Fish

[Ian Hawkins]

Do we have a complete protocol for glow in the dark bacteria? If so, where can I get the squid?

Thanks,

Ian Hawkins

617.784.9157 (m)

ianfh...@yahoo.com

Sent from my iPad

On Jul 7, 2011, at 6:36, diybio-bos...@googlegroups.com wrote:

  Today's Topic Summary

Group: http://groups.google.com/group/diybio-boston/topics

• Friday - replate of transformed E. coli stab. [3 Updates]

 Topic: Friday - replate of transformed E. coli stab.

Ian Hawkins <ianfh...@gmail.com> Jul 06 12:46PM -0400 ^

 
We didn't get any colonies on the new Amp IPTG LB plates, so I'm going in Friday around 7 pm to take a fresh sample from the refrigerated stab. I'll also be doing some brainstorming on modular small-scale biotech procedure automation widgets al la MakerBot toolheads.
 
Thanks,
Ian Hawkins
 
617.784.9157 (m)
ianfh...@yahoo.com
 
Sent from my iPad


 

Mackenzie Cowell <m...@diybio.org> Jul 06 12:52PM -0700 ^

 
I'll see you there.
 
Mac
 
 
--
+1.231.313.9062 / m...@diybio.org / @100ideas


 

Dakota Hamill <dko...@gmail.com> Jul 06 09:41PM -0400 ^

 
I'm heading into Sprout tomorrow to pick up the beer (if it's still
drinkable) as well as some Growlers, to bottle, then I'll be back Friday
night and bring you back a sterilized bucket and bubble chamber Mac. I
wanted to get in earlier but the convertible I was driving kept shutting
down on hot days from overheating, even with coolant, so I didn't want to
drive it in. Also couldn't figure out how to move 10 Growlers or a 5 Gallon
bucket back and forth on the T.
 
I'll bring in the saline agar plates material I have and pour some if it
makes sense, then if anyone wants to keep some and get their own fish for
the glow in the dark bacteria, go for it. It's another thing I've been
hanging onto for a year.
 
-Dakota


 

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[Avery louie]

I can help with that.  Will let you know after I get off work.

--Avery

[Mackenzie Cowell]

from my notes last time I tried:

13 Jan 2010

18:30 Got fresh, “uncleaned” squid from New Deal Fish Market in Union Square: $2.50

18:55: got 1L gatorade

Added 1/2 teaspoon of sea salt to the empty gatorade bottle

22:00 cut squid head in 1/2, added to tap water in gatorade bottle

14 Jan 2010

22:00 It GLOWS! wow!

Mac Cowell / @100ideas / +1.231.313.9062 / has100ideas.com /  DIYbio.org

[David Johnson]

mmmm.... glowing vibrio....


-Dave

[Jason Bobe]

I don't know if anyone listens to This Week in Microbiology, but last week they discussed the introduction of squid as a model organism and its relationship with vibrio, how it modulates light for camoflauge, etc.  

Check it out:

http://www.microbeworld.org/index.php?option=com_content&view=article&id=970:twim-10-a-symbiotic-cloaking-device&catid=107:this-week-in-microbiology&Itemid=275

Thx,

Jason Bobe 

[Dakota Hamill]

I'll bring the synthetic sea salt, peptone, yeast extract, glycerol, and agar if someone else can grab a squid.  I have a recipe written down for 500mL, which at ~ 20 mL per plate will use up the sleeve of 25 plates I have, or maybe the sleeve is 20 plates I don't know, but we'll be fine. Vibrio doesn't like the heat tho and Sprout's upstairs seems rather hot, at least for that strain from what I remember reading.  Cool stuff I can't wait!

I think I was looking at Cathal's recipe even though I've seen Mac's too just can't find it

http://letters.cunningprojects.com/wpcore/wp-content/uploads/2010/03/Homebrew%20Bacterial%20Media.pdf

http://letters.cunningprojects.com/?p=108

[Dakota Hamill]

I'm probably going to come in a little earlier, like 3-4pm maybe, and bring food and hang out for the night and drive home late to miss Friday night traffic, but look forward to a good day of science stuff!

[Ian F. Hawkins]

It took you an hour to drink 1L of Gatorade? :P

http://www.ncbi.nlm.nih.gov/pubmed/16750278 says 20 to 26°C. Not sure
how we'd accomplish that (high 70s°F).

I'll bring squid.

-Ian

[Dakota Hamill]

I bet downstairs in the library there are those temperatures, it was cooler downstairs.  Or a styrofoam beer cooler with an ice pack or two maybe

[Dakota Hamill]

I'm heading into Sommerville now, just have to go get a lawnmower and drop it off, probably be in there by 4:30 or so perhaps

[Ian F. Hawkins]

No uncleaned squid at the market today :(

[Avery louie]

I could get y'all a squid. You guys game? Let me know.

--Avery

On Jul 8, 2011 12:56 PM, "Dakota Hamill" <dko...@gmail.com> wrote:
> I bet downstairs in the library there are those temperatures, it was cooler
> downstairs. Or a styrofoam beer cooler with an ice pack or two maybe
>

[Ian F. Hawkins]

We are game.

[Dakota Hamill]

yeah!  I just got to Sprout I'm upstairs with all the stuff ready to make the saline broth.

[Avery louie]

Ok. I'll let you know if there are any in ~30 min.

--Avery

On Jul 8, 2011 4:28 PM, "Ian F. Hawkins" <ianfh...@gmail.com> wrote:
> We are game.

[Dakota Hamill]

cool, thanks.  i'll be here, going to just mix up the synthetic sea water mix

[Avery louie]

Ok.

[Avery louie]

Squid acquired!

[Mackenzie Cowell]

Avery got some.  He just arrived :)

On Fri, Jul 8, 2011 at 3:41 PM, Ian F. Hawkins <ianfh...@gmail.com> wrote:

No uncleaned squid at the market today :(

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[Dakota Hamill]

Well I had fun tonight, wish I got to hear Ian go over the stuff he took the time to write out on the board.  The plates are poured and labelled, squid is in the fridge, and Avery and I finished the little rocker thing for the tubes (he got the code and the servo and worked with the arduino, did most of the work!-- I just poked two holes in a paper plate and taped it on)  

The thing looks hilarious when it moves.  I got a little video but can't find the cable to connect to my camera.

Alright, keep in touch if anything glowy comes about, and I'll do the same.  Maybe Thursday we can reconvene on project night, or maybe sometime beforehand.

-Dakota

[Mac Cowell]

Lol!  Awesome!  Fingers crossed for Glowy!

Avery: please put the tube rocker code into the "code" folder in bosslab dropbox if you get a chance.

I'll swing by tomorrow to look for glowyness!

Great fun tonight.

Mac

231.313.9062 // @100ideas // iPhoned

--

[Mac Cowell]

Ps great photos here: http://bosslab.org/flickr

231.313.9062 // @100ideas // iPhoned

[cluckj]

That is really awesome. I hope they glow!

On Jul 9, 3:34 am, Mac Cowell <macow...@gmail.com> wrote:
> Ps great photos here:http://bosslab.org/flickr
>
> 231.313.9062 // @100ideas // iPhoned
>
> On Jul 9, 2011, at 3:27 AM, Mac Cowell <m...@diybio.org> wrote:
>
>
>
>
>
>
>
> > Lol!  Awesome!  Fingers crossed for Glowy!
>
> > Avery: please put the tube rocker code into the "code" folder in bosslab dropbox if you get a chance.
>
> > I'll swing by tomorrow to look for glowyness!
>
> > Great fun tonight.
> > Mac
>
> > 231.313.9062 // @100ideas // iPhoned
>
> > On Jul 9, 2011, at 12:27 AM, Dakota Hamill <dko...@gmail.com> wrote:
>
> >> Well I had fun tonight, wish I got to hear Ian go over the stuff he took the time to write out on the board.  The plates are poured and labelled, squid is in the fridge, and Avery and I finished the little rocker thing for the tubes (he got the code and the servo and worked with the arduino, did most of the work!-- I just poked two holes in a paper plate and taped it on)  
>
> >> The thing looks hilarious when it moves.  I got a little video but can't find the cable to connect to my camera.
>
> >> Alright, keep in touch if anything glowy comes about, and I'll do the same.  Maybe Thursday we can reconvene on project night, or maybe sometime beforehand.
>
> >> -Dakota
> >> --
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[Avery louie]

Hey guys, who is on squid duty tonight? Is it me?

--Avery

[Mackenzie Cowell]

I'll go in, I'm 5 min away.

[Mackenzie Cowell]

Here's the schedule from the whiteboard (http://www.flickr.com/photos/macowell/5916703441/in/photostream/)

Sat - photo squid: Mac

Sun - photo squid, streak first 2 plates: Ian
Mon - photo plates: Mac

Tue - replate glowing colonies from 1st plates onto new plates: Avery

Wed - photo plates: ? (unnecessary?)

Thu - inoculate liquid broth w/ glowing colonies: Avery + Dakota? + Mac

Check this schedule out and let the list know if you can't make a day.

Mac

[Avery louie]

Ok. I might stop by tonight

--Avert

[Avery louie]

When I stop by, it will be lateish, as it is laundry day :(.  Is there some way I can get in then?

--Avery

[Mackenzie Cowell]

yeah!  txt me!  I just arrived, i'm gonna email here for a bit, see if I can get my drunken-hack-foo going later.  231.313.9062

[Dakota Hamill]

Thanks for checking it.  There's nothing on the little piece I took, at least, nothing really visible.  Maybe I need to let my eyes adjust for a little while.  Perhaps it needs to be even colder then the basement.  

[Avery louie]

Any news on the squid?

[Avery louie]

I had to cook dinner tonight, so I will be over within an hour.

On Jul 9, 2011 8:00 PM, "Mackenzie Cowell" <m...@has100ideas.com> wrote:

[Dakota Hamill]

I was doing some reading and it sounds like it might be best to take it out of a plastic bag like that, because it can lead to smearing, whereas if its just resting in an open box the colonies can stay nice and tightly organized and won't get smudged every time you ruffle the bag closed.  just a thought tho, I'm hoping tomorrow i might see something on this piece of squid, otherwise i'm going to head down to the fish pier in gloucester and try and find something and put these 2 plates to use

[Nancy Ouyang]

(sidenote) my friend and i have had success getting nice glowing spots with cleaned squid (calamari) from new deal fish market (incubating with water+dash of salt covering most but not all of the squid, @ room temp for 24 hours, in tupperware). Phew that stunk though! 

new deal said next time we should call ahead and they can save some uncleaned squid.

we didn't have plates ready at the time and the spots seemed to have died after another 24 hours.

On Sat, Jul 9, 2011 at 6:56 PM, Dakota Hamill <dko...@gmail.com> wrote:

I was doing some reading and it sounds like it might be best to take it out of a plastic bag like that, because it can lead to smearing, whereas if its just resting in an open box the colonies can stay nice and tightly organized and won't get smudged every time you ruffle the bag closed.  just a thought tho, I'm hoping tomorrow i might see something on this piece of squid, otherwise i'm going to head down to the fish pier in gloucester and try and find something and put these 2 plates to use

--

[Dakota Hamill]

I'm about ready to just streak a plate and see if I can isolate glowing colonies from that, because if they don't show up tomorrow I don't think they'll show up, at least on my small little piece.

[Mackenzie Cowell]

It's not glowing yet.

photo set-up:

http://www.flickr.com/photos/macowell/5920714422/in/photostream/

Photo w/ reflections (15 sec f5.6 exposure):

http://www.flickr.com/photos/macowell/5920158125/in/photostream/

photo w/o/ ambient light (30 sec f5.6):

http://www.flickr.com/photos/macowell/5920736486/in/photostream/

When I did it (http://has100ideas.com/blog/culturing-bioluminescent-microbes-part-1), I incubated the squid for 1 day at room temperature (above my fridge).  I'm not sure if my culture was open or closed to the ambient air, but I think it was open.  The squid we have currently is inside a plastic bag, in a fridge.  The bag may be restricting airflow,  but it's also inside a refrigerator, which is also obviously restricting airflow and growth rate.

So we'll see what's up tomorrow (sunday night).  If nothing, than maybe we should take it out.

Cheers,

Mac

Perhaps we'll see glowing cultures tomorrow.

On Sat, Jul 9, 2011 at 10:20 PM, Dakota Hamill <dko...@gmail.com> wrote:

I'm about ready to just streak a plate and see if I can isolate glowing colonies from that, because if they don't show up tomorrow I don't think they'll show up, at least on my small little piece.

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[Ian F. Hawkins]

I'll be going in tonight around 9. If we have colonies, I'll streak
them on plates. If not... I'd like to incubate uncovered at room
temperature, but I'm not quite sure how to do that without stinking up
Sprout.

-Ian H.

[Mackenzie Cowell]

I think a tupperware sandwhich container would be just the right size.  We can leave the lid cracked, but not open.  That's what I did when I tried before.  I don't think it will smell too bad.

See you later

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[Mackenzie Cowell]

Also I think this is excellent grilling weather.  Anyone want to grill at my place with me around 6 or 7?

[Dakota Hamill]

Thanks for the invite, wish I was closer.  I had to throw out my squid just now, nothing was glowing and it smelled horrible and was filling three rooms in the basement.  I'm trying to work on a temp logger for a chemistry experiment about exo/endo thermic rxns and I'm trying to get my arduino set up again.  I found the code you used for the one at Sprout, and was wondering if you used that Pachube.com live data streaming site/software and if it was any good?  Also, did you just cover your temperature probe with some plastic/epoxy to make it water resistant?

Thanks, Dakota

[Ian F. Hawkins]

No Vibrio colonies, so moved squid + synthetic seawater to a plastic
container with the one edge left open.

No colonies on LB Amp IPTG plates. Probably used too weak of
Ampicillin concentration for initial selection. I'd like to order more
competent cells and a new RFP plasmid and try again; Mac wants to do a
kit.

[Mackenzie Cowell]

the squid saga continues: http://bosslab.org/post/7484301736/culturing-bioluminescent-microbes-from-squid

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[Avery louie]

we may need more air than that ...

[Mackenzie Cowell]

p.s. found some petri dishes + agar + ?? dated and labeled "LM".  These should have a more specific label and should go in the fridge.  I put them in the fridge.

p.p.s avery, where's my can'o'servos?

[Avery louie]

Lm. Is the ssw media for the v. Fischeri.  Can o servos ought be be back in the box

[Mackenzie Cowell]

I can't find the can o servos... next time you're at sprout have a look around.

thanks

mac

[Avery louie]

For sure.

[Dakota Hamill]

I just got some squid (cleaned) and divided it into two batches, one in regular NaCl and the other in that artificial sea water stuff-both in the refridgerator.  The tentacles still had the chromophore looking skin, but I got one uncleaned cap.  The market I went too didn't have any uncleaned.  I'll probably check them in 10-12 hours and put them down in the basement again where it's warmer but still cool.  I think I need to work on building my nightvision up too when I check for them.

[Avery louie]

I am going to go by sprout tonight and check the squid. Wil there be people there?

--Avery

[Avery louie]

En route.

--Avery

[Avery louie]

The squid was gone, but the smell lives on...

[Mac Cowell]

When were you there? Last night?

Mystery of the missing squid!

231.313.9062 // @100ideas // iPhoned

[Avery louie]

Yea...around 1030? unless it was hiding. Found the can o servos, it was in the bottom green box under the bench.

[Mackenzie Cowell]

did you look for the squid here: http://www.flickr.com/photos/macowell/5924697860/in/photostream ?

[Dakota Hamill]

The fresh (albeit cleaned) squid I bought yesterday is starting to turn a little milky and fresh smelling.  I read online the luminescence is density driven, but can also be circadian rhythm driven, so I'm giving them some ambient light time today at room temp.  I have a feeling they are just going to turn rotten again and I'll be left with a nice squid stew.  Maybe the broth, even if it's low level, keeps washing over the surface and taking any colonies with it.

How come it seems so easy when I read it online on peoples writeups?

[Avery louie]

Where does it say anything about circadian rhythms? I would like to read about it.

[Mackenzie Cowell]

Hello all,

Some people at sprout didn't like the smell of the squid, so they threw it away.  Also they are mildly annoyed with us.

Don't despair, third time's the charm.

Mac

[Mackenzie Cowell]

Wikipedia has the first clue:

http://en.wikipedia.org/wiki/Vibrio_fischeri#Bioluminescence

The luminescence appears to follow a circadian rhythm. That is, it is brighter during the nighttime than during the daytime.

But is it really a circadian rhythm?  A google search for "vibrio fischeri circadian rhythm" turns up some interesting papers.  I would strongly recommend the minireview "An Exclusive Contract: Specificity in the Vibrio fischeri-Euprymna scolopes Partnership": http://jb.asm.org/cgi/content/short/182/7/1779.  It's awesome.  It tells the fantastic story of how V. fischeri and a particular hawaiian squid, E. scolopes, manage to meet and work together.

Basically, E. scolopes has special tissue structures just for the V. fischeri.  Within hours of a baby squid's birth, it has found and nurtured a culture of V. fischeri.  It apparently can protect its bioluminescent "crypts" from microbes besides V. fischeri with a variety of defenses and countermeasures.  Interestingly, the V. fischeri are thought to bind to mannose sugars on the microvilli that line the crypt epithelium.  This makes me wonder, could we get V. fischeri to stick to arbitrary surfaces in salt water by coating them with mannose?  If we ever figure out how to culture them, we should try that out.

Anyway, back to the putative circadian rhythm.  Well, the authors of the minireview mention that circadian rhythm-like activity has been noted in the bioluminescence of S. scolopes bioluminescence organs.  But they clarify that experiments have shown that the cycle is not truly an "entrainable" circadian rhythm.  Although the cycle does seem to be light-dependent (changes in the light/dark cycles seem to cause changes in bioluminescnece), as of 2000, no one seems to be totally sure how the control actually works.  It could be controlled by oxygenation of the water, it could controlled by some small molecular, or other nutrients... more research and lit review will tell.

I've copied the relevant text from the minireview for you to read below.  Again, I strongly recommend reading the whole thing - it's very entertaining.

So what does this mean for us?  Well, we should probably figure out the species of squid from which we are trying to cultivate the v. fischeri and determine if it has it's own light-organs.  If so, we might have better luck directly sampling them.

Cheers

mac

---

Each morning, beginning with the first dawn following colonization of the organ, the squid vents 90% of its crypt contents into the body cavity (Fig. 5A), and the ventilatory movements of the squid carry this material out into the seawater (19, 31). The remaining 5 to 10% of the bacteria multiply and repopulate the light organ over the day. This behavioral pattern is not an entrainable, or circadian, rhythm; the animal responds each morning directly to the cue of an increasing level of ambient light (5, 31). Host-controlled muscular pressure imposed on the crypts results in the release of a thick exudate (Fig. 5B) containing not only bacteria but also macrophages, epithelial cell debris, and the protein-rich matrix of the crypt spaces (12, 31). This expulsion event may serve several functions. For example, this behavior may be a means of controlling symbiont number: while V. fischeri is not considered a pathogen, unchecked bacterial multiplication might harm the host. Also, it may be important for the squid to maintain a fresh culture ofV. fischeri cells. V. fischeri cells grown in culture induce luminescence in a density-dependent (autoinducer-dependent) manner (3). Coincident with stationary phase in culture, however, the level of luminescence drops. Furthermore, V. fischeri cells exhibit poor stationary-phase survival in the laboratory. Because bioluminescence is unnecessary in the daylight hours after dawn, during which time the animal is buried in the sand, expulsion of the bacteria may serve to maintain a healthy culture of bacteria that will be ready to bioluminescence the following evening. 

[Dakota Hamill]

Hey, thanks for the email and synopsis Mac.  Avery, When I mentioned the thing about the circadian rythm, I got it off the wiki Mac linked, but never followed up on it.  

That's crazy that perhaps the squid has evolved to flush out the  V. Fischeri just so the new fresh batch is ready to glow the next day!!  Avery even mentioned this problem, as does the paper, that "stationary phase culture" leads to the luminescence level dropping and then shutting off.  Nature rules.

I tried a few basic google searches but I don't know the species of squid we are getting, because I had the same exact type Avery bought, and by the looks of it, the same type Mac bought a while back.  I'll ask my buddy tomorrow who is a commercial fisherman if he's back, supposed to be back tomorrow or Thursday.  I've been meaning to ask him to bring a fresh fish back, as he says he sees crazy things brought up out there, from tropical fish to half rotten human bodies.  

Check out this badass fungus from the bioluminescence page,  http://en.wikipedia.org/wiki/Panellus_stipticus

What I want to know is what makes the water glow when I go to the beach in the summer and stomp on the sand?  Is it those dinoflagelites that you were talking about Avery?  Guess so according to this wiki.  Although all the ones I see are green not blue, but maybe the camera causes a color change because all the other ones online are blue too.  

http://en.wikipedia.org/wiki/Noctiluca_scintillans   

Before I threw the squid out I took a sterile swabber I had and just plated a rubdown of the squid solution--just to see if anything grows, or glows, or both.

Haha I bet it smelled great upstairs in Sprout with 100 degree temp and 100% humidity, but all in the name of science

[Dakota Hamill]

I swabbed some squid juice onto a plate last night, and stayed up until sunrise this morning, so I actually saw the bacteria grow in couple hour increments, pretty cool.

I knew I should have streaked it into 4 quadrants to get individual colonies, now I just have a big stinky smear that smells like dead squid still.

http://imageshack.us/f/52/img0026zb.jpg/

[Mac Cowell]

Keep it incubating in the light, see if there are any glowing patches tonight.

231.313.9062 // @100ideas // iPhoned

--

[Dakota Hamill]

This'll be a double post but for the sake of keeping this long thread going.

The plate I did yesterday alas has no glowing colonies.  But I did take a small sample of that plate, and replated it on the 2nd plate and did a 4 quadrant streak to maybe get down to single colonies on the 2nd plate.  My buddy just got back from fishing and he said he might be able to get really fresh stuff right off the boat tomorrow.  I shall not rest until V. fischeri is ours!

[Dakota Hamill]

Found a good thread about this on sciencemadness

http://www.sciencemadness.org/talk/viewthread.php?tid=1485&page=2

decent posts start a couple posts down

Seems people have success with liquid media, as well as really fresh fish, and the fridge for a couple days.  I don't know, it seems hit or miss, getting them at the right time and with the right amount of oxygen.

attached the.pdf i pulled off that thread

[Mac Cowell]

Good PDF.  Sounds like closed incubation at 7-14C for 72hrs could work.

I also am interested in looking for the luminescence crypts and harvesting directly from there, if possible.

What exactly is in the aquarium salt water mix you have?  Could you compare it to the specified recipe?  Any antibiotics?

Mac

231.313.9062 // @100ideas // iPhoned

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<Luminous bacteria.pdf>

[Dakota Hamill]

I went downstairs just 5 minutes ago  after having been outside all night, and no glowing colonies even with maximum nightvision.  I don't even know what bacteria is growing on the plates, but they all look to be the same type.  I think oxygen has a big part to play as well, and one of the successful stories I remember reading about was a fish in a big white Styrofoam beer cooler partially submerged in the saline mixture.

Here is the salt stuff we used.

http://www.petsmart.com/product/index.jsp?productId=2754897

I remember reading it after to make sure it didn't have antibacterial agents etc. but it didn't.  What you see on the description in that is the same thing written on the back label.

I don't think it's the culture medium, it's just squid that don't have the bacteria present in good numbers, or the incubaction technique.

I'd think the bacteria ARE on there, we just need to make sure they grow better then the other bacteria that are also on there.

That either means;

1. Refrigeration undisturbed for 2-3 days 

2. Culture in a liquid media with shaking/disturbance  check for luminescence. 

3.   Keep buying squid and fish until one of them produces nice colonies.


I'm going to soak the plates I have in bleach for a while and then toss them, maybe see if I can see anything in the microscope.

[Avery louie]

Im going to pick up some dead sea-critters today/tm.  I will find somewhere to incubate and see if I get glowies; if I do, may I plate them?

--Avery

On Jul 15, 2011 1:23 AM, "Dakota Hamill" <dko...@gmail.com> wrote:

[Mackenzie Cowell]

plate em!

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[Avery louie]

Got mo squid.

[Avery louie]

Notebook entry 9:00

Incubated squid on the red line...

[Avery louie]

I got bright, small colonies. Will plate today.

[Dakota Hamill]

huzzah

[Mackenzie Cowell]

photo!

what's the "9:00 notebook entry" all about?

On Sat, Jul 16, 2011 at 12:50 PM, Dakota Hamill <dko...@gmail.com> wrote:

huzzah

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[Avery louie]

That was facetious.  Hess to lab now.  Worried it wil be too bright to see colonies/they wil get smeared en route

[Dakota Hamill]

guard them with your life

[Avery louie]

Plated, in the fridge!

On Jul 16, 2011 1:35 PM, "Dakota Hamill" <dko...@gmail.com> wrote:
> guard them with your life

[Avery louie]

It grew...bigger.  After a few hours outside, the light from the squid is brighter.  Im going to try to photograph it.  I am leaving the plates out at the moment

[Avery louie]

Epic pix

[Dakota Hamill]

the suspense is killing me, where are they?

[Avery louie]

I took done pix out of range of the I fi, how do I get them off the card? They are on the camera...

On Jul 16, 2011 6:45 PM, "Dakota Hamill" <dko...@gmail.com> wrote:
> the suspense is killing me, where are they?
>

[Avery louie]

Fixed problem.

[Mac Cowell]

You have to leave the camera on for 30 - 60 sec

231.313.9062 // @100ideas // iPhoned

[Avery louie]

Took pix in RAW, going to take card to home computer to try to extract them

[Avery louie]

Will bring back to sprout tm morning.

[Avery louie]

Pix should auto upload to flkr

[Avery louie]

Leaving squid to incubate a bit more at rt

[Avery louie]

Plated once again; no growth on previous plates, which I find strange; you would think something would grow, our maybe I'm just pro at sterile techniques... Threw away squid, as most of it had stopped glowing, and the smell was growing. May check back tm to see if there is any growth. Left plates next to tablet.

Other thoughts: if we boil/blend some squid up, do you think we could use the liquid for media?

[Avery louie]

If no growth by mon night, reccomend we ought to get more squid.

[Mackenzie Cowell]

where are they incubating?

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[Dakota Hamill]

Wooooo awesome.  A couple of those pictures are really really amazing, that squid was covered!

It might be interesting to try some glowing colonies in a couple liquid media tubes, if that's still sterile and still sitting there.  It seems (from some pictures online) they glow the brightest in liquid media with some agitation for that extra kick of oxygen.  I'll be in to Sprout Monday or Tuesday to hang out and maybe donate some DNA to the Germans, and to hand out some beer Growlers.  I tasted the beer yesterday, and it's actually good.  When I tried the "wort" or whatever it was after 2 weeks of fermenting, it tasted like sour rotten poison, but it tastes like beer now.  I even took a video of the yeast in the microscope, a lotttttttt of yeast, you can't even see through the beer, and I was careful not to suck up any of the crud on the bottom too.  Oh well that's another story.

[Avery louie]

On the counter in front of the tablet

[Avery louie]

If anyone is in the lab today, please check and see if there is any growth on the plates.

[Dakota Hamill]

I was going to head in there to meet those German kids at 1 pm but I only have my car until 5:30 or so and it seems they are coming back at 9pm so I'm out of luck.  I still need to offload some of this beer.

  It sounded like another person or two were interested in coming because of that back pain genotyping thing, maybe they will stop by for another meeting.  Although, the Ginger beer/soda making thing Ian is doing is this Friday right?  So perhaps that's when I'll bring everything in.

[Mackenzie Cowell]

I checked the plates today and took some long exposures.  No glowing.  So I picked one the of the colonies and am trying to do PCR on a 16s region so we can do species identification w/ sequencing.  I'll let you know how it goes

On Tue, Jul 19, 2011 at 11:57 AM, Dakota Hamill <dko...@gmail.com> wrote:

I was going to head in there to meet those German kids at 1 pm but I only have my car until 5:30 or so and it seems they are coming back at 9pm so I'm out of luck.  I still need to offload some of this beer.

  It sounded like another person or two were interested in coming because of that back pain genotyping thing, maybe they will stop by for another meeting.  Although, the Ginger beer/soda making thing Ian is doing is this Friday right?  So perhaps that's when I'll bring everything in.

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[Avery louie]

Mac, I doubt they are a single species in that colony yet. Pcr will probably turn out poorly

[Mackenzie Cowell]

why do you think they are not single species colonies?

[Dakota Hamill]

new pics on flckr look cool.  Were you trying different DNA stains or something in that gel?  Those plates look similar to the plates I had, milky white smear of bacteria to milky white single colonies in the 4 quadrant smear.  I imagine a lot of bacteria have colonies that look like that though.  Be interesting to see if it is Vibrio or just E.Coli or something else that took over

Have you done the genotyping from the German's thing?

[Avery louie]

They came from a mixed sample and could very easily be a mixed colony.

[Dakota Hamill]

Google gave this picture of the unglowing colonies  http://www.biology.pl/bakterie_sw/bac_mf_en.html

so might be able to have a look at their morphology.  This nice Cannon camera I have doesn't seem to have a long exposure, which is crap.  I spent 30 mins hitting every button and reading through the manual.

I don't know very much about bacterial morphology or colony  phenotyping or whatever you call it, and my microscope I do not think would be of much use since I don't have any oil or cover slips for a 1000x zoom.  I'll upload what my plates look like in one minute

[Dakota Hamill]

Here are some pics.  Notice in the 1st and last one, there seems to be 3 different colonies (or at least 2 )

The one on the far left, tannish color, middle white color, right, browner color

http://imageshack.us/g/13/img0077hb.jpg/

[Dakota Hamill]

Also they are High Def not resized so the zoom in makes them really nice and easy to look at up close