Fast solidification of agarose gels

Hi there! Here is a simple but efficient trick to get your agarose gels to solidify much faster.  You have to weight the required amount of agarose as usual, but here it comes the trick. To dissolve this agarose, only pour HALF the amount of the required TAE 1% and save the rest (for example, to prepare a 1% gel, weight o.5 of agarose and instead of pouring the full 50ml, just pour 25ml and save the rest).

Proceed with the protocol boiling the mixture in the microwave until the agarose melts completely. Then, add the TAE you saved (25ml in our example) which will be at room temperature and will cool down your mixture  much faster to reach 60C-70C. Then you don’t need to wait any longer to add the ethidium bromide and to pour the gel in the tray.

You know you’ve worked too long in a lab when

Very funny! and very true! hehehe. I got it from a friend in facebook. There is this facebook group called You know you’ve worked too long in a lab when, Daniel Sutton from London is the admin, but I don’t really know who is/are the author/authors of these hilarious statements. LOL

1. You wonder what absolute alcohol tastes like with orange juice

2. You can tell what cheap and expensive white coats look like

3. You can’t watch CSI without cursing at least one scientific inaccuracy

4. You use acronyms for everything and never stop to elaborate

5. Liquid nitrogen is only about a 1/3 as dangerous as you thought

6. You always seem to use the microscope after the person with the impossible close together eyes

7. Accident reports are a badge of honor

8. You’ve wondered why you can’t drink distilled water in the lab – It should be clean?

9. You give the lab equipment motivational pep talks “Work for me today or i’ll reprogram you with a fire axe” is my favorite

10. You’ve worked out that a trained chimp could probably do 90% of your job

11. When a non-scientist asks you what you do for a living you roll your eyes and talk science at them until they’ve loss the will to live (mainly for fun)

12. You have to check the web to find out what the weather is outside

13. You realize that almost anything can be classed as background reading

14. People wearing shorts under a lab coat disturb you slightly as they look as though they might be naked underneath

15. Although all cooking is a glorified chemistry experiment you just still can’t seem to get it right

16. Safety equipment is optional unless it makes you look cool

17. Warning labels invoke curiosity rather than caution

18. The Christmas nightout reveals scientists can’t dance, although a formula for the movement of hands and feet combined with beats per min is found scrawled on a napkin by a waiter the next day

19. You know which part of the lab you can chill out undisturbed on friday afternoon

20. You decide the courses and conference you want to go on by the quality of the food served

21. You are strangely proud of the collection of junk you’ve stolen from vendors at trade shows

22. You’ve used dry ice to cool beer down

23. No matter what the timings in the experiment protocol there is always time for lunch in the middle

24. As has been pointed out to me on several occasions – You can no longer spell normal words but have no trouble with spelling things like immunohistochemistry or deoxyribonucleic acid.(Cheers Lesley)

25. Burning eyes, nose and throat indicate that you haven’t actually turned on the fumehood/downdraft bench

26. Your slightly too fond of the smell of (pick one or many) Xylene/Agar/Ethanol/Undergraduates/Alcoholic handwash

27. You’ve left the lab wearing a piece of PPE (personal protective equipment) because you forgot you had it on

28. You bitch about not being able to pipette by mouth any more (Not me but i’ve worked with people who do!)

Signs from others in the group (04-07-07)

29. Security come round at 2 am wondering why the lights are still on only to find you with your arms up to your elbows in a glovebox – Cheers James

30. you have made some kind of puppet out of a nitrile glove and kept it as a pet (I know this isn’t just me!) (Putting dry ice in makes for a rapidly expanding if short lived pet – DS) – Cheers Rachel

31. When at a Fall Out Boy gig you wonder why everyone is going round with Faecal Occult Blood (FOB) written on their head!!!! -Just for you Sarah

32. You have an irresistible urge to rip your shirt off superman stylie cos it has press stud fasteners just like your lab coat…..Most often occurring as you walk through a door just like exiting the lab…. (The worlds of strippers and lab workers collide, not pretty- DS) Thanks for that Carrie

33. You still get amusement out of “freezing” things in liquid nitrogen! – Not just you Tracey

A Few more from me (12-07-07)

34. Blinking real fast has saved your eyesight on more than one occasion.

35. You’ve removed your gloves to find a small hole which has left you with either – wrinkly old person hands, a brightly coloured finger (histologists especially) or a burning sensation and dermatitis and some point.

36. You’ve bent down to pick something up off the floor only to scatter the contents of your top pocket under the largest machine in the lab – Common problem i believe

More From you guys (19-07-07)

37.When you rejoice when grabbing a handfull of eppendorfs/bijous/anything and it turns outs to be the exact number you needed (always strangly good andy)

38.You can`t wait for lab clean-up coz you get to do random pointless “experiments” to figure out whats in all the dodgy unlabeled bottels (Sniff test is a bit of a gamble Nadia)

39. You hate having to change your lab coat to a new one because ‘it just won’t fit right’ and because the wrist bits are way too tight (They never get my ‘cut’ just right either Tom)

40. You know you have worked in a lab too long when you actually threaten your cells whilst waving a bottle of virkon (All been there Becky)

41.Your nose invariably itches when you’re doing mucky stuff with your hands so you develop the habit of scratching it on your upper arm. Unfortunately you sometimes carry this habit over to real life, where it looks like you’re sniffing your armpits (Trying to find a clean bit of lab coat can be fun as well, cheers Kate)

42. When as the senior of morphology you threaten each new registrar on their first day that oil and x10 dry objectives do not mix and will result in violence (Cheers Nichola)

43. when you say goodnight to your microscope on a friday night and tearfully hug it goodbye as you won’t see it all weekend (Cheers again Nichola)

44. When you start making patterns in your pipette tip box as you take the tips out. I made a beautiful spiral today (Could have been an art student Vicky)

45. When you wonder how much it will hurt if I pour just a smidge of this phenol:chloroform/trichloroacetic acid/any random chemical on myself (Best try it out on some one else first Mike)

46. You’ve seen how far away you can hit a target with a squirty water bottle or seeing how far away from the bin i can fire pipette tips. (Pinging gloves is also fun Ed)

47. The fire alarm ceases to bug you. You only evacuate when you see the fire. (Hand on the floor to check for heat is a good indicator)

48. You know when you’ve been in a lab too long when you make 6 litres of medium, but wonder why no one makes “high” or “low”.(Cheers Tom)

49. When you organise your kitchen cupboard contents the way you would your chemicals..all labeled in alphabetical order (Cheers Anggia)

50. When you’ve got that callus on the side of your thumb from opening PCR tubes (Cheers Chani)

A few more sent to me by a colleague, dam that’s another one – you call your friends colleagues “my colleague is just getting the beers in”

51. You open the toothpaste with one hand.

52.You wash your hands before and after using to the washroom.

53.When you hear tween, you think of the surfactant not the age group.

54.For you, media is something which increases your culture.

55.You can identify organs on road kills.

56.You have a callus on your thumb.

57.You use the word “aliquot” in regular sentences.

58.Sometimes you momentarily vanish from social activities because of a time point.

59.You’ve never worn a clean lab coat.

60.You don’t fear rodents, rodents fear you.

61.You say “orders of magnitude” in regular sentences.

62.You flinch when you hear the word “significant”.

63.Showing up at 10AM and having a coffee is a productive day.

64.You can’t stand god-like physicians, while secretly wishing you had their job.

65.You’re very good at diluting things.

66.You’re also very good at transferring small amounts of liquid between containers.

67.You are fed up of people saying alcohol, when they mean ethanol.

68.You say “conjugation” instead of “sex”, and “pili” sounds dirty.

69.SOB is not an insult; it’s what you grow your bugs in.

70.You say “mills” and “megs”.

71.No-one in your family has any idea what you do.

72.You can make a short film in power point.

73.You consider a green laser pointer to be science bling.

74.When your fruits go bad and you get fruit flies, you can’t help but check their eye colour

75.You own invitrogen t-shirts and actually wear them.

76.You refer to your children as the F1.

77.You’ve suffered carpal tunnel from the pipetman.

78.You’ve used Kimwipes as Kleenex.

79.A timer clipped to the hip is not only practical, but dead sexy.

80.You’ve played Battleship using tip boxes.

81.The front page of Science is your light reading.

82.You think the following is a quality insult: “I’ve seen cells more competent than you!”.

83.The scent of latex reminds you of work, not play.

84.You’ve used, “I’d like to get into your genes” as a pickup line.

Saving time and money doing maxi/midi preps

Here is a trick. It is very simple, only a bit messy. But it will save you one centrifugation step and 30 minutes.

You have to pellet your overnight culture as usual, ressuspend it, lysate it and stop the lysis. So far, as recommended in the protocol. The next step is normally a centrifugation to clear the lysate. HERE is where the trick comes.

You don’t need to centrifugate. You can use a clean coffee filter and the lysate will came out as clean as after centrifugation. If you don’t belived, try it. It works. Add the lysate to the coffee filter and collect it in a clean 50ml falcon tube. Fast, right?

At that point, you can add the filtrate straight to the equilibrated column and continue with the washings as written in the protocol.

The lucky ones that buy the filter cartridges provided by the companies probably won’t ever use this trick. But who knows, maybe you run’ll out of them on a Sunday afternoon,… and then is always good to have a plan B.

George W. Bush contribution to science

Ex-president George W. Bush inesperately contributed to science when he was able to duck flying shoes thrown at him by an Iraqi reporter in December in Bagdad as reveals the study published this month in the magazine Current Biology by two neurologists of Washington University.

Bush reflexes and the impassivity that al-Maliki, the Iraqi primer Minister, who was next to the US president, showed, support the theory of the two scientists that claims that two independent pathways in the human visual system exist.
One system guides the actions and the other the perception. The interesting part is that the first one allows the brain to “see” thing that the eyes don’t perceive, says Jeffrey Lin, first author of the study.

“When we throw two balls at you with very similar trajectories, they may look the same to your perceptual system, but your brain can automatically calculate which one is more threatening and trigger a dodging motion before you’ve even realized what has happened,” said Lin.

That explains why in the video al-Maliki doesn’t flinch “his brain has perceived that the shoe is not a threat for him. But Bush’s brain has categorized the shoe as threatening and triggers an evasive dodge, all within a fraction of a second”.

The scientists performed several experiments with students to demonstrate their theory, and the incident in Bagdad was an inesperate and welcomed example in real life.

Read the extended news at scientificblogging 2.0

Most commonly used lysis buffers

RIPA

150 mM NaCl
1 mM PMSF
1 mM EDTA
5 µg/ml Aprotinin
5 µg/ml Leupeptin
1% Triton x-100
1% Sodium deoxycholate
0.1% SDS
50 mM Tris, pH 7.4


NP-40

NaCl 150 mM
NP-40 1%
Tris, pH 8.0 50 mM


HEPES

Hepes pH 7.4 50mM
NaCl 150mM
Glycerol 10%
Triton X-100 1%
KCl 5mM
EDTA 1mM
Protease inhibitors (leupeptin 10ug/ul, aprotinin 10ug/ul, PMSF 1mM)
Phosphatase inhibitors (NaF 50mM, Na3VO4 0.5mM, Na4P2O7 5mM)

Odyssey infrared imaging system

Direct infrared fluorescence detection on the Odyssey Infrared Imaging System provides the established standard for Western blot analysis that can’t be equaled with chemiluminescence and visible fluorescence. Infrared detection gives you the quantitative analysis and wide linear dynamic range that chemiluminescence cannot. Strong and weak bands on the same blot are accurately detected without the uncertainty and inconvenience of multiple exposures, and without spending time in the darkroom. The Odyssey System gives you clear, sharp, reproducible bands without fuzziness or “blowout”. Bands hidden by overexposure with chemiluminescence are clear in Odyssey images.

Web site: http://www.licor.com/bio/odyssey/index.jsp

Spectrophotometric quantification of DNA and RNA

Because DNA and RNA absorb ultraviolet light, with a absorption peak at 260nm wavelength, spectrophotometers are commonly used to determine the concentration of DNA in a solution. Inside a spectrophotometer, a sample is exposed to ultraviolet light at 260 nm, and a photo-detector measures the light that passes through the sample. The more light absorbed by the sample, the higher the nucleic acid concentration in the sample.

Using the Beer-Lambert law it is possible to relate the amount of light absorbed to the concentration of the absorbing molecule. At a wavelength of 260 nm, the extinction coefficient for double-stranded DNA is 50 (μg/ml)-1 cm-1; for single-stranded DNA and RNA it is 38 (μg/ml)-1 cm-1. Thus, an optical density (or “OD”) of 1 corresponds to 50 µg/ml for double-stranded DNA, 38 µg/ml for single-stranded DNA and RNA. This method of calculation is valid for up to an OD of at least 2

Concentration of double stranded DNA = absorvance at 260nm * 50 * dilution (if you diluted 1 in 100 to quantify, the dilution value in this case will be 100)

Concentration of single-stranded DNA and RNA = absorvance at 260nm * 38 * dilution (if you diluted 1 in 50 to quantify, the dilution value in this case will be 50)

Protocol for Poly-D-Lysine Preparation and Plate Coating

1. Prepare Poly-D-Lysine
a. Add 100 ml of sterile tissue culture grade H2O to 5 mg poly-D-lysine (Sigma P6407). Final concentration of poly-D-lysine is 50 ug/ml.
b. Mix by pipetting several times.
c. Store at 2-8 ºC or –20 ºC.

2. Prepare Coated Plates
a. Add enough poly-D-lysine solution to cover culture plate surface.
b. Incubate at room temperature for 1-24 hours.
c. Aspirate the solution.
d. Rinse the plate surface thoroughly with sterile tissue culture grade H2O.
e. Dry the plate.
f. Store the plate at 4-8 ºC or room temperature.

protocol extracted from Gene Therapy Systems, Inc.

10X TBS (Tris Buffered Saline)

To prepare 1 liter of 10X TBS:

30 gr  Tris
88 gr NaCl
2 gr KCl

mix everything in dwater until it’s disolved. Adjust pH at 7.5

Paraformaldehide 4%

To prepare 100ml of PFA 4% stock solution:

- Add 86ml ddwater in an enlermeyer
- Boil water in the microwave
- Weigh 4g of PFA

From now on in the fume hood
- Transfer PFA in the erlenmeyer
- Add a clean stirring bar
- Add a drop of NaOH 10M
- Stir it and keep it hot (heat at #1-2) till it goes clear (DO NOT LET PFA BOIL)
- Place on ice to cool it down
- Adjust volume to 90ml
- Add 10ml PBS 10X
- Adjust pH to 7.4

Store at RT or 4C