2014

Listen while you look:





JANUARY

CHROMAWORLD

touché amoré
photo by honolulu pulse

LOVELY LADIES


FEBRUARY

Dad's Catch of the Day 2/27/14

Oversplit!


MARCH

Love Like Winter

I don't have a cat



APRIL

Aspergillus niger

MAY

F I S H B O N E S

t o e - t o u c h

JUNE

Tail Gating

Hanapa`a!

5:52 AM


JULY








AUGUST








SEPTEMBER





OCTOBER

THAT OLD SKOOL LIFE

playing in the rain

NOVEMBER

Koundinyasana

25 miles a-round

DECEMBER

IMPOSSIBLE R

SHOOT ME IN THE SMILE

everything else from 2014: https://www.flickr.com/photos/synonyme_c/archives/date-taken/2014/
and onwards to 2015!

GIVE WHAT YOU GET, AND YOU GET WHAT YOU DESERVE

here is the most recent friday5!
answer for yourself or see others answers here: http://f.riday5.com/?p=1116
sorry, guys these answers might be a little off point.  i like coincidences but don't believe in luck.  fuck luck, make things happen with hard work!

  1. When has something miraculous or seemingly miraculous happened to you or someone you know?
    my coworker had her layaway paid off by a stranger last week.
    kindness does exist. :)
  2. When were you last part of an amazing coincidence?
    i helped my pops wrap his secret santa gift for a friend.  he gave him ten packs of plastic glitter fishing lures and a bag of lead attached to a fishing hook.  he said, [lance] is going to love this.  i would even keep it for myself. 
    he came home that night with a gift from his secret santa.  the box got smashed in his backpack on the way home so he decided to open it... 
    his secret santa gave him 5 packs of the same plastic glitter lures, a bag of lead, and a bag of red beads for tying the lures.
  3. What have you recently accomplished against magnificent odds?
    i forgot to take one of my final exams.  
    not joking.
    it was for an online class and i had finished all my homework a week before and i totally forgot i had to take the final.  it was already past due and i was thinking, i have three exams this week.  one for every lecture... except i had four lectures... FUUUUUUUUUUUK...
    i emailed the professor right away.  i didn't think he would let me make it up but i decided to just let him know what happened.  
    his response was: without the final your grade is 91%  study for your other classes and have a nice winter break.
    yes guys, doing homework on time, all the time pays off.  study hard and you can still get an A without taking a final exam. OTL
  4. In what way this week has luck been in your favor?
    i was looking for a shirt for my sister's xmas present and couldn't find her size at ala moana.  i went to kahala and they had it there, hooray!
  5. What was the last outcome you consciously decided to leave up to chance?
    sunday at 4pm: I wonder if Nam Fong is still has duck left?  i will just go down and see... 
    (they weren't even open)





this post is in memory of Dr. Alam.




http://bdnews24.com/bangladesh/2014/12/21/jute-genome-scientist-maqsudul-alam-dies
i just heard the tragic news last night at our research group's christmas gathering.   i feel like i just saw him!  and even more unsettling, his remarks at the ASM meeting "which one of us will be next..." (which i wrote about here: http://www.liquoriceblack.blogspot.com/2014/10/electric-hum.html).  he was a genomic genius and had even more big ideas for the future.  he was known for sequencing projects, quorum sensing/pathogenicity islands, bioinformatics... the list goes on.  the last nice thing he did affecting me was paying for our virology class' time on the scanning electron microscope.  usually it's around $100/hr plus materials for sample preparation but he paid for our time at the facility and the ladies there let us take as many pictures as we want (and also let us drive around on the scope).  me and EmM and heathers and suz had such a good time!  i'm sure any of us would jump at a chance to do work or intern there or just sit at the scope and operate it again! here's a picture i took there:

"pickles" (bacteria SEM)
yes!  i actually took this!  it's bacteria from an oil pipe.  magnification is 18000x!

R.I.P.



besides the shocking news, we had a lovely time at our party.  lots of old and new faces congregating for food and fun.  yes, we talked about science and school and poop and med school interviews, but that's expected from bacteria scientists, no? ;)  it was a great!  super chill time, great food, talk story, and i also discovered sierra nevada coffee stout.  http://www.sierranevada.com/beer/variety-packs/coffee-stout




cheers!




VARIATIONS ON A THEME / REPLACEMENTS

my first impression of this song was

????????






but i want to... hear more?

so i listened again...
and again
and a few more times...

it grows on me.


i want to like it so bad because patrick kindlon is fucking funny!




"obviously brush your motherfucking teeth." 


but this song is really weird.
and the video is not exactly helping either.
i think... i have to listen to it a few more times...


------------------------------


i sort of like the replacements.  they have some good songs, but i could only name one off the top of my head (unsatisfied).  not sure if i'd go their concert if they ever showed up in hawaii.  but then i heard them playing LIVE on Jimmy Fallon: 


woahhhhhh, they sound really good live!  better than their recordings!  i would totally pay to be in the front row of a show like this!  

SO THE STORY GOES

hi!  friday 5 time! but, i'm answering last week's friday5, BECAUSE I CAN.
deadlines-schmedlines!  truthfully?  i was looking at the wrong post, haha.  i didn't notice the date was wrong, oh well.  i'll do this week's questions in another post. later. (maybe.)
answer for yourself or see others' answers here: http://f.riday5.com/?p=1113
and if you want to do THIS week's friday5 like an observant smarty-pants person would: http://f.riday5.com/?p=1116

  1. You are awake at 2:30 in the morning on a work night (or school night). What is the most likely reason?

    because I AM at work. ;D

  2. Time Magazine says you are a candidate for 2014’s Person of the Year. What is the most likely reason?

    "the most average person in the united states who happens to have strange and interesting hobbies!"

  3. Your hometown is naming a geographical or civil landmark after you, as in Mary’s Creek or David’s Corner. What is the most likely name of this spot?

    i actually do have a favorite spot on this island.  favorite spot in my whole known world, i suppose.  it's past baby makapuu before you get to makai pier, there is a stack of concrete barriers against the sand bank below the road.  you can climb up and sit here and look out on the ocean... 

    a feeling like flying..

    Vire au vent tournoie deploie tes ailes

    i would call it, codi's perch. :)

    if you are my friend i will take you to this place!

    iPod photo dump

    Au loin ton echo s'eloigne


  4. Your best friend from high school calls to ask a favor. What is the most likely nature of the favor?

    lately i've been getting a lot of:
    help, what do these lab results mean?!  or
    help, i need to borrow insert cooking utensil/tool here !

  5. Twenty-four hours from now, you’ve got a great smile on your face. What is the most likely reason for this smile?

    ye-ah, it's winter break! 

    weeeeeee!


some holiday cheer:



and a more musical version:



both make me equally :) 


I JUST WANT TO CATCH THE LAST LAUGH OF THAT SHOW

today i was at don quijote waiting in looooong check out line and the old lady in front of me was trying to chat me up: 
what does your shirt say? 
(la dispute) 
your basket looks heavy do you want to put it down? 
(microwave popcorn, choco pie, coffee. my diet is embarrassing.) 
this music is weird, don't you think they should play christmas carols? 
and i perked my ears up to hear... MODEST MOUSE! 


and i just smiled and said to her "well, it could've been would've been worse than you could ever know!" 

to play this particular song in the store... wow.  don quijote confirmed itself as weirdest grocery store in honolulu!  but i don't think it's a bad thing; you know how much i love weird stuff.  and it used to be worse!  with the bird sounds and fake plants (those are gone now, probably a fire hazard)!  i think they still have the giant roll of toilet paper hanging from the ceiling near the fruits and vegetables.  holiday mart/daiei used to be a nice normal store now it's like sensory overload while shopping! 

she sort of nodded so i continued, 
"the dashboard melted but we still have the radio..." 
now she is giving me a puzzled look. 
"we schemed and we schemed but we already blew it, we've yet to crash but you might as well stow it..." 
she turned around and stopped talking to me.  hahaha. 

BONS SONS 2014

things i heard and liked this year:

la dispute- first reactions after falling through the ice (rooms of the house)



but really, the whole album fills my heart with joy. my pick for album of the year!!
so good i have to give you another one from the album!:



and one more for good measure:



closely followed by

phantogram- the day you died



tycho- see (awake)



interpol- all the rage back home (el pintor)



the raveonettes- summer ends (pe'ahi)



ghost feet- Give me something (single off gem drops 4)



i'm partial to this remix because we did our aerial performance to it!  i still think in certain parts "straddle up, eagle wrap, hold..."
tove lo- habits (stay high) | hippie sabotage remix



shimmering stars- shadow visions (bedrooms of the nation)



i have to admit i was a super weezer fan for a long time but started losing interest around 2003. i didn't even listen to their last two albums out of disappointment.  but this single gives me hope!  it's like singing about In the Garage with the voice of Don't Let Go.
weezer- back to shack (everything will be alright in the end)



vales- dead wood (wilt and rise)



this single came out over a year ago but the full album finally arrived in 2014!
flight facilities- claire de lune (down to earth)



another single that came out in 2013, full album released in 2014 :D
lily allen- hard out here (sheezus)




a late breaker!  nearly missed the list!
mogwai- teenage exorcists (music industry 3 fitness industry 1)




CONSPICUOUS CONSUMPTION

i went to a great gatsby themed christmas party on the weekend.
(i don't have instagram but if you do, you might search #gatsbystraub for an idea of what it was like)

thanks to the instruction of Ms. Coffee Sunday, I managed to look like a nice person with makeup and stuff.  since she dedicated a post to me, i'm returning the favor here!

this is the stuff i used


  • liquid eyeliner from Faceshop (i have so much black eyeliner, this was just the first one i grabbed)
  • Falsies flared mascara (because i don't curl my eyelashes)
  • kat von d beethoven palette (my favorite palette)
  • a brush from target (because that's the only one i have)
  • kat von d hell bent (because i don't get to wear red lipstick often enough in regular life)

and a before shot:


(back lit, hand held, wide angle, SORRY FOR ALL THE PHOTOGRAPHY SINS!)

additional challenge: i have one double eyelid and one single so making things look balanced means putting more make up on one side of your face than the other, haha. 


first, eyeliner:


and then i went with the darkest purple:



then i went over everything with the gray-ish purple:


(hard to tell, i guess i should have squinted again)


mascara, lipstick and headband!




please check her blog for more tips, color swatches, humor, rants, outfits, cute pocky puppy, and coolness. http://coffeesundays.com

pre-party (a.k.a. ignoring stares while riding TheBus) music:



Anakin sounds awesome!  full album coming to no sleep records in 2015 !!!




P.S.
the dress.


i also had a sweater because the top half of this dress is too scandalous and i couldn't wear those heels besides for this photo due to my left toe injury.  and don't ask how much the dress was.  it was sort of a birthday splurge for myself and fuck if the laboratory wasn't going to be the most well dressed, bad-ass bitches department of the whole hospital party.


* bling ! *

DRIFTED LISTLESSLY THROUGH THE VELVET BLACKNESS OF OBLIVION

i just discovered DEAFHEAVEN !!!



what a great album!!  the sound is so beautiful.
it's often described as black metal shoegaze (so... blackgaze?) and i couldn't agree more.
sunbather was released in 2013 so too late for my best of 2014 list.  but they did release a single via adult swim this year:



download all the singles free here: http://www.adultswim.com/music/singles-2014/

i have the flu. :'(
the body ache, chills, nausea kind of flu.
and then i watched this video :



and i think my eyes exploded with mild vertigo. (holy crap is this game for real?! i can't tell what's going on at all here...)

so, i'm just going to listen to sunbather under the covers with my eyes closed and a cup of coffee and try not to think about impending final exams.

Determining localization of Dengue viral protein, NS1, in eukaryotic host cells by immunofluorescence

ABSTRACT
HEK 293T cells were co-transfected with Dengue viral protein, NS1, and various intracellular markers.  NS1 was conjugated pre-transfection with protein reporter, HIS/V5, while intracellular markers were conjugated to GFP.  Fluorescent labeled antibodies are used to detect the transfected proteins and are visualized by confocal microscopy under immunofluorescence.  Interaction between NS1 and intracellular structures can be hypothesized when the proteins are in close proximity however additional tests need to be performed to confirm this. 
INTRODUCTION!
Dengue is a single strand, positive-sense RNA flavivirus with four serotypes transmitted to humans by mosquito vectors.  Severe pathology in humans consists of malaise, fever, hemorrhaging, and can lead to shock and death.  Understanding the cause of its pathogenicity is key to development of dengue treatment and vaccine.
NS1 protein is a well-studied protein in the virology field. In the dengue virus, the non-structural proteins are named NS1 through NS5.  NS2 and NS3 genes are found to code for protease and helicase.  NS5 is the RNA dependent RNA Polymerase.  NS1 and NS4 have unknown functions.  Despite its unknown function, NS1 is an important protein because it can be used in diagnosing viral infection.  NS1 levels in serum are elevated at viral infection as it is secreted as a soluble hexamer from infected host cells.  As previously mentioned, the function and method of NS1 secretion is also unknown, however a hypothesis is proposed; monomeric NS1 (mNS1) protein is glycosylated and prepared in Endoplasmic Reticulum (ER) vesicle packets (GPI-NS1).  From the ER it is either passed to the Golgi where it takes hexameric form (sNS1) and then secreted as a soluble protein, sNS1-LP, or is associated with lipids and delivered to the cell surface by cholesterol/lipid rafts where it becomes a membrane associated protein. 
This experiment uses immunofluorescence to visualize a protein marker, HIS/V5, conjugated to NS1 in host cells.  Proteins of certain organelle-associated proteins are conjugated with protein markers to visualize in conjunction with NS1 protein.  Fluorescent-labeled antibodies against these proteins are added to detect the proteins’ presence in the cell.  If NS1 is synthesized and secreted as hypothesized, we expect co-localization of NS1 in the host cell with ER, Golgi, and in the plasma membrane as it is being prepared for release from the cell.

MATERIALS AND METHODS
            A 60% confluent culture of HEK293T cells on cover slips were transfected using Polyfect, a transfection kit optimized for transfection with small lipid particles.  Prior to transfection, genes for NS1 protein were conjugated to genes for HIS/V5 protein marker while known intracellular protein genes were conjugated to green fluorescent protein (GFP) genes.  See Table 1 for list of conjugated proteins used.  Polyfect reagent and conjugated proteins were incubated together to form complexes between the lipid and the proteins were then added drop-wise to cell cultures.  Gene expression in cells took place over 24 hours at 37ºC.  After transfection, cells were fixed with 3.7% paraformaldehyde.
            For the immunostaining fixed cells were first blocked with 5% BSA, then monoclonal antibodies were added: mouse anti-V5 IgG to detect NS1 protein and rabbit anti-GFP IgG to detect cell proteins and incubated for 48 hours at 4ºC.  Cells were then washed 3 to 5 times with phosphate buffered saline (PBS).  Secondary antibodies conjugated to fluorochromes were then added.  See Table 1 for list of antibodies used.  Cells were incubated with secondary antibodies for 45 minutes, protected from light, at room temperature. Cells were washed 3 to 5 more times with PBS to remove excess antibody and reduce background.
            The cover slips were then removed and dried.  A nuclear stain, DAPI, was applied and then the cover slips were attached to glass slides with low viscosity enamel for viewing.  Confocal fluorescent microscopy was used to visualize cells.

Table 1: Conjugated proteins used for cell treatment
Protein and conjugate pairings
Primary Antibodies
Secondary Antibodies with fluorescent conjugate (color at excitation)
NS1-V5 + ARF1-GFP
Mouse anti-V5 +
Rabbit anti-GFP
Goat anti-mouse Alexafluor 555 (red) +
Goat anti-rabbit Alexafluor 488 (green)
NS1-V5 + FAPP1-GFP
Mouse anti-V5 +
Rabbit anti-GFP
Goat anti-mouse Alexafluor 555 (red) +
Goat anti-rabbit Alexafluor 488 (green)
NS1-V5 + SEC31-GFP
Mouse anti-V5 +
Rabbit anti-GFP
Goat anti-mouse Alexafluor 555 (red) +
Goat anti-rabbit Alexafluor 488 (green)




RESULTS
ARF1 is a protein that is associated with the golgi.  ARF1 exits the golgi and either returns to the ER or shuttles proteins to the plasma membrane.  NS1 proteins will be associated with vesicle proteins as it is moved from ER to golgi for hexamerization and from golgi to plasma membrane for secretion.  If NS1 becomes a membrane associated protein it will be transferred by cholesterol/lipid raft to the cell membrane.

Figure 1: NS1-V5 and ARF-GFP labeled HEK 293T cells
(L-R): DAPI & ARF-GFP & NS1-V5 | ARF-GFP | NS1-V5
In Figure 1 the ARF proteins are indicated with green color and NS1 is indicated with red color.  In the merged image co localization is visualized by yellow color produced by overlapping green and red light.


FAPP1 is a protein that localizes in the golgi where lipid synthesis takes place.  After GPI-NS1 leaves the ER in vesicles, it is either associated with lipids and taken to the cell surface to become a membrane associated protein or processed in the golgi as a hexamer to become a soluble secreted protein.  Two NS1 treatments were performed.  Figure 2 shows FAPP1 in green and NS1 in red where the host cell has been premeablized before fluorescent antibodies were applied.  In Figure 3, cells were not permeablized before antibody application in order to visualize the NS1 in the plasma membrane as a membrane associated protein or congregating before secretion. 




Figure 2: NS1-V5 and FAPP1-GFP (permeable)
DAPI & FAPP-GFP & NS1-V5 | FAPP1-GFP | NS1-V5
In Figure 2 golgi associated protein, FAPP1 is green.  There is a lot of background protein in the cell but the golgi is prominently shown as a large green mass.  NS1 has entered the permeablized cell but is not co-localizing with FAPP1 proteins as evidenced by the lack of yellow color in the merged image. 


Figure 3: NS1-V5 and FAPP1-GFP (non permeablized)
DAPI & FAPP-GFP & NS1-V5 | FAPP1-GFP | NS1-V5
In Figure 3 golgi protein FAPP1 is indicated by green color and NS1 is indicated by red color.  These cells were not permeablized before antibody addition.  Therefore we do not expect co localization of the two proteins as there are not golgi proteins in the plasma membrane. 


The third cellular protein used is SEC31-GFP.  SEC31 protein localizes in vesicles exiting the ER.  After mNS1 is glycosylated in the ER (GPI-NS1) it is taken by vesicles to the golgi for hexamerization and association with lipids.  Two NS1 treatments were performed.  Figure 4 shows SEC31 in green and NS1 in red where the host cell has been premeablized before fluorescent antibodies were applied.  In Figure 5, cells were not permeablized before antibody application in order to visualize the NS1 in the plasma membrane as a membrane associated protein or congregating before secretion. 


Figure 4: NS1-V5 and SEC31-GFP (permeable)
DAPI & SEC31-GFP & NS1-V5 | SEC31-GFP | NS1-V5
In Figure 4 merged image, there is one cell with a large amount of cytoplasm and strong NS1 signal.  The yellow dots indicate areas of co-localization of NS1 in SEC31-GFP stained vesicles.  Compare to the cell to the left in the merged image where there is no co-localization.


Figure 5: NS1-V5 and SEC31-GFP (non-permeablized)
DAPI & SEC31-GFP & NS1-V5 | SEC31-GFP | NS1-V5
In Figure 5 the non permeablized cells show no co-localization [yellow] areas.  This is expected because vesicles are located intracellular, in the cytoplasm between the ER and the golgi, but the labeled NS1 is restricted to the plasma membrane of the cell, where there are no vesicles.


Figure 6: HEK293T Cells Infected with Dengue Virus
A positive control was performed.  Figure 6 shows HEK293T cells that were infected with Dengue virus instead of transfected with viral protein.  DAPI is also applied for nuclear identification and is shown in blue.  ER is shown in green and NS1 protein is red.  This control demonstrates NS1 protein is present in infections. 

DISCUSSION AND CONCLUSION
In the non-permeablized cells there was no co localization of the NS1 protein and the labeled cell structures which was an expected result.  In non permeablized cells, antibodies are unable to get into the cell so only NS1 on the plasma membrane is labeled.  This demonstrates that NS1 is not only found intracellularly but on the surface and extracellularly as it is secreted.  In the permeablized cells there was co localization of the NS1 proteins with the cellular structures, as evidenced by the yellow color produced by the overlapping red and green light.  However, not much co localization is seen with NS1 and FAPP1.  Since FAPP1 is associated with the golgi this could indicate NS1 does not go to the golgi for synthesis as much as it goes to other areas.  However, NS1 is a known secreted protein and would require some assembly at the golgi for this to happen.  The lack of visible co localization could be from too much background from either fluorochrome masking the others’ color or not enough protein aggregated in one place to be visible.  Errors in transfection causing this problem is not likely because the transfection was preformed at an off site lab proficient in viral proteins.  Controls indicate the antibodies are satisfactorily labeling proteins as well.  A human error or factors concerning excitation of fluorochromes and the background proteins, as previously mentioned, may have caused this aberration.  This experiment may be repeated to confirm the lack of association since other results confirm a secretion pathway from ER to golgi to cell surface. 

This experiment shows only the localization of NS1 protein and does confer actual interaction between the organelle and the viral protein.  To confirm interaction, additional assays would have to be performed such as an Eastern blot on transfected cells to detect lipid or carbohydrate modified NS1 protein.


* * * AND NOW, DANCE PARTY ! * * *


they all try to keep up, while we fuck this shit up! 

SOME RANDOM THINGS .

just have to stay awake one more week...

C O F F E E




process some fluorescent protein photos



writing two papers at the same time so i don't get bored


i missed the king street bike path opening because i was spending time with a friend i haven't seen for many months.  we did yoga and ate ramen.  

crow pose


blackbird

that ramen place: http://www.frolichawaii.com/urbanmixplate/something-new-man-ichi/ 
i still like the flavor of the ramen at that place outside of don quijote on kaheka.  (the restaurant name is in japanese?  i don't know what it's called.)  although, manichi had amazingly tender pork slices and gyoza.  also, the workers were... overly enthusiastic about our presence.  by don quijote they leave you alone. 


i finished reading this book 

╮ (. ❛ ᴗ ❛.) ╭

i feel better about myself because i know there are people crazier than i am in the world! :D 

next on my reading list (gotta start after this semester is over... maybe even next year) is  The Checklist Manifesto: How to Get Things Right

The Checklist Manifesto: How to Get Things Right

what else planned for next year?  my dentist said i need braces. :|
twenty six fucking years old and i need braces.
i feel like he's trying to rip me off but i'm not sure... 


rather spend my $$$ on comic books.
i haven't been to the comic book store in over a month... T___T  so many cliff hangers need to be resolved! 

alas, i spent a large chunk of money on some xmas gifts
got this cool shirt for my dad! 


and some really nice bowls from the UH Manoa Art department glass sale

(sorry for the face i was sending this via imessage to my sister)



several people at work are calling out one hour before their shift starts because of "school".  i want to say to them, SUCK IT UP, PUSSIES, OR QUIT YOUR JOB.  geez, at least have the courtesey of calling out the day before so we can find a replacement!  13 credits and 40 hours of work and i haven't called in sick since 2012.  BITCHES.  WHATEVER.  WE DON'T NEED YOUR HELP.  THE MORNING ROUNDS GET DONE WITH OR WITHOUT YOU.


have a nice day, the end, so long pard'ner, see you next week after i turn in all my homeworks and become a sane person again.

INSTANT TIME MACHINE!

CERCA TROVA:

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for a pulse sed rate seeking a friend for the end of the world self defense family self published separation sequins seven psychopaths shadowplay shaloha sheezus shimmering stars shoes shout out louds show sierra nevada sikdorak silly girl sinclair library skateboard skyfall slipshift small brown bike smart db smoke gets in your eyes snl somnio sorority noise soundcloud speech and debate sphere spyral stamps staph staphylococci star trek steam stellastarr* straddle ups string db suetonius suicide squad suis la lune sunday morning at a funeral sunny day real estate surya namaskar sushi sushi king swim swoon taco tuesday taiyaki tantalus target taste tea taunt tcb technical writing tempura tf2 tguk thanksgiving the angels the audition the bougies the calm blue sea the chase the get up kids the hives the killers the kingsmen the kinks the pipettes the quick the raconteurs the russian the saddest landscape the supremes the things we think we're missing the voices the weight of the world 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