Difference between revisions of "Diy-transilluminator"
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Primers for this test and the protocol were obtained from Delphine Ducoulombier of l'Eprouvette. <br> Originally, these are based on a Cold Spring Harbour lab test (in their DNA Learning Center). | Primers for this test and the protocol were obtained from Delphine Ducoulombier of l'Eprouvette. <br> Originally, these are based on a Cold Spring Harbour lab test (in their DNA Learning Center). | ||
Q5 polymerase was used for the amplification, and DNA from Sam (a non-taster) and Rachel (a taster) was used for the test. | Q5 polymerase was used for the amplification, and DNA from Sam (a non-taster) and Rachel (a taster) was used for the test. | ||
| − | The DNA was a quick GnG prep that had been concentrated by pptn, after apparently no product was obtained the first PCR attempt, (but Rachel's sample had less DNA than Sam's... also less cells to start!). Stages of looking at the first gel are shown in images 6-8 of the gallery.<br> | + | The DNA was a quick GnG prep that had been concentrated by pptn, after apparently no product was obtained the first PCR attempt, (but Rachel's sample had less DNA than Sam's... also less cells to start!). Stages of looking at the first gel (2% agarose in TAE) are shown in images 6-8 of the gallery.<br> |
| − | + | A new set of PCR reactions with concentrated DNA was run on 11dec, and some of the reactions was cut with HaeIII. Images from this expt are shown as 9-10 in gallery, and it is clear that this DIY transilluminator is not as sensitive as the UNIL 'SafeImager' from Promega - image 12... <br> | |
| − | The non-taster band that does not cut with HaeIII (predicted size 221bp) is visible from Sam's DNA, but no virtually no product | + | The non-taster band that does not cut with HaeIII (predicted size 221bp) is visible from Sam's DNA, but no virtually no product is visible from Rachel's... (tasters are expected to have the band cut for 177 + 44bp - perhaps there is a difference in the HaeIII cut lane of RA, vs the uncut, with something smaller perhaps just there??)<br> |
| + | The PCR test should be done again with more input DNA from RA, but also the midi size trans-illuminator should be built. | ||
Revision as of 13:21, 12 December 2017
Transilluminators are especially useful to see DNA bands on a gel. While there is an old upcycled imager in the P1, this has detector problems, so sometimes much of the gel is not even visible... Therefore, we have been trying out new 'DIY-transilluminators' with a little help from some Hackteria friends and others (htgaa)!
DIY Transilluminators
Urs brought his GaudiLabs kit to Hackuarium on 2dec, and he helped Rachel do the soldering and put it all together.
He said it needs a diffuser of some sort to help avoid seeing the whole circuit board below, and that the circuit board itself fluoresces in this blue LED light, which is an unsolved problem.
The first five pictures in the gallery show various stages of this process.
Dan also ordered bigger filters and a string of bright blue LEDs to try making a new 'midi' size transilluminator, soon!
11dec17 gel - second PTC PCR test
The chemical PTC can be tasted or not tasted, depending on a GC polymorphism in the TAS2R38 bitter taste receptor.
Primers for this test and the protocol were obtained from Delphine Ducoulombier of l'Eprouvette.
Originally, these are based on a Cold Spring Harbour lab test (in their DNA Learning Center).
Q5 polymerase was used for the amplification, and DNA from Sam (a non-taster) and Rachel (a taster) was used for the test.
The DNA was a quick GnG prep that had been concentrated by pptn, after apparently no product was obtained the first PCR attempt, (but Rachel's sample had less DNA than Sam's... also less cells to start!). Stages of looking at the first gel (2% agarose in TAE) are shown in images 6-8 of the gallery.
A new set of PCR reactions with concentrated DNA was run on 11dec, and some of the reactions was cut with HaeIII. Images from this expt are shown as 9-10 in gallery, and it is clear that this DIY transilluminator is not as sensitive as the UNIL 'SafeImager' from Promega - image 12...
The non-taster band that does not cut with HaeIII (predicted size 221bp) is visible from Sam's DNA, but no virtually no product is visible from Rachel's... (tasters are expected to have the band cut for 177 + 44bp - perhaps there is a difference in the HaeIII cut lane of RA, vs the uncut, with something smaller perhaps just there??)
The PCR test should be done again with more input DNA from RA, but also the midi size trans-illuminator should be built.
Gallery
<gallery mode=packed widths=300 heights=300> File:IMG 2911.JPG File:IMG 2913.JPG File:IMG 2919.JPG File:IMG 2920.JPG File:IMG 2921.JPG File:IMG 3017.JPG File:IMG 3021.JPG File:IMG 3022.JPG File:IMG 3362.JPG File:IMG 3370.JPG File:1409taw4.jpg File:IMG 3373.JPG