Haynes Lab:Notebook/Jan/2013/09/27: Difference between revisions
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|style="background-color: #EEE"|[[Image:owwnotebook_icon.png|128px]]<span style="font-size:22px;"> Project name</span> | |style="background-color: #EEE"|[[Image:owwnotebook_icon.png|128px]]<span style="font-size:22px;"> Project name</span> | ||
|style="background-color: #F2F2F2" align="center"| | |style="background-color: #F2F2F2" align="center"|[[File:Report.png|frameless|link={{#sub:{{FULLPAGENAME}}|0|-11}}]][[{{#sub:{{FULLPAGENAME}}|0|-11}}|Main project page]]<br />{{#if:{{#lnpreventry:{{FULLPAGENAME}}}}|[[File:Resultset_previous.png|frameless|link={{#lnpreventry:{{FULLPAGENAME}}}}]][[{{#lnpreventry:{{FULLPAGENAME}}}}{{!}}Previous entry]] }}{{#if:{{#lnnextentry:{{FULLPAGENAME}}}}|[[{{#lnnextentry:{{FULLPAGENAME}}}}{{!}}Next entry]][[File:Resultset_next.png|frameless|link={{#lnnextentry:{{FULLPAGENAME}}}}]]}} | ||
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** Results for Streaks 1 and 2: Growth in both areas along zig-zag line. Bottom left of agar plate dried out. Success. | ** Results for Streaks 1 and 2: Growth in both areas along zig-zag line. Bottom left of agar plate dried out. Success. | ||
* Sample 1: fshPCD/V0120 colony 1; BBa_J176002; part = 186 bp; vector = | * Sample 1: fshPCD/V0120 colony 1; BBa_J176002; part = 186 bp; vector = 3200 bp | ||
* Sample 2: fshPCD/V0120 colony 2; same | * Sample 2: fshPCD/V0120 colony 2; same | ||
* Sample 3: fshPCD/V0120 (control DNA); BBa_J176002; part = 186 bp; vector = | * Sample 3: fshPCD/V0120 (control DNA); BBa_J176002; part = 186 bp; vector = 3200 bp | ||
'''DNA Concentration Data''' | '''DNA Concentration Data''' | ||
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| bgcolor=#cfcfcf | Reagent | | bgcolor=#cfcfcf | Reagent | ||
| bgcolor=#cfcfcf | Volume | | bgcolor=#cfcfcf | Volume | ||
| rowspan="7" | <u>Expected:</u><br>1. Plasmid 1 = 186 bp, | | rowspan="7" | <u>Expected:</u><br>1. Plasmid 1 = 186 bp, 3200 <br>2. Plasmid 2 = 186, 3200 <br> | ||
| rowspan="7" | [[Image: | | rowspan="7" | [[Image:Jan Simper fshPCD.jpg|400px|Today's gel]]<br>15 μL/lane; 1% agarose; [http://openwetware.org/wiki/Image:KAH_Fermentas_GeneRuler_1kbplus.jpg Ladder] | ||
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| DNA(plasmid) || 3.0 μL | | DNA(plasmid) || 3.0 μL | ||
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'''Conclusion:''' All samples had a band just under the 5000 bp band (the brightest top band), which was probably the 3200 bp vector. Also, the expected part base pair was 186 bp, and the band in the picture is slightly above the 200 bp band, but all three samples including the control DNA had the band at the same location, so overall, the restriction digest was successful. However, additional help about the two bands around the 1500 bp location that appeared in samples 1 and 2 but not in the control DNA will be received before continuing. After talking with the professor, a possible explanation is that this band was supercoiled undigested plasmid DNA. It was decided to submit samples 1 and the control DNA for DNA sequencing. | |||
'''DNA Sequencing Samples''' | '''DNA Sequencing Samples''' | ||
<!-- * signs create an automatically bulleted list. Replace 'date' with the date you submitted the samples --> | <!-- * signs create an automatically bulleted list. Replace 'date' with the date you submitted the samples --> | ||
<!-- The Order Number is shown at the top of your sequencing order form. Record in the indicated spot below --> | <!-- The Order Number is shown at the top of your sequencing order form. Record in the indicated spot below --> | ||
* Submitted to DNASU on: ' | * Submitted to DNASU on: '10/8/13' | ||
* Order Number: | * Order Number: 8215 | ||
<!-- # signs create an automatically numbered list. Replace Plasmid 1 & 2 with the names of your plasmids. Replace 'name' with the appropriate name of each primer. --> | <!-- # signs create an automatically numbered list. Replace Plasmid 1 & 2 with the names of your plasmids. Replace 'name' with the appropriate name of each primer. --> | ||
# | # fshPCD - forward primer 'P0001' 186/186 matches. 100% alignment across the sequence. | ||
# | # fshPCD - reverse primer 'P0002' 186/186 matches. 100% alignment across the sequence. | ||
# | # fshPCD-control DNA - forward primer 'P0001' 186/186 matches. 100% alignment across the sequence. | ||
# | # fshPCD-control DNA - reverse primer 'P0002' 186/186 matches. 100% alignment across the sequence. | ||
Latest revision as of 23:23, 26 September 2017
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Plasmid Extraction and Validation of fshPCD
DNA Concentration Data
Conclusion: All samples had a band just under the 5000 bp band (the brightest top band), which was probably the 3200 bp vector. Also, the expected part base pair was 186 bp, and the band in the picture is slightly above the 200 bp band, but all three samples including the control DNA had the band at the same location, so overall, the restriction digest was successful. However, additional help about the two bands around the 1500 bp location that appeared in samples 1 and 2 but not in the control DNA will be received before continuing. After talking with the professor, a possible explanation is that this band was supercoiled undigested plasmid DNA. It was decided to submit samples 1 and the control DNA for DNA sequencing. DNA Sequencing Samples
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