Gel Electrophoresis Documentation and links ODIN http://www.kerafast.com/PDF/sybr-green-gel-staining.pdf http://people.virginia.edu/~owp3a/docs/protocols/Preparing%20agarose%20gels.pdf https://www.researchgate.net/post/I_am_using_Sybre_Safe_to_stain_the_gel_electropheresisi_5microlitre_in_each_welll_but_donot_got_any_band_i_just_saw_ladder_band_i_am_adding_6x_dy http://www.science.smith.edu/cmbs/wp-content/uploads/sites/36/2015/09/Gel-Electrophoresis-Protocol.pdf https://www.biorxiv.org/content/10.1101/568253v1.full.pdf https://www.thermofisher.com/ca/en/home/brands/thermo-scientific/molecular-biology/molecular-biology-learning-center/molecular-biology-resource-library/spotlight-articles/8-DNA-ladder-tips.html (Choosing the optimal agarose gel concentration) https://www.researchgate.net/post/What_is_the_minimum_DNA_quantity_ng_required_to_visualize_DNA_on_Agarose_gel for how much DNA https://www.goldbio.com/articles/article/AgaroseGelFAQs for reusig th gel Bioengineering 102 - Class 3 - Analyzing DNA Using Agarose Gels the-odin video What is needed: Devices: Gel Box + Lid Tray Tape Power supply parafilm or alum foil alum foil 2 mm comb depth spacer --reagents etc--- 25 ml TAE .25 or .5 g Agarose powder for shorter DNA 3 ul Gel Stain TAE - a lot 2 times about .5 liter loading buffer 10ul of each DNA sample DNA ladder water tips large and small pipette ========== START ========== prepare the box tape the tray ------------------ take 25 ml TAE bufer(0.125 g TAE for 25 ml distilled water) add .25g Agarose powder which is 1% OR (2% so .5g for gels with 100–2,000 bp). We can increase the agarose to .75g? for greater line distace in 100bp ladders. Microwave let it cool to 50 Celsius until hand acceptable. Do't leave it too long!!! while hot add and mix by gently stirring 3ul of DNA Safe Gel Stain (Sybre Safe?) ---------this will give about 7 mm total height of the gel and a 5 mm depth of the wells which at 1 mm x 4 mm is about 20 ul volume. For 10ul sample +10 ul loading die with NO space on top so it's a problem!! It will work for 5ul sample +5ul loadig dye=10ul total take the tray out of the box and put in the position and direction where the box will finally be use the 2 mm comb depth spacer to screw in the comb at the proper depth load in the tray gently to avoid bubbles making sure is leveled. If bubbles appear burst them with a pipette tip LETS DO 10 ul sample and 10ul loading buffer per each WELL (ecept the ladder 5 ul) for a total of 20ul per well* AND 100 bp DNA ladder which varies from 100bp to 1500 bp hich means the Agarose should be 2% so !!!! (https://www.thermofisher.com/ca/en/home/brands/thermo-scientific/molecular-biology/molecular-biology-learning-center/molecular-biology-resource-library/spotlight-articles/8-DNA-ladder-tips.html) This should yield a gel about 6.5 mm. It is best to make the gels as thin as possible. Gels 0.75 cm thick or less are appropriate for general applications (less than 25 µL sample volume per well). Thicker gels may be required if you have a large sample volume, e.g., when purifying DNA fragments" Add comb cover the tray with aluminum because the stain is light sensitive place tray in fridge or air cool and wait until fully cured. The gel becomes slightly opaque then. About 20 minutes. make exactly 450 ml TAE buffer for filling the box. That for 5g/liter is 2.25 grams. Stir it manually with magnetic stirrer for 5 minutes to fully dissolve. It's easier to put all solids and little water to stir it manually. fill box with TAE buffer until it slightly covers the top of the gel about 450 ml 0 put in 10ul* DNA ladder in lane 1 (it already has the loading buffer) pipette 3ul loading buffer (6x) on parafilm or aluminum foil (should be about 1ul or more for each 5 ml of samples and it can be more if ou want to) pipette and pipette mix 10ul of sample(that is 200g!!) 0 with each drop of loading buffer then reload and put in comb weel. Repeat for all aliquots, changing the tip. The gel with dye has a shelf life of only a few days and needs to be in the dark (From one of the links for TBE not TAE. For example, if you run TBE gels and require 30 mL of molten agarose for your tray, add 3 µL of 10,000X SYBR® Safe stain concentrate to 30 mL of 1X TBE and mix well. Add this stain–TBE mixture to the powdered agarose. The agarose/SYBR® Safe stain mixture may be heated in the microwave. ) ===== Cost TAE 1$ Ladder at 7ul .7$ Stain @3ul=.3 $ Loading dye @3ul=0.05S Agarose 1x=.25g or .5g for 2x(100bp) =.5$ or .25$ Total =2.5$ per run or 2 $ when reusing gel =============Last try=== 100ul ladder from Odin https://www.the-odin.com/dna-ladder-100bp/ which seems to be similar to (TODO check the actual photos) http://bioland-sci.com/index.php?main_page=product_info&products_id=385 5ug Ladder 5ug Gel Stain (works with 3ul too) https://www.the-odin.com/dna-stain-for-gels-10-000x/ which is the SAME as http://bioland-sci.com/index.php?main_page=product_info&cPath=123_187&products_id=947 80 volts old TAE 2mm height comb spacer 25 ml TAE+.5 g Agarose =>65 mm gel about 25 minutes for ceter position? camera Manual FOcus ISO 100, 1/6 NO TAE over =========== https://docs.google.com/document/d/10Y4NgXjMRvG_Vd5D4o2lPJp1ehY2R516gpfrh2PZWfw/edit http://bioland-sci.com/index.php?main_page=index&cPath=123_187 which is the same description as ODIN "Product Description so ODIN sells Bioland!!!! the one I have is http://bioland-sci.com/index.php?main_page=product_info&products_id=385 "This DNA stain is a safe class of fluorescent dyes, used for agarose or polyacrylamide gel staining to visualize DNA or RNA. You can use it the same way as you would use ethidium bromide. It emits green fluorescence when bound to DNA and red fluorescence when bound to RNA when exposed to UV light or green fluorescence when exposed to blue light. It has two excitation maxima: 290nm and 490nm. Most blue LEDs work great." And the same for Safe DNA Gel Stain http://bioland-sci.com/index.php?main_page=index&cPath=123_187 http://bioland-sci.com/PDF/BLooK.pdf The blue LED lights are arranged under the viewing area (200 × 120 mm). An amber filter, on hinges, is lowered into position once your gel is mounted. (http://bioland-sci.com/index.php?main_page=product_info&cPath=123_188&products_id=952)