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1. Get 3 fresh buccals and Lyse-n-Go them
2. While the lysing is occurring prepare the PCR rxns, below is for 1 tube:
25ul go green
1ul keratin sense
1ul keratin as
FV = 50ul
3. Once lysis is holding at 80C add the 5ul to the PCR mix, pulse vortex and pellet
4. Run the PCR cycles at the highest temp that still gave good pdt previously
5. Load and run 15ul, hope for a single, strong band
-if no bands appear don’t use the sample
-if a second band appears over 50bp don’t use the sample
-if a second band is below 50bp use the sample
-if you have a single band use the sample
6. Take any good samples to column purification
7. In the last step of column purification add 30ul of elution buffer and let it sit in the column for 1
minute to allow for a greater concentration (rather than the usual 50ul)
8. Take 5ul of eluate to spectroscopy and hope we have enough at a high enough concentration
to move onto sequencing
Now we get to the sticky part of the class. Our experiments should work, yet in the past I've had poor overall success. What we will be doing is indeed trial and error, such is science. On Tuesday I will lecture a little on DNA and then we'll set up restriction digestions of our genomic DNA. There is no quiz.
So I need you to calculate the following:
1. How much genomic is needed to put 2ug in a tube to reach a FV of 15ul with the addition of water
2. How to set up a restriction digestion given the following information:
2 ug of genomic DNA
1ul of EcoRI
Xul of 10x buffer
FV = 15 ul
3. Set up a SalI digestion:
10ug of genomic DNA
15u/ug of enzyme to DNA if the enzyme comes at a concentration of 10u/ul
how much 10x buffer?
how much water?
reach some final volume (it should be around 150ul)
So in other words, review how to set up digestions and we'll do it at the end of class. I've mapped out a strategy for the next 3 classes, cross your fingers!
Alan Wong's Restaurant + Surf Lanai | Honolulu, Hawaii: "Last week Friday we had the most epic date night. Thanks to a generous gift certificate for Alan Wong's Restaurant and our Waikiki Gold Card, we were able to indulge in a little bit of awesomeness, without over-spending.
Alan Wong's Restaurant
1857 S King St
Honolulu, HI 96826
The Royal Hawaiian - Surf Lanai Restaurant
2259 Kalakaua Ave
Dinner was at the prestigious Alan Wong's Restaurant.
Wifey started with the Lychee Colada cocktail. It's a spin on the traditional piña colada with lychee in place of pineapple juice.
As the designated driver, I opted for the Plantation Iced Tea.
We started with the Hamakua Springs Tomato, Beet and Avocado Salad. The Li Hing Mui Ume Vinaigrette is amazing. Neither flavor was overly stated and it brought a whole new level of complexity to the dish.
Appetizer number 2 was the warm asparagus. It was good, but not extraordinary.
They were more than helpful in accommodating wifey's vegetarian diet. Here is a special Ginger Crusted Tofu (basically, it's the Ginger Crusted Onaga off the menu, with tofu instead of fish). Thanks to @ryankanno for suggesting we ask about this specific dish.
Along with wifey's dish came this side of potato.
My entrée was one of their specials that evening: the Surf and Turf. It was a petite tenderloin (cooked medium-rare, of course) with seafood risotto, topped with a poached egg. You can tell from the flavor that the beef was grass-fed and the seafood was fresh. Everything was cooked perfectly; nothing was overdone.
Dessert was Five Spoons of Brulees. An interesting concept, but not quite as satisfying as a other crème brûlée dishes we've had.
We spend the night at the Royal Hawaiian Hotel - a mini stay-cation, if you will. So for breakfast the next day, we decided to check out the Surf Lanai restaurant. The started us off with a nice sized basket of various breads.
Wifey ordered the vegetarian pizza. The crust was very thin and she said it tasted like everything came straight from the farmer's market.
Here are the fries that came with my sandwich.
I ordered the Kalua Pork Sliders. There was a bonus slice of Portuguese Sausage in each one. I love the 'gourmet' take on local favorites."
Bonnie Burton, our crafty pal over at the official Star Wars blog, shares an interview with Star Wars devotee, Damian Dayton, who customizes his shoes based on his favorite characters from the films. He explains his tips and tricks for making your own. I'm thinking I may need to make a pair of these for my Star Wars-obsessed son, and then .... a pair for myself.
May 1940. "Untitled." Mr. U and the rest of the Untitleds somewhere near Durham, North Carolina. 35mm nitrate negative by Jack Delano. View full size.