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Team:KIT-Kyoto/Notebook/ATF1/september
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<h2>ATF1</h2> | <h2>ATF1</h2> | ||
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| + | <p class=MsoNormal><b><span lang=EN-US style='font-size:14.0pt'>September 1<sup>st</sup></span></b></p> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>Picked 32 colonies | ||
| + | of <i>ATF1 </i>transformants.</span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US> </span></p> | ||
| + | |||
| + | <p class=MsoNormal><b><span lang=EN-US style='font-size:14.0pt'>September 3<sup>rd</sup></span></b></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>Checked the | ||
| + | colonies by colony cracking.</span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>No appropriate | ||
| + | plasmid DNA was detected.</span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>Digested <i>ATF1</i> | ||
| + | and pET-15b with <i>Xho</i>I and <i>Bpu</i>1102I at 37</span><span | ||
| + | style='font-size:11.0pt;font-family:"ＭＳ 明朝","serif"'>˚</span><span lang=EN-US | ||
| + | style='font-size:11.0pt'>C overnight.</span></p> | ||
| + | |||
| + | <table class=MsoTableGrid border=1 cellspacing=0 cellpadding=0 | ||
| + | style='border-collapse:collapse;border:none'> | ||
| + | <tr> | ||
| + | <td width=111 valign=top style='width:83.2pt;border:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>ATF1</span></p> | ||
| + | </td> | ||
| + | <td width=55 valign=top style='width:41.2pt;border:solid windowtext 1.0pt; | ||
| + | border-left:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>24µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td width=111 valign=top style='width:83.2pt;border:solid windowtext 1.0pt; | ||
| + | border-top:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><i><span lang=EN-US style='font-size:11.0pt'>Xho</span></i><span | ||
| + | lang=EN-US style='font-size:11.0pt'>I</span></p> | ||
| + | </td> | ||
| + | <td width=55 valign=top style='width:41.2pt;border-top:none;border-left:none; | ||
| + | border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>1µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td width=111 valign=top style='width:83.2pt;border:solid windowtext 1.0pt; | ||
| + | border-top:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><i><span lang=EN-US style='font-size:11.0pt'>Bpu</span></i><span | ||
| + | lang=EN-US style='font-size:11.0pt'>1102I</span></p> | ||
| + | </td> | ||
| + | <td width=55 valign=top style='width:41.2pt;border-top:none;border-left:none; | ||
| + | border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>1µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td width=111 valign=top style='width:83.2pt;border:solid windowtext 1.0pt; | ||
| + | border-top:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>BSA</span></p> | ||
| + | </td> | ||
| + | <td width=55 valign=top style='width:41.2pt;border-top:none;border-left:none; | ||
| + | border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>1µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td width=111 valign=top style='width:83.2pt;border:solid windowtext 1.0pt; | ||
| + | border-top:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>NEB Buffer 2</span></p> | ||
| + | </td> | ||
| + | <td width=55 valign=top style='width:41.2pt;border-top:none;border-left:none; | ||
| + | border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>3µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | </table> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'> </span></p> | ||
| + | |||
| + | <table class=MsoTableGrid border=1 cellspacing=0 cellpadding=0 | ||
| + | style='border-collapse:collapse;border:none'> | ||
| + | <tr> | ||
| + | <td width=111 valign=top style='width:83.2pt;border:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>pET-15b</span></p> | ||
| + | </td> | ||
| + | <td width=55 valign=top style='width:41.2pt;border:solid windowtext 1.0pt; | ||
| + | border-left:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>33µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | <tr style='height:20.15pt'> | ||
| + | <td width=111 valign=top style='width:83.2pt;border:solid windowtext 1.0pt; | ||
| + | border-top:none;padding:0mm 5.4pt 0mm 5.4pt;height:20.15pt'> | ||
| + | <p class=MsoNormal><i><span lang=EN-US style='font-size:11.0pt'>Xho</span></i><span | ||
| + | lang=EN-US style='font-size:11.0pt'>I</span></p> | ||
| + | </td> | ||
| + | <td width=55 valign=top style='width:41.2pt;border-top:none;border-left:none; | ||
| + | border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt;height:20.15pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>1µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td width=111 valign=top style='width:83.2pt;border:solid windowtext 1.0pt; | ||
| + | border-top:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><i><span lang=EN-US style='font-size:11.0pt'>Bpu</span></i><span | ||
| + | lang=EN-US style='font-size:11.0pt'>1102I</span></p> | ||
| + | </td> | ||
| + | <td width=55 valign=top style='width:41.2pt;border-top:none;border-left:none; | ||
| + | border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>1µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td width=111 valign=top style='width:83.2pt;border:solid windowtext 1.0pt; | ||
| + | border-top:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>BSA</span></p> | ||
| + | </td> | ||
| + | <td width=55 valign=top style='width:41.2pt;border-top:none;border-left:none; | ||
| + | border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>1µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td width=111 valign=top style='width:83.2pt;border:solid windowtext 1.0pt; | ||
| + | border-top:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>NEB Buffer 2</span></p> | ||
| + | </td> | ||
| + | <td width=55 valign=top style='width:41.2pt;border-top:none;border-left:none; | ||
| + | border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>4µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | </table> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US> </span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US> </span></p> | ||
| + | |||
| + | <p class=MsoNormal><b><span lang=EN-US style='font-size:14.0pt'>September 4<sup>th </sup></span></b></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>Applied pET-15b | ||
| + | and <i>ATF1</i> to the blue gel electrophoresis.</span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>No pET-15b was | ||
| + | detected.</span></p> | ||
| + | |||
| + | <p class=MsoNormal><i><span lang=EN-US style='font-size:11.0pt'>ATF1</span></i><span | ||
| + | lang=EN-US style='font-size:11.0pt'> was extracted from the blue gel.</span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US> </span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US> </span></p> | ||
| + | |||
| + | <p class=MsoNormal><b><span lang=EN-US style='font-size:14.0pt'>September 11<sup>th</sup></span></b></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>PCR reaction was | ||
| + | carried out using the primer. </span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>(Forward : CGATCCTCGAGTGGCCTGGTATTGTCATCGCGTTG</span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>Reverse : CGATCGCTCAGCAACCAACCAAAGCCGAGGGAGTG)</span></p> | ||
| + | |||
| + | <table class=MsoTableGrid border=1 cellspacing=0 cellpadding=0 | ||
| + | style='border-collapse:collapse;border:none'> | ||
| + | <tr> | ||
| + | <td width=162 valign=top style='width:121.45pt;border:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>Buffer</span></p> | ||
| + | </td> | ||
| + | <td width=162 valign=top style='width:121.45pt;border:solid windowtext 1.0pt; | ||
| + | border-left:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>50µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td width=162 valign=top style='width:121.45pt;border:solid windowtext 1.0pt; | ||
| + | border-top:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>dNTP</span></p> | ||
| + | </td> | ||
| + | <td width=162 valign=top style='width:121.45pt;border-top:none;border-left: | ||
| + | none;border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>20µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td width=162 valign=top style='width:121.45pt;border:solid windowtext 1.0pt; | ||
| + | border-top:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>Primer mix</span></p> | ||
| + | </td> | ||
| + | <td width=162 valign=top style='width:121.45pt;border-top:none;border-left: | ||
| + | none;border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>1µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td width=162 valign=top style='width:121.45pt;border:solid windowtext 1.0pt; | ||
| + | border-top:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>Genome DNA</span></p> | ||
| + | </td> | ||
| + | <td width=162 valign=top style='width:121.45pt;border-top:none;border-left: | ||
| + | none;border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>0.5µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td width=162 valign=top style='width:121.45pt;border:solid windowtext 1.0pt; | ||
| + | border-top:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>KOD-FX</span></p> | ||
| + | </td> | ||
| + | <td width=162 valign=top style='width:121.45pt;border-top:none;border-left: | ||
| + | none;border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>2µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td width=162 valign=top style='width:121.45pt;border:solid windowtext 1.0pt; | ||
| + | border-top:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>H<sub>2</sub>O</span></p> | ||
| + | </td> | ||
| + | <td width=162 valign=top style='width:121.45pt;border-top:none;border-left: | ||
| + | none;border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>26.5µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | </table> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>Electrophoresed in | ||
| + | 1% agarose gel. Verified as <i>ATF1.</i></span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>Purified the PCR | ||
| + | products of <i>ATF1.</i></span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>Digested <i>ATF1</i> | ||
| + | and pET-15b with <i>Xho</i>I and <i>Bpu</i>1102I at 37</span><span | ||
| + | style='font-size:11.0pt;font-family:"ＭＳ 明朝","serif"'>˚</span><span lang=EN-US | ||
| + | style='font-size:11.0pt'>C for 2 hours.</span></p> | ||
| + | |||
| + | <p class=MsoNormal><i><span lang=EN-US style='font-size:11.0pt'> </span></i></p> | ||
| + | |||
| + | <table class=MsoTableGrid border=1 cellspacing=0 cellpadding=0 | ||
| + | style='border-collapse:collapse;border:none'> | ||
| + | <tr> | ||
| + | <td width=111 valign=top style='width:83.2pt;border:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><i><span lang=EN-US style='font-size:11.0pt'>ATF1</span></i></p> | ||
| + | </td> | ||
| + | <td width=55 valign=top style='width:41.2pt;border:solid windowtext 1.0pt; | ||
| + | border-left:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>15.5µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td width=111 valign=top style='width:83.2pt;border:solid windowtext 1.0pt; | ||
| + | border-top:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><i><span lang=EN-US style='font-size:11.0pt'>Xho</span></i><span | ||
| + | lang=EN-US style='font-size:11.0pt'>I</span></p> | ||
| + | </td> | ||
| + | <td width=55 valign=top style='width:41.2pt;border-top:none;border-left:none; | ||
| + | border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>1µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td width=111 valign=top style='width:83.2pt;border:solid windowtext 1.0pt; | ||
| + | border-top:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><i><span lang=EN-US style='font-size:11.0pt'>Bpu</span></i><span | ||
| + | lang=EN-US style='font-size:11.0pt'>1102I</span></p> | ||
| + | </td> | ||
| + | <td width=55 valign=top style='width:41.2pt;border-top:none;border-left:none; | ||
| + | border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>1µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td width=111 valign=top style='width:83.2pt;border:solid windowtext 1.0pt; | ||
| + | border-top:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>BSA</span></p> | ||
| + | </td> | ||
| + | <td width=55 valign=top style='width:41.2pt;border-top:none;border-left:none; | ||
| + | border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>0.5µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td width=111 valign=top style='width:83.2pt;border:solid windowtext 1.0pt; | ||
| + | border-top:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>NEB Buffer 2</span></p> | ||
| + | </td> | ||
| + | <td width=55 valign=top style='width:41.2pt;border-top:none;border-left:none; | ||
| + | border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>2µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | </table> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>Added 1µL of BAP | ||
| + | to the solution of pET-15b and incubated it at 37</span><span style='font-size: | ||
| + | 11.0pt;font-family:"ＭＳ 明朝","serif"'>˚</span><span lang=EN-US style='font-size: | ||
| + | 11.0pt'>C for 30 minutes.</span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>Applied pET-15b | ||
| + | and <i>ATF1</i> to the blue gel electrophoresis.</span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>Isolated and | ||
| + | purified them.</span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>Ligated pET-15b and | ||
| + | <i>ATF1 </i>and transformed.</span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US> </span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US> </span></p> | ||
| + | |||
| + | <p class=MsoNormal><b><span lang=EN-US style='font-size:14.0pt'>September 12th</span></b></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>Picked 32 colonies | ||
| + | of <i>ATF1 </i>transformants.</span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US> </span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US> </span></p> | ||
| + | |||
| + | <p class=MsoNormal><b><span lang=EN-US style='font-size:14.0pt'>September 13th</span></b></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>Checked the | ||
| + | colonies by colony cracking.</span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>Picked up the | ||
| + | appropriate colonies and cultured in 3mL LB medium with ampicillin at 37</span><span | ||
| + | style='font-size:11.0pt;font-family:"ＭＳ 明朝","serif"'>˚</span><span lang=EN-US | ||
| + | style='font-size:11.0pt'>C for 5 hours.</span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>Miniprepped plasmid | ||
| + | DNA.</span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>Digested plasmid | ||
| + | DNA with <i>Xho</i>I and Bpu1102I at 37</span><span style='font-size:11.0pt; | ||
| + | font-family:"ＭＳ 明朝","serif"'>˚</span><span lang=EN-US style='font-size:11.0pt'>C | ||
| + | for 2 hours.</span></p> | ||
| + | |||
| + | <table class=MsoTableGrid border=1 cellspacing=0 cellpadding=0 | ||
| + | style='border-collapse:collapse;border:none'> | ||
| + | <tr> | ||
| + | <td width=150 valign=top style='width:112.25pt;border:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><i><span lang=EN-US style='font-size:11.0pt'>ATF1 </span></i><span | ||
| + | lang=EN-US style='font-size:11.0pt'>into pET-15b</span></p> | ||
| + | </td> | ||
| + | <td width=55 valign=top style='width:41.2pt;border:solid windowtext 1.0pt; | ||
| + | border-left:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>15.5µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td width=150 valign=top style='width:112.25pt;border:solid windowtext 1.0pt; | ||
| + | border-top:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><i><span lang=EN-US style='font-size:11.0pt'>Xho</span></i><span | ||
| + | lang=EN-US style='font-size:11.0pt'>I</span></p> | ||
| + | </td> | ||
| + | <td width=55 valign=top style='width:41.2pt;border-top:none;border-left:none; | ||
| + | border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>1µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td width=150 valign=top style='width:112.25pt;border:solid windowtext 1.0pt; | ||
| + | border-top:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><i><span lang=EN-US style='font-size:11.0pt'>Bpu</span></i><span | ||
| + | lang=EN-US style='font-size:11.0pt'>1102I</span></p> | ||
| + | </td> | ||
| + | <td width=55 valign=top style='width:41.2pt;border-top:none;border-left:none; | ||
| + | border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>1µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td width=150 valign=top style='width:112.25pt;border:solid windowtext 1.0pt; | ||
| + | border-top:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>BSA</span></p> | ||
| + | </td> | ||
| + | <td width=55 valign=top style='width:41.2pt;border-top:none;border-left:none; | ||
| + | border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>0.5µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td width=150 valign=top style='width:112.25pt;border:solid windowtext 1.0pt; | ||
| + | border-top:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>NEB Buffer 2</span></p> | ||
| + | </td> | ||
| + | <td width=55 valign=top style='width:41.2pt;border-top:none;border-left:none; | ||
| + | border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>2µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | </table> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>Applied pET-15b to | ||
| + | the agarose gel electrophoresis.</span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>No band was | ||
| + | detected.</span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>Plasmid DNA was | ||
| + | digested with <i>Xho</i>I and <i>Bpu</i>1102I at 37</span><span | ||
| + | style='font-size:11.0pt;font-family:"ＭＳ 明朝","serif"'>˚</span><span lang=EN-US | ||
| + | style='font-size:11.0pt'>C (overnight).</span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'> </span></p> | ||
| + | |||
| + | <table class=MsoTableGrid border=1 cellspacing=0 cellpadding=0 | ||
| + | style='border-collapse:collapse;border:none'> | ||
| + | <tr> | ||
| + | <td width=150 valign=top style='width:112.25pt;border:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><i><span lang=EN-US style='font-size:11.0pt'>ATF1 </span></i><span | ||
| + | lang=EN-US style='font-size:11.0pt'>into pET-15b</span></p> | ||
| + | </td> | ||
| + | <td width=55 valign=top style='width:41.2pt;border:solid windowtext 1.0pt; | ||
| + | border-left:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>15.5µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td width=150 valign=top style='width:112.25pt;border:solid windowtext 1.0pt; | ||
| + | border-top:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><i><span lang=EN-US style='font-size:11.0pt'>Xho</span></i><span | ||
| + | lang=EN-US style='font-size:11.0pt'>I</span></p> | ||
| + | </td> | ||
| + | <td width=55 valign=top style='width:41.2pt;border-top:none;border-left:none; | ||
| + | border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>1µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td width=150 valign=top style='width:112.25pt;border:solid windowtext 1.0pt; | ||
| + | border-top:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><i><span lang=EN-US style='font-size:11.0pt'>Bpu</span></i><span | ||
| + | lang=EN-US style='font-size:11.0pt'>1102I</span></p> | ||
| + | </td> | ||
| + | <td width=55 valign=top style='width:41.2pt;border-top:none;border-left:none; | ||
| + | border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>1µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td width=150 valign=top style='width:112.25pt;border:solid windowtext 1.0pt; | ||
| + | border-top:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>BSA</span></p> | ||
| + | </td> | ||
| + | <td width=55 valign=top style='width:41.2pt;border-top:none;border-left:none; | ||
| + | border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>0.5µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td width=150 valign=top style='width:112.25pt;border:solid windowtext 1.0pt; | ||
| + | border-top:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>NEB Buffer 2</span></p> | ||
| + | </td> | ||
| + | <td width=55 valign=top style='width:41.2pt;border-top:none;border-left:none; | ||
| + | border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>2µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | </table> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'> </span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US> </span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US> </span></p> | ||
| + | |||
| + | <p class=MsoNormal><b><span lang=EN-US style='font-size:14.0pt'>September 14<sup>th</sup></span></b></p> | ||
| + | |||
| + | <p class=MsoNormal><i><span lang=EN-US style='font-size:11.0pt'>ATF1</span></i><span | ||
| + | lang=EN-US style='font-size:11.0pt'> and pET-15b were applied<i> </i>to | ||
| + | electrophoresis to the agarose gel.</span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>No band was | ||
| + | detected.</span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>Digested <i>ATF1</i> | ||
| + | and pET-15b with <i>Bpu</i>1102I at 37</span><span style='font-size:11.0pt; | ||
| + | font-family:"ＭＳ 明朝","serif"'>˚</span><span lang=EN-US style='font-size:11.0pt'>C | ||
| + | overnight.</span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'> </span></p> | ||
| + | |||
| + | <table class=MsoTableGrid border=1 cellspacing=0 cellpadding=0 | ||
| + | style='border-collapse:collapse;border:none'> | ||
| + | <tr> | ||
| + | <td width=98 valign=top style='width:73.65pt;border:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><i><span lang=EN-US style='font-size:11.0pt'>ATF1</span></i></p> | ||
| + | </td> | ||
| + | <td width=55 valign=top style='width:41.2pt;border:solid windowtext 1.0pt; | ||
| + | border-left:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>26µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td width=98 valign=top style='width:73.65pt;border:solid windowtext 1.0pt; | ||
| + | border-top:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><i><span lang=EN-US style='font-size:11.0pt'>Bpu</span></i><span | ||
| + | lang=EN-US style='font-size:11.0pt'>1102I</span></p> | ||
| + | </td> | ||
| + | <td width=55 valign=top style='width:41.2pt;border-top:none;border-left:none; | ||
| + | border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>1µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td width=98 valign=top style='width:73.65pt;border:solid windowtext 1.0pt; | ||
| + | border-top:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><i><span lang=EN-US style='font-size:11.0pt'>Bpu</span></i><span | ||
| + | lang=EN-US style='font-size:11.0pt'>1102I Buffer</span></p> | ||
| + | </td> | ||
| + | <td width=55 valign=top style='width:41.2pt;border-top:none;border-left:none; | ||
| + | border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>3µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | </table> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'> </span></p> | ||
| + | |||
| + | <table class=MsoTableGrid border=1 cellspacing=0 cellpadding=0 | ||
| + | style='border-collapse:collapse;border:none'> | ||
| + | <tr> | ||
| + | <td width=98 valign=top style='width:73.65pt;border:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>pET-15b</span></p> | ||
| + | </td> | ||
| + | <td width=55 valign=top style='width:41.2pt;border:solid windowtext 1.0pt; | ||
| + | border-left:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>26µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td width=98 valign=top style='width:73.65pt;border:solid windowtext 1.0pt; | ||
| + | border-top:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><i><span lang=EN-US style='font-size:11.0pt'>Bpu</span></i><span | ||
| + | lang=EN-US style='font-size:11.0pt'>1102I</span></p> | ||
| + | </td> | ||
| + | <td width=55 valign=top style='width:41.2pt;border-top:none;border-left:none; | ||
| + | border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>1µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td width=98 valign=top style='width:73.65pt;border:solid windowtext 1.0pt; | ||
| + | border-top:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><i><span lang=EN-US style='font-size:11.0pt'>Bpu</span></i><span | ||
| + | lang=EN-US style='font-size:11.0pt'>1102I Buffer</span></p> | ||
| + | </td> | ||
| + | <td width=55 valign=top style='width:41.2pt;border-top:none;border-left:none; | ||
| + | border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>3µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | </table> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'> </span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US> </span></p> | ||
| + | |||
| + | <p class=MsoNormal><b><span lang=EN-US style='font-size:14.0pt'>September 15<sup>th</sup></span></b></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>Purified <i>ATF1</i> | ||
| + | and pET-15b.</span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>Digested <i>ATF1</i> | ||
| + | and pET-15b with <i>Xho</i>I.</span></p> | ||
| + | |||
| + | <table class=MsoTableGrid border=1 cellspacing=0 cellpadding=0 | ||
| + | style='border-collapse:collapse;border:none'> | ||
| + | <tr> | ||
| + | <td width=98 valign=top style='width:73.65pt;border:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><i><span lang=EN-US style='font-size:11.0pt'>ATF1</span></i></p> | ||
| + | </td> | ||
| + | <td width=55 valign=top style='width:41.2pt;border:solid windowtext 1.0pt; | ||
| + | border-left:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>26µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td width=98 valign=top style='width:73.65pt;border:solid windowtext 1.0pt; | ||
| + | border-top:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><i><span lang=EN-US style='font-size:11.0pt'>Xho</span></i><span | ||
| + | lang=EN-US style='font-size:11.0pt'>I</span></p> | ||
| + | </td> | ||
| + | <td width=55 valign=top style='width:41.2pt;border-top:none;border-left:none; | ||
| + | border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>1µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td width=98 valign=top style='width:73.65pt;border:solid windowtext 1.0pt; | ||
| + | border-top:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><i><span lang=EN-US style='font-size:11.0pt'>Bpu</span></i><span | ||
| + | lang=EN-US style='font-size:11.0pt'>1102I Buffer</span></p> | ||
| + | </td> | ||
| + | <td width=55 valign=top style='width:41.2pt;border-top:none;border-left:none; | ||
| + | border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>3µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | </table> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'> </span></p> | ||
| + | |||
| + | <table class=MsoTableGrid border=1 cellspacing=0 cellpadding=0 | ||
| + | style='border-collapse:collapse;border:none'> | ||
| + | <tr> | ||
| + | <td width=98 valign=top style='width:73.65pt;border:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>pET-15b</span></p> | ||
| + | </td> | ||
| + | <td width=55 valign=top style='width:41.2pt;border:solid windowtext 1.0pt; | ||
| + | border-left:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>26µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td width=98 valign=top style='width:73.65pt;border:solid windowtext 1.0pt; | ||
| + | border-top:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><i><span lang=EN-US style='font-size:11.0pt'>Xho</span></i><span | ||
| + | lang=EN-US style='font-size:11.0pt'>I</span></p> | ||
| + | </td> | ||
| + | <td width=55 valign=top style='width:41.2pt;border-top:none;border-left:none; | ||
| + | border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>1µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td width=98 valign=top style='width:73.65pt;border:solid windowtext 1.0pt; | ||
| + | border-top:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><i><span lang=EN-US style='font-size:11.0pt'>Bpu</span></i><span | ||
| + | lang=EN-US style='font-size:11.0pt'>1102I Buffer</span></p> | ||
| + | </td> | ||
| + | <td width=55 valign=top style='width:41.2pt;border-top:none;border-left:none; | ||
| + | border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>3µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | </table> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'> </span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>pET-15b and <i>ATF1</i> | ||
| + | were applied to electrophoresis in the Blue gel and purified.</span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>Ligated <i>ATF1</i> | ||
| + | and pET-15b and transformed them into <i>E. coli </i>cells.</span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US> </span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US> </span></p> | ||
| + | |||
| + | <p class=MsoNormal><b><span lang=EN-US style='font-size:14.0pt'>September 16th</span></b></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>Picked 32 colonies | ||
| + | of <i>ATF1 </i>transformants.</span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US> </span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US> </span></p> | ||
| + | |||
| + | <p class=MsoNormal><b><span lang=EN-US style='font-size:14.0pt'>September 18th</span></b></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>Checked the | ||
| + | colonies by colony cracking.</span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>Picked up the | ||
| + | colonies and cultured in 3mL LB medium with ampicillin at 37</span><span | ||
| + | style='font-size:11.0pt;font-family:"ＭＳ 明朝","serif"'>˚</span><span lang=EN-US | ||
| + | style='font-size:11.0pt'>C (overnight).</span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'> </span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US> </span></p> | ||
| + | |||
| + | <p class=MsoNormal><b><span lang=EN-US style='font-size:14.0pt'>September 19<sup>th</sup></span></b></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>plasmid DNA was | ||
| + | purified and digested with <i>Xho</i>I and<i> Bpu</i>1102I at 37</span><span | ||
| + | style='font-size:11.0pt;font-family:"ＭＳ 明朝","serif"'>˚</span><span lang=EN-US | ||
| + | style='font-size:11.0pt'>C for 2 hours.</span></p> | ||
| + | |||
| + | <table class=MsoTableGrid border=1 cellspacing=0 cellpadding=0 | ||
| + | style='border-collapse:collapse;border:none'> | ||
| + | <tr> | ||
| + | <td width=150 valign=top style='width:112.25pt;border:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><i><span lang=EN-US style='font-size:11.0pt'>ATF1 </span></i><span | ||
| + | lang=EN-US style='font-size:11.0pt'>into pET-15b</span></p> | ||
| + | </td> | ||
| + | <td width=55 valign=top style='width:41.2pt;border:solid windowtext 1.0pt; | ||
| + | border-left:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>15.5µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td width=150 valign=top style='width:112.25pt;border:solid windowtext 1.0pt; | ||
| + | border-top:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><i><span lang=EN-US style='font-size:11.0pt'>Xho</span></i><span | ||
| + | lang=EN-US style='font-size:11.0pt'>I</span></p> | ||
| + | </td> | ||
| + | <td width=55 valign=top style='width:41.2pt;border-top:none;border-left:none; | ||
| + | border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>1µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td width=150 valign=top style='width:112.25pt;border:solid windowtext 1.0pt; | ||
| + | border-top:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><i><span lang=EN-US style='font-size:11.0pt'>Bpu</span></i><span | ||
| + | lang=EN-US style='font-size:11.0pt'>1102I</span></p> | ||
| + | </td> | ||
| + | <td width=55 valign=top style='width:41.2pt;border-top:none;border-left:none; | ||
| + | border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>1µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td width=150 valign=top style='width:112.25pt;border:solid windowtext 1.0pt; | ||
| + | border-top:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>BSA</span></p> | ||
| + | </td> | ||
| + | <td width=55 valign=top style='width:41.2pt;border-top:none;border-left:none; | ||
| + | border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>0.5µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td width=150 valign=top style='width:112.25pt;border:solid windowtext 1.0pt; | ||
| + | border-top:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>NEB Buffer 2</span></p> | ||
| + | </td> | ||
| + | <td width=55 valign=top style='width:41.2pt;border-top:none;border-left:none; | ||
| + | border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>2µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | </table> | ||
| + | |||
| + | <p class=MsoNormal><i><span lang=EN-US style='font-size:11.0pt'>ATF1</span></i><span | ||
| + | lang=EN-US style='font-size:11.0pt'> and pET-15b were applied<i> </i>to | ||
| + | electrophoresis to the agarose gel.</span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>No band was | ||
| + | detected.</span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>Digested <i>ATF1</i> | ||
| + | and pET-15b with <i>Xho</i>I and <i>Blp</i>I.</span></p> | ||
| + | |||
| + | <table class=MsoTableGrid border=1 cellspacing=0 cellpadding=0 | ||
| + | style='border-collapse:collapse;border:none'> | ||
| + | <tr> | ||
| + | <td width=150 valign=top style='width:112.25pt;border:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><i><span lang=EN-US style='font-size:11.0pt'>ATF1</span></i></p> | ||
| + | </td> | ||
| + | <td width=55 valign=top style='width:41.2pt;border:solid windowtext 1.0pt; | ||
| + | border-left:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>15.5µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td width=150 valign=top style='width:112.25pt;border:solid windowtext 1.0pt; | ||
| + | border-top:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><i><span lang=EN-US style='font-size:11.0pt'>Xho</span></i><span | ||
| + | lang=EN-US style='font-size:11.0pt'>I</span></p> | ||
| + | </td> | ||
| + | <td width=55 valign=top style='width:41.2pt;border-top:none;border-left:none; | ||
| + | border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>1µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td width=150 valign=top style='width:112.25pt;border:solid windowtext 1.0pt; | ||
| + | border-top:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><i><span lang=EN-US style='font-size:11.0pt'>Blp</span></i><span | ||
| + | lang=EN-US style='font-size:11.0pt'>I</span></p> | ||
| + | </td> | ||
| + | <td width=55 valign=top style='width:41.2pt;border-top:none;border-left:none; | ||
| + | border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>1µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td width=150 valign=top style='width:112.25pt;border:solid windowtext 1.0pt; | ||
| + | border-top:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>BSA</span></p> | ||
| + | </td> | ||
| + | <td width=55 valign=top style='width:41.2pt;border-top:none;border-left:none; | ||
| + | border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>0.5µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td width=150 valign=top style='width:112.25pt;border:solid windowtext 1.0pt; | ||
| + | border-top:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>Buffer</span></p> | ||
| + | </td> | ||
| + | <td width=55 valign=top style='width:41.2pt;border-top:none;border-left:none; | ||
| + | border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>2µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | </table> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'> </span></p> | ||
| + | |||
| + | <table class=MsoTableGrid border=1 cellspacing=0 cellpadding=0 | ||
| + | style='border-collapse:collapse;border:none'> | ||
| + | <tr> | ||
| + | <td width=150 valign=top style='width:112.25pt;border:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>pET-15b</span></p> | ||
| + | </td> | ||
| + | <td width=55 valign=top style='width:41.2pt;border:solid windowtext 1.0pt; | ||
| + | border-left:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>15.5µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td width=150 valign=top style='width:112.25pt;border:solid windowtext 1.0pt; | ||
| + | border-top:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><i><span lang=EN-US style='font-size:11.0pt'>Xho</span></i><span | ||
| + | lang=EN-US style='font-size:11.0pt'>I</span></p> | ||
| + | </td> | ||
| + | <td width=55 valign=top style='width:41.2pt;border-top:none;border-left:none; | ||
| + | border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>1µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td width=150 valign=top style='width:112.25pt;border:solid windowtext 1.0pt; | ||
| + | border-top:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><i><span lang=EN-US style='font-size:11.0pt'>Blp</span></i><span | ||
| + | lang=EN-US style='font-size:11.0pt'>I</span></p> | ||
| + | </td> | ||
| + | <td width=55 valign=top style='width:41.2pt;border-top:none;border-left:none; | ||
| + | border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>1µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td width=150 valign=top style='width:112.25pt;border:solid windowtext 1.0pt; | ||
| + | border-top:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>BSA</span></p> | ||
| + | </td> | ||
| + | <td width=55 valign=top style='width:41.2pt;border-top:none;border-left:none; | ||
| + | border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>0.5µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td width=150 valign=top style='width:112.25pt;border:solid windowtext 1.0pt; | ||
| + | border-top:none;padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>Buffer</span></p> | ||
| + | </td> | ||
| + | <td width=55 valign=top style='width:41.2pt;border-top:none;border-left:none; | ||
| + | border-bottom:solid windowtext 1.0pt;border-right:solid windowtext 1.0pt; | ||
| + | padding:0mm 5.4pt 0mm 5.4pt'> | ||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>2µL</span></p> | ||
| + | </td> | ||
| + | </tr> | ||
| + | </table> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'> </span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>Added 1µL of BAP | ||
| + | to the solution of pET-15b and incubated it at 37</span><span style='font-size: | ||
| + | 11.0pt;font-family:"ＭＳ 明朝","serif"'>˚</span><span lang=EN-US style='font-size: | ||
| + | 11.0pt'>C for 30 minutes.</span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>Applied pET-15b | ||
| + | and <i>ATF1</i> to the blue gel electrophoresis.</span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>Isolated and | ||
| + | purified them.</span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>The <i>ATF1 </i>was | ||
| + | ligated with pET-15b, and transformed <i>into</i> <i>E. coli</i> cells.</span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'> </span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='color:red'> </span></p> | ||
| + | |||
| + | <p class=MsoNormal><b><span lang=EN-US style='font-size:14.0pt'>September 20<sup>th</sup></span></b></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>Picked 64 colonies | ||
| + | of <i>ATF1 </i>transformants.</span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US> </span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US> </span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US> </span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US> </span></p> | ||
| + | |||
| + | <p class=MsoNormal><b><span lang=EN-US style='font-size:14.0pt'>September 21st</span></b></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>Checked the | ||
| + | colonies by colony cracking.</span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>Picked up colonies | ||
| + | and cultured in the LB in 3mL LB medium with ampicillin at 37</span><span | ||
| + | style='font-size:11.0pt;font-family:"ＭＳ 明朝","serif"'>˚</span><span lang=EN-US | ||
| + | style='font-size:11.0pt'>C (overnight).</span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US> </span></p> | ||
| + | |||
| + | <p class=MsoNormal><b><span lang=EN-US style='font-size:14.0pt'>September 22nd</span></b></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>Miniprepped plasmid | ||
| + | DNA and digested it with <i>Xho</i>I and<i> Blp</i>I at 37</span><span | ||
| + | style='font-size:11.0pt;font-family:"ＭＳ 明朝","serif"'>˚</span><span lang=EN-US | ||
| + | style='font-size:11.0pt'>C for 2 hours.</span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>Applied<i> ATF1</i> | ||
| + | into pET-15b to the agarose gel electrophoresis.</span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'>No band was | ||
| + | detected.</span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'> </span></p> | ||
| + | |||
| + | <p class=MsoNormal><span lang=EN-US style='font-size:11.0pt'> </span></p> | ||
</div> | </div> | ||
Revision as of 07:30, 27 September 2013
ATF1
September 1st
Picked 32 colonies of ATF1 transformants.
September 3rd
Checked the colonies by colony cracking.
No appropriate plasmid DNA was detected.
Digested ATF1 and pET-15b with XhoI and Bpu1102I at 37˚C overnight.
|
ATF1 |
24µL |
|
XhoI |
1µL |
|
Bpu1102I |
1µL |
|
BSA |
1µL |
|
NEB Buffer 2 |
3µL |
|
pET-15b |
33µL |
|
XhoI |
1µL |
|
Bpu1102I |
1µL |
|
BSA |
1µL |
|
NEB Buffer 2 |
4µL |
September 4th
Applied pET-15b and ATF1 to the blue gel electrophoresis.
No pET-15b was detected.
ATF1 was extracted from the blue gel.
September 11th
PCR reaction was carried out using the primer.
(Forward : CGATCCTCGAGTGGCCTGGTATTGTCATCGCGTTG
Reverse : CGATCGCTCAGCAACCAACCAAAGCCGAGGGAGTG)
|
Buffer |
50µL |
|
dNTP |
20µL |
|
Primer mix |
1µL |
|
Genome DNA |
0.5µL |
|
KOD-FX |
2µL |
|
H2O |
26.5µL |
Electrophoresed in 1% agarose gel. Verified as ATF1.
Purified the PCR products of ATF1.
Digested ATF1 and pET-15b with XhoI and Bpu1102I at 37˚C for 2 hours.
|
ATF1 |
15.5µL |
|
XhoI |
1µL |
|
Bpu1102I |
1µL |
|
BSA |
0.5µL |
|
NEB Buffer 2 |
2µL |
Added 1µL of BAP to the solution of pET-15b and incubated it at 37˚C for 30 minutes.
Applied pET-15b and ATF1 to the blue gel electrophoresis.
Isolated and purified them.
Ligated pET-15b and ATF1 and transformed.
September 12th
Picked 32 colonies of ATF1 transformants.
September 13th
Checked the colonies by colony cracking.
Picked up the appropriate colonies and cultured in 3mL LB medium with ampicillin at 37˚C for 5 hours.
Miniprepped plasmid DNA.
Digested plasmid DNA with XhoI and Bpu1102I at 37˚C for 2 hours.
|
ATF1 into pET-15b |
15.5µL |
|
XhoI |
1µL |
|
Bpu1102I |
1µL |
|
BSA |
0.5µL |
|
NEB Buffer 2 |
2µL |
Applied pET-15b to the agarose gel electrophoresis.
No band was detected.
Plasmid DNA was digested with XhoI and Bpu1102I at 37˚C (overnight).
|
ATF1 into pET-15b |
15.5µL |
|
XhoI |
1µL |
|
Bpu1102I |
1µL |
|
BSA |
0.5µL |
|
NEB Buffer 2 |
2µL |
September 14th
ATF1 and pET-15b were applied to electrophoresis to the agarose gel.
No band was detected.
Digested ATF1 and pET-15b with Bpu1102I at 37˚C overnight.
|
ATF1 |
26µL |
|
Bpu1102I |
1µL |
|
Bpu1102I Buffer |
3µL |
|
pET-15b |
26µL |
|
Bpu1102I |
1µL |
|
Bpu1102I Buffer |
3µL |
September 15th
Purified ATF1 and pET-15b.
Digested ATF1 and pET-15b with XhoI.
|
ATF1 |
26µL |
|
XhoI |
1µL |
|
Bpu1102I Buffer |
3µL |
|
pET-15b |
26µL |
|
XhoI |
1µL |
|
Bpu1102I Buffer |
3µL |
pET-15b and ATF1 were applied to electrophoresis in the Blue gel and purified.
Ligated ATF1 and pET-15b and transformed them into E. coli cells.
September 16th
Picked 32 colonies of ATF1 transformants.
September 18th
Checked the colonies by colony cracking.
Picked up the colonies and cultured in 3mL LB medium with ampicillin at 37˚C (overnight).
September 19th
plasmid DNA was purified and digested with XhoI and Bpu1102I at 37˚C for 2 hours.
|
ATF1 into pET-15b |
15.5µL |
|
XhoI |
1µL |
|
Bpu1102I |
1µL |
|
BSA |
0.5µL |
|
NEB Buffer 2 |
2µL |
ATF1 and pET-15b were applied to electrophoresis to the agarose gel.
No band was detected.
Digested ATF1 and pET-15b with XhoI and BlpI.
|
ATF1 |
15.5µL |
|
XhoI |
1µL |
|
BlpI |
1µL |
|
BSA |
0.5µL |
|
Buffer |
2µL |
|
pET-15b |
15.5µL |
|
XhoI |
1µL |
|
BlpI |
1µL |
|
BSA |
0.5µL |
|
Buffer |
2µL |
Added 1µL of BAP to the solution of pET-15b and incubated it at 37˚C for 30 minutes.
Applied pET-15b and ATF1 to the blue gel electrophoresis.
Isolated and purified them.
The ATF1 was ligated with pET-15b, and transformed into E. coli cells.
September 20th
Picked 64 colonies of ATF1 transformants.
September 21st
Checked the colonies by colony cracking.
Picked up colonies and cultured in the LB in 3mL LB medium with ampicillin at 37˚C (overnight).
September 22nd
Miniprepped plasmid DNA and digested it with XhoI and BlpI at 37˚C for 2 hours.
Applied ATF1 into pET-15b to the agarose gel electrophoresis.
No band was detected.
