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− | The entire process of the DNA extraction using the MN patches takes less than 1 min, orders of magnitude faster and simpler than the conventional extraction protocol.</p> </div>
| + | We compared our DNA extraction method using microneedles with the conventional extraction method, by characterizing |
− | <h5 style="color:indigo">Comparison of extraction efficiency between the MN and the Dneasy methods. </h5> | + | the quantity and purity of DNA extracted with both methods.</p> </div> |
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| <div class="post-content"><p style="font-size:17px; align="justify"> | | <div class="post-content"><p style="font-size:17px; align="justify"> |
− | We compared our DNA extraction method using microneedles with the conventional extraction method, by characterizing the quantity and purity of DNA extracted with both methods. The total extracted DNA and the purity of DNA obtained by both extraction methods were characterized by a Nano- Drop spectrophotometer. From the spectral data, it is clear that the MN patch extracted a significant amount of DNA from plant leaves based on the appearance of A260 absorption for all samples tested. In contrast, the solution from the blank MN patch did not show any significant absorption at 260 nm. The purity of MN-extracted DNA samples was compared with those obtained by conventional extraction method. <br> We can also note that the conventional method takes 1 to 2 hours and requires a lab, and is therefore more than 100 times more time consuming than MN extraction, which is non neglectable. </p> | + | We compared our DNA extraction method using microneedles with the conventional extraction method, by characterizing |
| + | the quantity and purity of DNA extracted with both methods. The total extracted DNA and the purity of DNA obtained by |
| + | both extraction methods were characterized by a Nano- Drop spectrophotometer. From the spectral data, it is clear |
| + | that the MN patch extracted a significant amount of DNA from plant leaves based on the appearance of A260 absorption |
| + | for all samples tested. In contrast, the solution from the blank MN patch did not show any significant absorption at |
| + | 260 nm. The purity of MN-extracted DNA samples was compared with those obtained by conventional extraction method. <br> |
| + | We can also note that the conventional method takes 1 to 2 hours and requires a lab, and is therefore more than 100 times |
| + | more time consuming than MN extraction, which is non neglectable. </p></div> |
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| <div class="post-header"> | | <div class="post-header"> |
− | <h1 class="post-title" style="color:purple">Successful DNA Extraction</h1> | + | |
− | | + | <h5 style="color:indigo">Analysis with a Nanodrop spectrophotometer </h5> |
| </div> | | </div> |
− | <div class="post-content"><p style="font-size:17px;" align="justify">
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− | <p style="font-size:17px; align="justify">
| + | <div class="post-content"><p style="font-size:17px;" align="justify"> |
− | We amplified different the EC (Endogeneous Control) by PCR and ran a gel electrophoresis to analyze the results. We loaded the gel from left to right as following, the ladder, synthetic EC DNA sequence, DNA extracted with a microneedle patch, DNA extracted with a standard kit, control with primers but no DNA. Bands are observed in all the lanes containing DNA, no band is present in the control. This shows that the microneedle patch successfully extracted DNA from the plant, which was then amplified by PCR. </p> | + | We used the Nanodrop to analyze the DNA extracted with a microneedle patch (red). The positive control (green) |
| + | is EC synthetic DNA sequence and the negative control (blue) is the elution buffer applied on an unused microneedle patch. </br> |
| + | As expected, the negative control absorption at 260nm is close to zero, no DNA is present in the microneedle patch. |
| + | In contrast, the positive control has the typical DNA Nanodrop readout. </br> |
| + | The DNA extracted by microneedle patch shows significant absorption at A260. Its line higher than the negative |
| + | control but lower than the positive control. This means that some DNA was extracted but that it is not pure, |
| + | which is normal since, as mentioned previously, the patch extracts DNA and other molecules on the leaf. |
| + | </p> |
| | | |
− | | + | <h5 style="color:indigo">PCR amplification and gel electrophoresis </h5> |
| <img src="https://static.igem.org/mediawiki/2019/c/c9/T--EPFL--3courbes.png" > | | <img src="https://static.igem.org/mediawiki/2019/c/c9/T--EPFL--3courbes.png" > |
| <p style="font-size:17px; align="justify"> | | <p style="font-size:17px; align="justify"> |
− | We used the NanoDrop to analyse the DNA extracted with a microneedle patch (red line). The positive control (green line) is EC synthetic DNA sequence and the negative control (blue line) is the elution buffer applied on an unused microneedle patch. </br> | + | We amplified different EC DNA by PCR and ran a gel electrophoresis to analyze the results. |
− | | + | We loaded the gel from left to right as following, the ladder, synthetic EC DNA sequence, |
− | As expected, the negative control line is close to zero, no DNA is present in the microneedle patch. The positive control has the typical DNA nanodrop readout. The DNA extracted by microneedle patch shows a line higher than the negative control but lower than the positive control. This means that some DNA was extracted but that it is not pure, which is normal since the patch doesn’t only extract the DNA in the leaf.
| + | DNA extracted with a microneedle patch, DNA extracted with a standard kit, control with primers but no DNA. |
− | | + | Bands are observed in all the lanes containing DNA, no band is present in the control. |
| + | This shows that the microneedle patch successfully extracted DNA from the plant, which was then amplified by PCR. |
| </p> | | </p> |
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| <div class="post-content"><p style="font-size:17px; align="justify"> | | <div class="post-content"><p style="font-size:17px; align="justify"> |
− | To sum up, we demonstrated that microneedle DNA extraction can be a successful, nondestructive, cell-lysis-free, and purification-free plant DNA extraction method. We managed to reduce the time of the conventional few hour extraction to a few minutes extraction using microneedle patches. <p>
| + | We demonstrated that microneedle patches can be used to extract DNA from leaves in less than 1 minute. |
| + | The extracted DNA could be then amplified by PCR and analyzed by gel electrophoresis. |
| + | We can also note that the conventional method takes 1 to 2 hours and requires a lab and is |
| + | therefore more than 100 times more time consuming than microneedle extraction, which is non neglectable. |
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