned in 1988<br /> |-<br /> | RNase H- available in 1987 || RNase H- available in 1998<br /> |}<br /> <br /> ==Problems associated with RT enzymes==<br /> <br /> #High Error Rate<br /> #Sluggish and inefficient (at best 50% efficient)<br /> #*RNase H degradation<br /> #**Competition for degradation vs. extension during initial primer binding<br /> #**Premature termination<br /> #*Prone to Pausing<br /> #**Sites rich in Secondary Structure<br /> #**Homopolymer runs (particularily A)<br /> #**Sensitive to [dNTP]<br /> <br /> <br /> ==Alternatives==<br /> Thermus thermophilus DNA polymerase (Tth pol)<br /> <br /> #Possesses RT activity in the presence of MnCl <br /> #*Synthesizes DNA from both RNA and DNA templates<br /> #*Mn+ lowers the fidelity of DNA synthesis during PCR<br /> #Thermostable<br /> #Requires sequence specific primer to insure stable binding at elevated temperatures.<br /> #Only capable of producing short products (~ 1-2 kb)</div>

Jason R. Kelly