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Zach T's Content

There have been 72 items by Zach T (Search limited from 20-January 19)



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#112284 Insert magically shortening???

Posted by Zach T on 10 June 2011 - 07:18 AM in Molecular Biology

The PCRs only amplify small segments of the insert. The idea was that with a gel purified 12kb insert, there shouldn't be anyway to modify it once its in the cell so if we can amplify the 3' and 5' end, it should all be there. Do you think there are issues with that thought?

So I have used a few different enzymes to digest my recovered plasmid from the cells and all the fragments from each digestion add up to around 8kb instead of the 16kb (12kb insert+4kb vector) that I'm expecting.



#112236 Insert magically shortening???

Posted by Zach T on 09 June 2011 - 02:47 PM in Molecular Biology

Hey everyone,

I am trying to clone a 12kb insert into a pBS vector (~4kb) using In-Fusion cloning which uses 15bp homologous ends instead of TA cloning. I linearized the vector with EcoRV which is a blunt end digestion enzyme and gel purify it. I am also using Takara's LAPCR to amplify the 11kb insert and gel purify it as well. I go through the ligation, trans, and plating, however when I screen the colonies I get some very weird results.

First, using digestion enzymes to linearize the vector and vizualize it on a gel, I only get a length of around 7-8 kb which would mean that my insert is only ~3-4kb instead of 12kb. I"m not sure why this is happening, or how this would even happen. Also, to confuse things even more, I have been screening these inserts by doing a basic PCR using primers that target the 3', 5' and middle section of the insert and they all amplify at the correct lengths which would suggest that the entire insert is present in the vector. But the digestion says otherwise.

Any suggestions/comments/experiences?

Thanks!



#108920 Help a couple of snowboard bums ;)

Posted by Zach T on 05 May 2011 - 10:21 AM in Chit Chat


Yeah, I already have gym that I like but I think I'll be able to handle the altitude or at least gut it out. Its nice being able to look at the mtn every day from my desk window...its a beast!!

:P Gut it out?.....I hope that it's not only your guts talking....anyways, if you think you can handle the beast then you probably can....
and you have an awesome view of Mt Rainier everyday? Lucky you...

Yeah, I have a great view of the mtn but sadly on overcast days, I can't see it at all...so about most the time I can't see it at all. :lol:



#108804 Help a couple of snowboard bums ;)

Posted by Zach T on 04 May 2011 - 11:10 AM in Chit Chat

Yeah, I already have gym that I like but I think I'll be able to handle the altitude or at least gut it out. Its nice being able to look at the mtn every day from my desk window...its a beast!!



#108677 Help a couple of snowboard bums ;)

Posted by Zach T on 03 May 2011 - 08:13 AM in Chit Chat


Haha, that was really funny. There was some pretty old footage in there too!

Yeah, we are planning on doing a 3 day trip for the Rainier hike. I'm not sure how I'm going to prepare for the elevation. I grew up at 6600 ft (2000m) and routinely went up to 11000 ft (3350m) to go snowboarding and hiking but now I've been living at sea level for nearly 5 years. It'll be interesting. Mt Whitney will be a good warmup cuz its only a day hike but is actually taller than Rainier so I'll get a gist of the altitude and oxygen levels

funny yup...but most were pretty stupid....:P
as to the altitude training....well, if Zach T can't come to the mountain then the mountain has to come down to Zach T ...how about some kind of hypoxic training (special tent, bed, mask etc)? The IOC hasn't banned this yet...the "live/breathe/sleep high but train low" regimen....

Oh thats pretty awesome!! I had never really heard of that but it completely makes sense. Too bad those tents/machines probably cost a pretty penny...probably not worth it for my "fun" hikes and triathlons :lol: but it would be pretty cool to just sleep at simulated 22,000 ft elevation!

I need to get sponsored by some supplement company or something, that would make things so much easier :P



#108389 Help a couple of snowboard bums ;)

Posted by Zach T on 29 April 2011 - 10:32 AM in Chit Chat

Haha, that was really funny. There was some pretty old footage in there too!

Yeah, we are planning on doing a 3 day trip for the Rainier hike. I'm not sure how I'm going to prepare for the elevation. I grew up at 6600 ft (2000m) and routinely went up to 11000 ft (3350m) to go snowboarding and hiking but now I've been living at sea level for nearly 5 years. It'll be interesting. Mt Whitney will be a good warmup cuz its only a day hike but is actually taller than Rainier so I'll get a gist of the altitude and oxygen levels



#107996 Help a couple of snowboard bums ;)

Posted by Zach T on 26 April 2011 - 09:02 AM in Chit Chat

ice axes and crampons? be careful dude....but we would like to see a vid of you sliding.....:) you guys are serious ice mountaineers?

and btw, are we gonna learn a new lingo again- clearing the wake...scarecrows and air raleys....but you haven't "stepped up to the 3s"? You mean 360 degree spin in the air? Your buddies can do this? ppfftt...really sick...


Yeah, the Rainier summit will be interesting for sure. I'll def get some footy from that.

Haha, yeah wakeboarding has its own ridiculous jargon...but for any board sport they are all now just shortening how they call out 360s and 540s etc to 3s, 5s, 7s, 9s, 10s, etc.

Check out this video I found last night, makes my life seems extremely boring...
http://www.youtube.com/watch?v=jNcXn9VPk1U



#107935 Help a couple of snowboard bums ;)

Posted by Zach T on 25 April 2011 - 01:55 PM in Chit Chat

You're definitely a rider, Zach T....so can you also do some wakeboarding tricks? :P And your two mountains to summit- I had to do the metric conversion though...you'd take your go pro, wouldn't ya? Post some pix/vids...


I can clear the wake and usually throw in some grabs and 180s. Haven't stepped up to the 3s or bigger spins but I only get out a couple times a year opposed to snowboarding where I get up 20-30 times. A couple of my buddies compete tho, so they can throw the backflips and scarecrows and such, pretty sick riders. :o

I'll definitely bring my cameras up for the hikes. Mt Whitney trip is June 29th and then Mt Rainier is August 12-14th...so I have a bit of time. Need to start training tho! :D Whitney should be pretty simple and straight forward, just a day hike but Rainier will be intense with the glaciers and all (fun with ice axes!! :lol: )



#107890 Help a couple of snowboard bums ;)

Posted by Zach T on 25 April 2011 - 06:37 AM in Chit Chat

but the summer season should start soon? You've got something else going there or is it really just for winter activities (I've never been to Stevens Pass) and Seattle sucks during winter....dark, cold, rainy and the city stops when snow falls.....:lol:..


Steven's pass just got approved to build their bike park for summer activities, so that'll hopefully open end of summer, but pretty stoked on that. You're right tho, winter in seattle blows, and nobody here knows how to drive in the rain, and panic terribly when they see one snowflake. :lol: But Saturday was our first glimpse of spring/summer type weather so hopefully good weather will start to roll in. I pretty much just wakeboard, mtn bike and hike in the summer. I have two mtns to summit this summer, both around 14.4 kfeet! :D



#107738 Cloning two inserts at a time in a vector

Posted by Zach T on 22 April 2011 - 02:30 PM in Molecular Cloning

Not 100% sure on your inserts but I know that Infusion cloning from clontech allows you to have a 15bp overhang on each of your inserts so you can insert multiple into one vector. I'd check out their kit, its pretty good.



#107737 Help a couple of snowboard bums ;)

Posted by Zach T on 22 April 2011 - 02:23 PM in Chit Chat

hey Zach T....I just watched this one: :P

http://www.youtube.com/watch?v=RGTB-wNO85o&feature=related

it's a pretty decent pond....and it was snowing, right? Any of you guys swallowed a rubber ducky? And of course, skiers always better at hydroplaning....:lol:...

Haha, yeah I watched that video the other day, pretty cool angles they had, solid crashes too. We were so bummed tho that they didn't put us in the video (guess we were just too good and classy :P ) I think they mainly wanted to show off the crashes. One guy did email us some pics so we at least got some photos of us on the water but they are on facebook and not flickr.

So sad the season is over tho :( lol



#107265 Help a couple of snowboard bums ;)

Posted by Zach T on 19 April 2011 - 01:11 PM in Chit Chat




I just saw your group pic, Zach T...with the fat cigars and classy suits....really gangsta.....:P


Hahaha, yeah we looked good!! :lol: Got those magnificent clothes all for under $20 too, clutch! Glad you approve tho :P

yup, not bad, not bad...the only thing missing was the getaway car (or snowmobile) :D...but the funky chicken was probably better....

The chicken costume was pretty funny, there was also a pink gorilla...both of them didn't do too well tho. I think the best ones were the guys who didn't care about getting wet and would just spray the crowd instead :D no free shows :lol:



#107222 Help a couple of snowboard bums ;)

Posted by Zach T on 19 April 2011 - 06:42 AM in Chit Chat

I just saw your group pic, Zach T...with the fat cigars and classy suits....really gangsta.....:P


Hahaha, yeah we looked good!! :lol: Got those magnificent clothes all for under $20 too, clutch! Glad you approve tho :P



#107142 Help a couple of snowboard bums ;)

Posted by Zach T on 18 April 2011 - 11:49 AM in Chit Chat


It was decent sized, at least about 50 ft. Had to straight line it from the top to make it across. Anyone who speed checked at all was soaked. :lol:

Sadly, you'll have to wait until tomorrow to see our amazing outfits (sry, don't have the pic uploaded to anything yet so can't insert a link).

But tomorrow I'll also upload a pic of the pond and the run where everyone was starting!

can't open your photo from here but the participants used only snowboards and skis for the pondskimming? and how was the after party? Now those pix or vids would be more interesting...:P

There were both skiers and boarders in it. Skiers definitely had the advantage in it, but we represented the snowboarding community very well, haha. We didn't really stay too long after we crossed, I just snapped a few pics of other ppl crashing and then we took off. Sadly i didn't get any video of the event but I'm really hoping the resort posts a video of the event b/c I know i saw them filming it and a ton of ppl were taking pics. I'm hoping i can find some of us crossing it in our suits, would be dope!



#107124 Help a couple of snowboard bums ;)

Posted by Zach T on 18 April 2011 - 10:37 AM in Chit Chat

It was decent sized, at least about 50 ft. Had to straight line it from the top to make it across. Anyone who speed checked at all was soaked. :lol:

Sadly, you'll have to wait until tomorrow to see our amazing outfits (sry, don't have the pic uploaded to anything yet so can't insert a link).

But tomorrow I'll also upload a pic of the pond and the run where everyone was starting!



#107113 Help a couple of snowboard bums ;)

Posted by Zach T on 18 April 2011 - 08:38 AM in Chit Chat

So pond skimming was this last weekend!! And all four of us actually made it all the way across the pond, and didn't get wet! sry to disappoint :lol:

But since I know you wanted to see a splash, here is someone who didn't make it:
http://www.flickr.co...tar/5629391693/

And I'll probably throw the picture of our group up tomorrow. We were by far the classiest bunch there, we were all dressed in gross old school suits and had fatty cigars in, definitely a crowd favorite (mainly b/c of the classiness and that we all killed it hydroplaning!) :D



#106239 Help a couple of snowboard bums ;)

Posted by Zach T on 08 April 2011 - 11:56 AM in Chit Chat




Haha, true. Cross dressing would be too far tho :P

but you did say epic, right?

I was just reminded of one of the episodes of The Big Bang Theory...when they lost a bowling bet and had to pay up by wearing such costumes...really hilarious...but yup, better not to go that far...:P...how about a more sciencey or nerdy theme? Darwinian? Survival of the fittest and all that stuff?

haha yeah, we'll see what we end up doing. I want like a full animal costume to ride in for the day but those are probably expensive :( Would be pretty funny tho!

well, you can make them- chimaeras would be cool.....and there won't be any riding, right? or is this another one of your fancy words? :P

Wayyy too much effort to make a costume especially since its just gonna get gross and wet! :lol: I was just saying to ride in as in 'snowboard in' rather than ride on or something. I need the bear/hunter costume from super troopers :D that would be classic!
Posted Image :blink:

I don't think that I could ride on the chairlift with that, now that i think about it :lol:



#106226 Help a couple of snowboard bums ;)

Posted by Zach T on 08 April 2011 - 10:49 AM in Chit Chat


Haha, true. Cross dressing would be too far tho :P

but you did say epic, right?

I was just reminded of one of the episodes of The Big Bang Theory...when they lost a bowling bet and had to pay up by wearing such costumes...really hilarious...but yup, better not to go that far...:P...how about a more sciencey or nerdy theme? Darwinian? Survival of the fittest and all that stuff?

haha yeah, we'll see what we end up doing. I want like a full animal costume to ride in for the day but those are probably expensive :( Would be pretty funny tho!



#106141 Help a couple of snowboard bums ;)

Posted by Zach T on 07 April 2011 - 02:38 PM in Chit Chat

Haha, true. Cross dressing would be too far tho :P



#106107 Help a couple of snowboard bums ;)

Posted by Zach T on 07 April 2011 - 08:34 AM in Chit Chat

yup, you're an adrenaline junkie, Zach T :P....nitro circus...warrior dash...I saw the scahedule of Warrior Dash this year and it's coming to Ontario.....pretty extreme obstacle courses, eh e.g. mud bath with barbed wire......anyways, don't forget- keep you body attached to your head and the go pro to your body so we can have that first-hand experience of the event....:lol:...


I would look like such a dork with the go pro mounted during that race...would be slightly entertaining tho. My roommates and I need to come up with some epic outfits to wear to this, you know...body paint, viking helmets etc :D



#106005 Words, Words, Words

Posted by Zach T on 06 April 2011 - 01:19 PM in Chit Chat

soggy

figured i'd break this up a bit



#106004 Help a couple of snowboard bums ;)

Posted by Zach T on 06 April 2011 - 01:18 PM in Chit Chat

:) I've seen those crazy events.... the contraptions fall apart during the run and then all you've got are the drivers in skimpy clothes jumping into the pond...really crazy....

flaming logs and barbed wire? So you wanna have a second career in the circus (in case your research career doesn't pan out)...great...post some pix...


Haha :lol: i'd be a part of nitro circus (you might have to look that one up) :P

I'll just strap my go pro to myself for the warrior dash (did you see the go pro's in the toboggan races?!) :D Get you a first person experience of the event!



#105969 Help a couple of snowboard bums ;)

Posted by Zach T on 06 April 2011 - 08:58 AM in Chit Chat


At least they are fun carbon footprints :lol:

I'm glad you have so much faith in me to make it across the pond...but maybe you're right. Gotta go for style points B)

oh but I've got a lot of faith in you, Zach T......but style ain't worth much if you're sinking in the slush........but yup, at least sink with style...:P...and here's another fun carbon footprinting (with the crazy engineers):

The Great Northern Crazy Concrete Toboggan Race:

http://www.youtube.com/watch?v=CMaUrb9Y4kA

Haha thats awesome! I've always wanted to do some of these crazy events like the red bull one where they build the contraptions and then send them off the platform into the water, that would be fun. This summer, i'm doing the warrior dash where you have to jump over flaming logs and crawl under barbed wire :lol:



#105856 Help a couple of snowboard bums ;)

Posted by Zach T on 05 April 2011 - 12:59 PM in Chit Chat

At least they are fun carbon footprints :lol:

I'm glad you have so much faith in me to make it across the pond...but maybe you're right. Gotta go for style points B)



#105837 Help a couple of snowboard bums ;)

Posted by Zach T on 05 April 2011 - 11:16 AM in Chit Chat


Of course! :lol: gotta stay ahead of the times! Anyways, april 17th is the last day of the season so they always have the pond skimming challenge. So I think we are going to buy some cheap suits and ride in those for springfest! :P As usual, I'll take some pics.

Here is one of the other dude I was riding with:

http://www.flickr.co...tar/5590437733/

Quite the pose, huh?? :blink:

only snowboards and skis? so how long is the pond? and the basic rule of pond skimming: the tackier and the cheaper the costume, the better....and make a big splash, ok? :D

Idk, I've never done it at Steven's. I'd say its going to probably be about 30-40 ft. And i'm trying to stay dry, screw splashes.. :rolleyes:

Oh and at least I'm getting some street cred on another website...just not about my snowboarding, just about my car pics :P

Check it:
http://www.gtspirit....-car-show-2011/




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