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From 2013.igem.org

Contents 1 Cooperation with Freiburg(Hanna) 2 Lyon (Hanna, Philipp, Tania) 3 Human Practice 4 Financing, Karl Steinbruch Stiftung, Software Project 5 Lab Update 5.1 Delftibactin 5.2 Indigoidine 5.3 Tyrocidine 5.4 Methyl-malonyl CoA 6 NRPS

Cooperation with Freiburg(Hanna)

• arrive on Monday(14.7.) want to show us Gibbson Assembly
• accomodation: Philipp, Julia, Hanna/Ralf
• Would go out for Dinner at Schmidt's, Hanna is going to send an Email
• we should take away all the things in the lab indicating our actual project with Delftibactin

Lyon (Hanna, Philipp, Tania)

• hotel: 7x 3 people in one room
• train to Lyon is booked, need to book the ride back on 15.7. (Monday)

Human Practice

• Konrad offered to take care of the soil samples to send to the UK (?)
  • want to get some info on Streptomycces from them.
• Fanny and bts take care of the discussion round

Financing, Karl Steinbruch Stiftung, Software Project

• Anja: we need three people's CVs that have worked in Bioinfomatics and have more than 120 ECTS
• they want to know about where we spent the money
• max ammount is app. 830 €/month
• dead line is 31.7.

Lab Update

Delftibactin

• Del H
  • problems with 1B bands
    • new amplification worked
    • were digested and purified today
    • are going to be ligated tomorrow

Dominiks suggestion:

• divide ligation after 1 hour into two, one is kept at -20°C the other is let continue over night at 4°C
  • pool both together and electroporate this batch
• gibbson primers arrived today; this will be the new approach, if ligation is unsuccessful
  • parallel gibbson does not make sense, to expensive

---> gel is running right now and will influence decision on further proceeding

• Del Rest
  • 7-8 fragments of 32 are still missing
  • backbone has been amplified
  • Flo designed new primers
    • for DelOP

Indigoidine

• pptase could be amplified, but not indigoidine synthetase
  • brainstorming
    • maybe the gene is not in this sub-species?
• plamsid from UTAH arrived
• want to write to other labs if they can send us the plasmid
• there is a different Streptomyces (Stamm) that has this enzyme but need to finde the correct name
• burocracy discussion.. MTA, Hauspost, Eils signature
• primer strategy with BPSA

Tyrocidine

• Stamm come from Marahije (?)
• Glycerol stocks,colony PCRs
• new strategy has be developed
• ordering of primers by end of the week

Methyl-malonyl CoA

• want genomic integration of substrate synthesis complex for DelF
• got the insert from the USA
• just need to amplify the insert with appropritate primers with some extra DNA
• first tries didn't work
• has several primers for accurate testing/screening
• if integration worked only one resistance gene copy
• gets fragment PLUS other bands of something else...
• did electroporation and purification this week
  • some altered conditions
• some good colonies
• screening

NRPS

• primary goals of the NRPS etc
• NRPS are modulary
• three domains, CA-T-optional
• flexible linkers between domains
• can be put into different orders
• wetlab:
  • goal: library of physical DNA of single modules to be able to put into different orders
  • start with tyrocidine pathway

• software:
• tell what you want
• has info on different pathways and domains
• tells me how to ligate everything together etc
• specifity is dependent on CA domain

• look at what is most probable to work later

--> calculate evolutionary distance according to the taxon ids in the database

• 4 nodes:
  • b.brevis
  • e.coli.
  • p. aeruginosa
  • b.subtilis

• Discussion about Linkers:
  • should they be standardized?
  • how should they look like?
  • domains are highly conserved among species and linkers are NOT conserved at all