August 2016 | Science

[DIYbio] Re: extracting DNA from bacteria

6:41 PM |

huh, so it actually works. That's definitely cheaper than a kit. Thanks for sharing!

On Wednesday, August 31, 2016 at 4:46:52 PM UTC-4, Mega [Andreas Stuermer] wrote:

http://openwetware.org/wiki/User:Andrew_Perry/Protocols/DNA_gel_extraction

I once gave a protocol like this a shot. It seemed to have worked. The DNA gets diluted but it should be possible to work with.

Attached are photographs, from when I tried it many many years ago.

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RE: [DIYbio] Re: Perth (Australia) Group

3:19 PM |

G'day all,

Fortuitously, I'm moving back to Perth today and might tag along on Saturday if that's alright?

Cheers,

Mike

Michael Strack
PhD candidate, Macquarie University
Department of Biological Sciences
Ecotoxicology and Freshwater Ecology lab
+61 434 920 666
http://www.twitter.com/mixtrak

From: Nathan Thompson
Sent: Wednesday, 31 August 2016 12:49
To: diybio@googlegroups.com
Subject: RE: [DIYbio] Re: Perth (Australia) Group

Hi Benedict, Philip, Leon and all other Perth based Outsider Biologists.

This Saturday the 3rd, 2pm at the Artifactory looks interesting.
Oron Catts, Christine, Ito and myself from SymbioticA UWA will be attending to check things out.

Looking forward to it.
Regards
Nathan

Date: Sun, 28 Aug 2016 21:50:13 -0700
From: bjnoel@gmail.com
To: diybio@googlegroups.com
Subject: [DIYbio] Re: Perth (Australia) Group

Hey all,

Just a reminder that we've got a meeting at Artifactory this Saturday (3rd) at 2pm - all are welcome. Leon will be walking us through building a magnetic stirrer (I'll bring some liquid cultures to be stirred). I was hoping we'd get to use the laser cutter to create some equipment but I need to get some training and the acrylic we need is on order - we'll do it next time instead.

If anyone else has something they'd like to share, please bring it along.

Cheers,

Benedict

On Friday, 12 August 2016 18:07:06 UTC+8, Philip Wijesinghe wrote:

Hi All,

Andrew - thanks for the kind words. Student engagement and the whole atmosphere seems to be more energetic at UWA than when I was an undegrad : I have high hopes for the new bioengineering and data science degrees, and all that will come with it. Thanks for taking the time to mentor and nurture us, really appreciated.

Steve - are you very active at the Arifactory? I'm asking because I'm really interested in checking out the space.

With biohack academy: there was the first organiser's conference call last Tuesday.
It will run from the 31st Jan 2017 for 10 weeks. Each week we would be expected to put in no less than 25 hours (2hrs lectures+2hrs discussion \ 5 hrs organisation \ 16 hours in a lab/makerspace) - this is the first hurdle.
The second hurdle is the costs. For recommended ~10 participants per 2 organisers, the costs are roughly $1k per head. $10k total. Not many people, especially students, have that kind of money, especially if they are giving up so much time.
I'm applying for funding from the UWA to cover most of the costs. I've started outlining a detailed budget estimate - I'll share it once it is done, and we'll see if there is space to trim.
Third is space - we need 3 days\week access to a somewhat clean lab with basic sink/fridge/labcoats etc. This is the least of the problems as there are a few places where we may be able to do it.

Cheers
Philip

On Thursday, 11 August 2016 15:22:11 UTC+8, BigSteve wrote:
Hi Philip,

Thanks for reaching out! I've involved with the Artifactory group
The Biohack academy sounds amazing! What would it take to make it happen?

It sounds like there will be another Artifactory meet-up in September, it would be great to collaborate!

-Steve

On Tuesday, 26 July 2016 11:08:14 UTC+8, Philip Wijesinghe wrote:
[cross-post from facebook]

Hi All. Anyone interested in a meetup in the near future? There seem to be 3-4 different diyBio/maker related groups and projects in Perth, running somewhat independently (and on different social media and team comms apps). It would be good to join in collective efforts.

Also, would anyone be keen on participating in the BiohackAcademy 2017 (~30Jan17 for 2.5 month), if I start looking at feasibility and sorting logistics to host it in Perth?
https://waag.org/en/project/biohack-academy-biofactory

Also, anyone keen on doing the upcoming GovHack? https://www.govhack.org/

Cheers,
Philip

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[DIYbio] Re: extracting DNA from bacteria

1:46 PM |

http://openwetware.org/wiki/User:Andrew_Perry/Protocols/DNA_gel_extraction

I once gave a protocol like this a shot. It seemed to have worked. The DNA gets diluted but it should be possible to work with.

Attached are photographs, from when I tried it many many years ago.

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Need IBM Tivoli ; @ Morristown, NJ ; Long term contract

9:32 AM |

Hi,

Title : IBM Tivoli

Location : Morristown, NJ

Duration : Long term contract

Skill :

· Oracle IDM or Tivoli IDM with strong development knowledge in Core Java

· Experience with Identity & Access management tools is a plus, such as IBM Security Access Manager for Web and Mobile, Tivoli Directory Server, Tivoli Federated Identity Manager, Tivoli Identity Manager and DataPower IND2016

Thanks and Best Regards,

Praveen Kumar

SRI Tech Solutions Inc.

praveen@sritechsolutions.com

(T) 1-813-423-6500 X Ext: 149 (M) 813-440-2778 (F) 813.423.6520

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[DIYbio] Re: extracting DNA from bacteria

5:07 AM |

thanks guys! A few people also sent me private messages recommending the colony PCR route as well, so colony PCR I shall try.

@Cathal

Thanks! I have developed a odd relationship with these bacteria. They are just so cool. The Odin tutorial on e. coli transformed with pVIB said that they like cool and dark, so I left one of my cultures at room temp at my lab desk and another in the fridge wrapped in aluminum foil, and the later was brighter. I didn't know that they "officially" don't like light and it's definitely something interesting to experiment with. In terms of the squishy gel extraction method, where does the dna end up? The Odin kit came with the green dye

@koeng

avoiding the use of gels would help me keep the cost down haha but how do I go around confirming and cleaning out the DNA? Do the PCR clean up kit and hope there's something in there? It would be a bummer to send the result of a failed PCR reaction in for sequencing :(

@Patrick

Doh! a direct link from the page I was at... thanks for pointing it out. I think I'll take the opportunity to learn and practice lab skillz.

On Tuesday, August 30, 2016 at 12:26:53 AM UTC-4, Towa wrote:

hey guys! I was able to culture some bioluminescent bacteria from squid and I want to know what it is. My guess is that its V. Phosphoreum but to confirm, I want to extract it's DNA and try the 16S rRNA identification with primers from Odin (http://www.the-odin.com/bacterial-16s-rdna-primers-for-bacterial-identification/). Should I be able to get PCR-able DNA out with dish detergent and ethanol/isopropanol? Any good protocols that are DIY friendly?

on a side note, does anyone have either a) good DIY gel purification method or b) someone that will sell kits to ronin-hackers (do not belong to any institution)?

as always, thanks!

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RE: [DIYbio] Re: Perth (Australia) Group

7:49 PM |

Hi Benedict, Philip, Leon and all other Perth based Outsider Biologists.

This Saturday the 3rd, 2pm at the Artifactory looks interesting.
Oron Catts, Christine, Ito and myself from SymbioticA UWA will be attending to check things out.

Looking forward to it.
Regards
Nathan

Date: Sun, 28 Aug 2016 21:50:13 -0700
From: bjnoel@gmail.com
To: diybio@googlegroups.com
Subject: [DIYbio] Re: Perth (Australia) Group

Hey all,

Just a reminder that we've got a meeting at Artifactory this Saturday (3rd) at 2pm - all are welcome. Leon will be walking us through building a magnetic stirrer (I'll bring some liquid cultures to be stirred). I was hoping we'd get to use the laser cutter to create some equipment but I need to get some training and the acrylic we need is on order - we'll do it next time instead.

If anyone else has something they'd like to share, please bring it along.

Cheers,

Benedict

On Friday, 12 August 2016 18:07:06 UTC+8, Philip Wijesinghe wrote:

Hi All,

Andrew - thanks for the kind words. Student engagement and the whole atmosphere seems to be more energetic at UWA than when I was an undegrad : I have high hopes for the new bioengineering and data science degrees, and all that will come with it. Thanks for taking the time to mentor and nurture us, really appreciated.

Steve - are you very active at the Arifactory? I'm asking because I'm really interested in checking out the space.

With biohack academy: there was the first organiser's conference call last Tuesday.
It will run from the 31st Jan 2017 for 10 weeks. Each week we would be expected to put in no less than 25 hours (2hrs lectures+2hrs discussion \ 5 hrs organisation \ 16 hours in a lab/makerspace) - this is the first hurdle.
The second hurdle is the costs. For recommended ~10 participants per 2 organisers, the costs are roughly $1k per head. $10k total. Not many people, especially students, have that kind of money, especially if they are giving up so much time.
I'm applying for funding from the UWA to cover most of the costs. I've started outlining a detailed budget estimate - I'll share it once it is done, and we'll see if there is space to trim.
Third is space - we need 3 days\week access to a somewhat clean lab with basic sink/fridge/labcoats etc. This is the least of the problems as there are a few places where we may be able to do it.

Cheers
Philip

On Thursday, 11 August 2016 15:22:11 UTC+8, BigSteve wrote:
Hi Philip,

Thanks for reaching out! I've involved with the Artifactory group
The Biohack academy sounds amazing! What would it take to make it happen?

It sounds like there will be another Artifactory meet-up in September, it would be great to collaborate!

-Steve

On Tuesday, 26 July 2016 11:08:14 UTC+8, Philip Wijesinghe wrote:
[cross-post from facebook]

Hi All. Anyone interested in a meetup in the near future? There seem to be 3-4 different diyBio/maker related groups and projects in Perth, running somewhat independently (and on different social media and team comms apps). It would be good to join in collective efforts.

Also, would anyone be keen on participating in the BiohackAcademy 2017 (~30Jan17 for 2.5 month), if I start looking at feasibility and sorting logistics to host it in Perth?
https://waag.org/en/project/biohack-academy-biofactory

Also, anyone keen on doing the upcoming GovHack? https://www.govhack.org/

Cheers,
Philip

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[DIYbio] Re: extracting DNA from bacteria

2:00 PM |

The Odin actually links to a very simple protocol for 16s rDNA PCR, from the page that you referenced:

16s rDNA Amplfiication and Sequencing

They're actually recommending to scrape a single colony straight into the PCR reaction. No separate DNA extraction step whatsoever - how simple can you get?

If you're more interested in getting the answer asap than in teaching yourself how to do 16S PCR, many sequencing services will also do 16S PCR and sequencing straight from a colony or liquid culture.

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Re: [DIYbio] extracting DNA from bacteria

11:34 AM |

I'd recommend to just avoid gels altogether. Restriction enzymes are great, but gel purification just kills that method for me. I always gibson/goldengate, with one of the main reasons I hate cutting gels. i don't know if it is just the kit I use, but I always end up with highly salted un-pure DNA. If anyone knows of a good kit to purify DNA from gels, I'd love to know what it is.

@Cathal I am actually going to try the method with filter paper. I will report back in a while if it works or not.

-Koeng

On Tuesday, August 30, 2016 at 8:03:30 AM UTC, cathal...@cathalgarvey.me wrote:

Congratulations on your new glowing minions! :)

Watch out for a small gotcha with V.phosphoreum; at least with the strain that I isolated years back, when grown under light they do not glow. They need to be insulated from the light to produce the luciferin. Whether this was photobleaching or transcriptional regulation, I do not know; would make an interesting (and perhaps publishable) project, though. Even finding out which frequencies it responds to would be fun.

For DNA extraction, you're dealing with a gram-negative without much peptidoglycan, so there are plenty of handy options. Classic alkaline lysis will work, as would ultra-dirty "boiling lysis" methods, and in either case you should get plenty of 16S to work with IMO. I won't comment on the procedure for 16S identification itself because I've never done it. Just make sure you clean up any detergent and salt from your prep method; dishwashing detergent for example may have tons of unidentified enzyme inhibitors, and good luck getting the company to tell you what's in it! Ethanol is apparently better than isopropanol for de-salting DNA after extraction, I'm not sure how it compares on detergents.

I'm *told* that the best gel extraction method is to slice your DNA fragment out as small as you can make it, then put it in a PCR tube with a hole cut in the end and some filter paper covering the hole..then spin it inside a larger tube in a centrifuge, so the gel fragment gets squished flat and the liquid escapes into the larger tube. I have also never done this, however. :)

The best advice I can give for gel extraction is..never, ever use Ultraviolet. Not even once. Go blue or go home. :) So, SYBR dyes or similar would be fine, but an Ultraviolet dye will lead to shredded DNA and poor PCR outcomes.

August 30, 2016 5:27 AM, "Towa" <tow...@gmail.com> wrote:

hey guys! I was able to culture some bioluminescent bacteria from squid and I want to know what it is. My guess is that its V. Phosphoreum but to confirm, I want to extract it's DNA and try the 16S rRNA identification with primers from Odin (http://www.the-odin.com/bacterial-16s-rdna-primers-for-bacterial-identification). Should I be able to get PCR-able DNA out with dish detergent and ethanol/isopropanol? Any good protocols that are DIY friendly?

on a side note, does anyone have either a) good DIY gel purification method or b) someone that will sell kits to ronin-hackers (do not belong to any institution)?

as always, thanks!

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Need IBM Tivoli ; @ Philly , NJ ; Long term contract

10:34 AM |

Hi,

Title : IBM Tivoli

Location : Philly , NJ

Duration : Long term contract

Skill :

· Oracle IDM or Tivoli IDM with strong development knowledge in Core Java

· Experience with Identity & Access management tools is a plus, such as IBM Security Access Manager for Web and Mobile, Tivoli Directory Server, Tivoli Federated Identity Manager, Tivoli Identity Manager and DataPower IND2016

Thanks and Best Regards,

Praveen Kumar

SRI Tech Solutions Inc.

praveen@sritechsolutions.com

(T) 1-813-423-6500 X Ext: 149 (M) 813-440-2778 (F) 813.423.6520

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Position: LTE Test Engineer , TX or WA

10:33 AM |

Hi,

Please find the below position and respond back to me at praveen@sritechsolutions.com

Position: LTE Test Engineer

Location: Austin, TX or Redmond, WA

Duration: 6-12 months + Extendable

Description:

· 3-5+ years of LTE testing experience.

· Experience with OpenStack

· Experience with Virtualization.

· Excellent communication skills.

Thanks and Best Regards,

Praveen Kumar

SRI Tech Solutions Inc.

praveen@sritechsolutions.com

(T) 1-813-423-6500 X Ext: 149 (M) 813-440-2778 (F) 813.423.6520

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2 hot Requirements || Salesforce Developer & Salesforce Apex developer || USC / GC / EAD-GC Only || Phone hire

8:52 AM |

Hope you are doing great!!

Please add me on yahoo at arpittechwire and do share me your hotlist also at Arpit@tresourceinc.com.

Please go through the requirement and let me know if you have any consultant for the same position.

Note: Looking for USC or GC or GC-EAD ………

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Job Description :

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Job Description :

Someone with several years' experience doing Salesforce and Apex development, configuration, and customization work. Specific experience

with nCino Bank Operating System would be a huge bonus. Experience

working in the credit and loan processing financial sector would be "nice to have", but not a replacement for the technical skills needed.

Thanks & Regards,

Arpit Arora
Technology Resource Group Inc.
1700 Park St, Unit 212, and Naperville IL, 60563
Direct: 909-859-1312 | Office:408-709-1760 Ext : 9961
Fax: 408-884-2409
Arpit@tresourceinc.com | www.tresourceinc.com

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Re: [DIYbio] extracting DNA from bacteria

1:03 AM |

Congratulations on your new glowing minions! :)

Watch out for a small gotcha with V.phosphoreum; at least with the strain that I isolated years back, when grown under light they do not glow. They need to be insulated from the light to produce the luciferin. Whether this was photobleaching or transcriptional regulation, I do not know; would make an interesting (and perhaps publishable) project, though. Even finding out which frequencies it responds to would be fun.

For DNA extraction, you're dealing with a gram-negative without much peptidoglycan, so there are plenty of handy options. Classic alkaline lysis will work, as would ultra-dirty "boiling lysis" methods, and in either case you should get plenty of 16S to work with IMO. I won't comment on the procedure for 16S identification itself because I've never done it. Just make sure you clean up any detergent and salt from your prep method; dishwashing detergent for example may have tons of unidentified enzyme inhibitors, and good luck getting the company to tell you what's in it! Ethanol is apparently better than isopropanol for de-salting DNA after extraction, I'm not sure how it compares on detergents.

I'm *told* that the best gel extraction method is to slice your DNA fragment out as small as you can make it, then put it in a PCR tube with a hole cut in the end and some filter paper covering the hole..then spin it inside a larger tube in a centrifuge, so the gel fragment gets squished flat and the liquid escapes into the larger tube. I have also never done this, however. :)

The best advice I can give for gel extraction is..never, ever use Ultraviolet. Not even once. Go blue or go home. :) So, SYBR dyes or similar would be fine, but an Ultraviolet dye will lead to shredded DNA and poor PCR outcomes.

August 30, 2016 5:27 AM, "Towa" <towa.ee@gmail.com> wrote:

hey guys! I was able to culture some bioluminescent bacteria from squid and I want to know what it is. My guess is that its V. Phosphoreum but to confirm, I want to extract it's DNA and try the 16S rRNA identification with primers from Odin (http://www.the-odin.com/bacterial-16s-rdna-primers-for-bacterial-identification). Should I be able to get PCR-able DNA out with dish detergent and ethanol/isopropanol? Any good protocols that are DIY friendly?

on a side note, does anyone have either a) good DIY gel purification method or b) someone that will sell kits to ronin-hackers (do not belong to any institution)?

as always, thanks!

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[DIYbio] extracting DNA from bacteria

9:26 PM |

hey guys! I was able to culture some bioluminescent bacteria from squid and I want to know what it is. My guess is that its V. Phosphoreum but to confirm, I want to extract it's DNA and try the 16S rRNA identification with primers from Odin (http://www.the-odin.com/bacterial-16s-rdna-primers-for-bacterial-identification/). Should I be able to get PCR-able DNA out with dish detergent and ethanol/isopropanol? Any good protocols that are DIY friendly?

on a side note, does anyone have either a) good DIY gel purification method or b) someone that will sell kits to ronin-hackers (do not belong to any institution)?

as always, thanks!

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Position: LTE Test Engineer , WA

7:53 AM |

Hi,

Please find the below position and respond back to me at praveen@sritechsolutions.com

Position: LTE Test Engineer

Location: Austin, TX or Redmond, WA

Duration: 6-12 months + Extendable

Description:

· 3-5+ years of LTE testing experience.

· Experience with OpenStack

· Experience with Virtualization.

· Excellent communication skills.

Thanks and Best Regards,

Praveen Kumar

SRI Tech Solutions Inc.

praveen@sritechsolutions.com

(T) 1-813-423-6500 X Ext: 149 (M) 813-440-2778 (F) 813.423.6520

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Position: Hadoop Architect , WA

7:52 AM |

Hi,

Please find the below position and respond back to me at praveen@sritechsolutions.com

Position: Hadoop Architect

Location: Seattle, WA

Job Type: Contract

Required:

• Design and Solution Architect experience in Hadoop platforms.

• Strong experience in Hadoop Core components – HDFS, Pig , Hive on HDP platform.

• Hands-on experience on JAVA, SQL, Hbase/ Streaming. Would be required to do POC on various aspects of Hadoop work apart from creating strategic vision.

• Working experience in different analytics tools on Hadoop

• Work under client team guidance on evolving big data architecture.

Thanks and Best Regards,

Praveen Kumar

SRI Tech Solutions Inc.

praveen@sritechsolutions.com

(T) 1-813-423-6500 X Ext: 149 (M) 813-440-2778 (F) 813.423.6520

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[DIYbio] Magnetic stirrer ...

9:54 PM |

http://www.instructables.com/id/Magnetic-Stirrer-2/

On Aug 28, 2016 9:50 PM, "Benedict" <bjnoel@gmail.com> wrote:

>

> Hey all,
>
> Just a reminder that we've got a meeting at Artifactory this Saturday (3rd) at 2pm - all are welcome. Leon will be walking us through building a magnetic stirrer (I'll bring some liquid cultures to be stirred). I was hoping we'd get to use the laser cutter to create some equipment but I need to get some training and the acrylic we need is on order - we'll do it next time instead.
>
> If anyone else has something they'd like to share, please bring it along.
>
> Cheers,
> Benedict
>

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[DIYbio] Re: Perth (Australia) Group

9:50 PM |

Hey all,

Just a reminder that we've got a meeting at Artifactory this Saturday (3rd) at 2pm - all are welcome. Leon will be walking us through building a magnetic stirrer (I'll bring some liquid cultures to be stirred). I was hoping we'd get to use the laser cutter to create some equipment but I need to get some training and the acrylic we need is on order - we'll do it next time instead.

If anyone else has something they'd like to share, please bring it along.

Cheers,

Benedict

On Friday, 12 August 2016 18:07:06 UTC+8, Philip Wijesinghe wrote:

Hi All,

Andrew - thanks for the kind words. Student engagement and the whole atmosphere seems to be more energetic at UWA than when I was an undegrad : I have high hopes for the new bioengineering and data science degrees, and all that will come with it. Thanks for taking the time to mentor and nurture us, really appreciated.

Steve - are you very active at the Arifactory? I'm asking because I'm really interested in checking out the space.

With biohack academy: there was the first organiser's conference call last Tuesday.
It will run from the 31st Jan 2017 for 10 weeks. Each week we would be expected to put in no less than 25 hours (2hrs lectures+2hrs discussion \ 5 hrs organisation \ 16 hours in a lab/makerspace) - this is the first hurdle.
The second hurdle is the costs. For recommended ~10 participants per 2 organisers, the costs are roughly $1k per head. $10k total. Not many people, especially students, have that kind of money, especially if they are giving up so much time.
I'm applying for funding from the UWA to cover most of the costs. I've started outlining a detailed budget estimate - I'll share it once it is done, and we'll see if there is space to trim.
Third is space - we need 3 days\week access to a somewhat clean lab with basic sink/fridge/labcoats etc. This is the least of the problems as there are a few places where we may be able to do it.

Cheers
Philip

On Thursday, 11 August 2016 15:22:11 UTC+8, BigSteve wrote:
Hi Philip,

Thanks for reaching out! I've involved with the Artifactory group
The Biohack academy sounds amazing! What would it take to make it happen?

It sounds like there will be another Artifactory meet-up in September, it would be great to collaborate!

-Steve

On Tuesday, 26 July 2016 11:08:14 UTC+8, Philip Wijesinghe wrote:
[cross-post from facebook]

Hi All. Anyone interested in a meetup in the near future? There seem to be 3-4 different diyBio/maker related groups and projects in Perth, running somewhat independently (and on different social media and team comms apps). It would be good to join in collective efforts.

Also, would anyone be keen on participating in the BiohackAcademy 2017 (~30Jan17 for 2.5 month), if I start looking at feasibility and sorting logistics to host it in Perth?
https://waag.org/en/project/biohack-academy-biofactory

Also, anyone keen on doing the upcoming GovHack? https://www.govhack.org/

Cheers,
Philip

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