Notebook

On this page, you can see a summary of what we did each day/week over the course of the year to make our project happen. As it is sometimes hard to keep track of everything, this page is meant as a high-level overview.

January

1st	2nd	3rd	4th	5th	6th	7th

8th	9th	10th	11th	12th	13th	14th

15th	16th	17th	18th	19th	20th	21st
					Kick-off meeting
22nd	23rd	24th	25th	26th	27th	28th
					First regular team meeting (weekly)
29th	30th	31st

February

1st	2nd	3rd	4th	5th	6th	7th
	Wiki & Design Meeting: Planning of Design Theme, Portfolio, and Website		Finance Introduction Meeting: Mentors from last year introduced us into our responsibilities and gave us useful advice.
8th	9th	10th	11th	12th	13th	14th
		Wetlab: first inventory of lab consumables from former iGEM teams	Wiki & Design, Drylab Meeting with Elias from last year’s team: Tips for building the Website and Wiki, Tips for Work in the Drylab
15th	16th	17th	18th	19th	20th	21st
			Finance: communication with old team members to organize the team bank accounts	Wiki & Design Meeting with Lukas from previous iGEM Tuebingen teams: Tips on mentor search, Website building, and Wiki building
22nd	23rd	24th	25th	26th	27th	28th
					Project finding day	We got in contact with our university bank account managers

March

1st	2nd	3rd	4th	5th	6th	7th

8th	9th	10th	11th	12th	13th	14th
Wiki & Design Meeting: Creating the Portfolio using our sponsored Affinity Suite and further discussions about the Portfolio; Website Hosting discussion
15th	16th	17th	18th	19th	20th	21st
	Wiki & Design Meeting: Working on the Portfolio and Design Theme	Meeting with Prof. Schäffer		Meeting with Prof. Forchhammer		Social media meeting: kick-off & brainstorming about plans for our different social media channels
22nd	23rd	24th	25th	26th	27th	28th
		Meeting with Prof. Groß: Questions about genome mining			Finance meeting: Brainstorming about sponsoring. Discussion with mentors about raising money.	Social media meeting: introduction into Affinity software for editing our design scheme
29th	30th	31st

April

1st	2nd	3rd	4th	5th	6th	7th
				HP Meeting: Brainstorming of general project ideas	Finance Meeting: Application for Promega grant; List of all possible sponsors
8th	9th	10th	11th	12th	13th	14th
			Drylab Meeting: Brainstorming ideas and searching for local experts HP Meeting: Podcast Conceptualisation I	Meeting with student council regarding sponsoring for registration fee	HP Meeting: Podcast Conceptualisation II
15th	16th	17th	18th	19th	20th	21st
Finance Meeting: Application for sponsorship by the student council was accepted; Created excel sheet to keep track pf expenses; Created Skype account for receiving calls					Social media Meeting: internal organisation, responsibilities for the different social media channels
22nd	23rd	24th	25th	26th	27th	28th
Wiki & Design: Working on our Website Finance Meeting: Contact to biotechnology companies for sponsorship			Drylab Meeting with Elias from last year's team, he gave us tips on genome mining, molecular dynamics simulations and Drylab collaboration ideas HP Meeting: Getting in contact with Wüstewelle (radio channel) as well as Podcast Conceptualisation III		Meeting with Prof. Groß: Questions about genome mining
29th	30th
Finance Meeting: IDT registration; Discussion of the ordering list.	Drylab Meeting with Prof. Ziemert: Questions about genome mining and our project in general

May

1st	2nd	3rd	4th	5th	6th	7th
				Finance meeting: Created first list of laboratory material to order.
8th	9th	10th	11th	12th	13th	14th
		Drylab Meeting: Beginning work on Hidden Markov Models and Translating Genomes, further research of possible cyclotide candidates, and finding previous iGEM teams who did similar things		Finance meeting: phone calls sponsors; Contacted university administration for bank account. Conceptualisation of further sponsoring strategies. Application of second Promega grant. HP Meeting: Meeting with Team Ioaninna I		German meet-up organized by iGEM Bonn & Kaiserslautern
15th	16th	17th	18th	19th	20th	21st
					Social media meeting: talk about possible collaborations, planning of thank you posts for sponsoring HP Meeting: Scheduling Podcast Episodes	Finance Meeting: Planning of Meeting with the company brand. Organisation of applications for other iGEM grants.
22nd	23rd	24th	25th	26th	27th	28th
				Drylab: Collecting further HMM profiles from sites such as PFAM, beginning genome mining	HP Meeting: Conceptualisation and creation of the Podcast Intro	HP Meeting: Elias and Erik try to explain Biology to Kira
29th	30th	31st
		Video: Kick-off meeting

June

1st	2nd	3rd	4th	5th	6th	7th
	Drylab Meeting: Looking into the options of using Prosite profiles for genome mining, further work on HMMs and genome mining HP Meeting: Podcast Description, Logo Design and finalization of the Intro	Meeting with iGEM Ioannina Finance Meeting: Staying in contact with sponsors		Social media sub team: team building movie night
8th	9th	10th	11th	12th	13th	14th
Finance Meeting: Impact grand, Contacted Roche, Twist, Benchling and Corning	Drylab Meeting: Keeping it up with genome mining		Meeting with Alex Grundmann from Twist Bioscience to talk about sponsoring possibilities
15th	16th	17th	18th	19th	20th	21st
	HP Meeting: Setting tasks and roles for the Podcast, solving problems in the concept and preparing the next interview	Safety instruction for the Forchhammer lab Finance Meeting: Contacted Microsynth for sequencing labels	HP Meeting: Meeting with Team Ioaninna II	Organization Meeting: Plans for Sub teams HP Meeting: Trial interview with our Team
22nd	23rd	24th	25th	26th	27th	28th
Video: First day of shooting the promotion video (lab) HP Meeting: Meeting to schedule radio interview	HP Meeting: Regular discussion, setting new tasks, brainstorming more ideas	Video: Second day of shooting the promotion video (botanical garden, forest, lab) Finance Meeting: Impact grand, List of possible expenses; Ordering of gloves	HP: interview at Radio Wüste Welle	Drylab Meeting: Possible collaboration opportunity with iGEM Team IISER Kolkata; Thinking about how to edit cyclotides for different bioactivity; Molecular Dynamics Planning	Wiki & Design Meeting: Portfolio Printing; Postcard Collaboration Project from iGEM Team Düsseldorf; Wiki building begins	Video: Third day of shooting the promotion video (greenhouse); video editing begins
29th	30th
	HP Meeting: Discussion about modifications for the podcast, reflection on last meetings brainstorm

July

1st	2nd	3rd	4th	5th	6th	7th
Wetlab: bring inventory up to date and stuff we need to our lab space Finance meeting: Staying in contact with excellence cluster; List of chemicals HP: "Real" Interview with our Team	HP Meeting: Briefing on Üner Kolukisaoglu for the Interview	Drylab Meeting: Refocusing of project towards cyclotide grafting; Idea of pre-screening Wetlab constructs through simulations		HP: Interview Üner Kolukisaoglu		Video meeting: editing of the promotion video HP Meeting: Finalizing legal matters for the podcast, decision upon new HP Projects: field trip & panel discussion
8th	9th	10th	11th	12th	13th	14th
Finance Meeting: Ordering postcards for collaboration HP Meeting: Conceptualization of panel discussion in greater detail		Drylab Meeting: Homology Modeling of constructs; Learning about Molecular Dynamics with GROMACS		HP Meeting: Briefing on Nadine Ziemert for the Interview		HP Meeting: General organization, research for more interview partners as well as the field trip & panel discussion
15th	16th	17th	18th	19th	20th	21st
Wetlab: meeting with Prof. Brötz-Oesterhelt to talk about design and planning of the antimicrobial assays Interview with MolecularCloud to present our project Finance Meeting: Contacted Promega, IDT and Twist for laboratory materials		Drylab Meeting: Idea of coarse-grained simulations in conjunction to all-atom simulations Team building: barbeque		HP Meeting: Interview Nadine Ziemert
22nd	23rd	24th	25th	26th	27th	28th
First day in the lab. We prepared a lot of media and buffers.	Second day in the lab. We finished our preparations. The lab work can start! Social media meeting: planning of social media content for Promega silver sponsoring			Golden gate cloning level I (Assembly with BbsI-HF and transformation in competent E. coli) Inserts cloned:CtAEP1 (AEP) and AEP precursor (prec.)	Overnight culture of Level 1 constructs from Monday	Social media: video filming and editing for Promega as part of sponsorship contract for the Silver Grant Contact other possible sponsors, Ordering for Assays Miniprep of clones of the overnight culture; control digest with BAmHI and control agarose gel electrophoresis of level 1 constructs
29th	30th	31st
Sequencing of AEP and precursor constructs	Drylab Meeting with Dr. Thiel: Questions about Molecular Dynamics simulations; Supercomputer Access Autoclaving of consumables and media HP Meeting: Interview Andreas Dräger	Drylab Meeting: Getting access to supercomputer Cluster from Dr. Thiel; Planning of Molecular Dynamics simulations; Building of structures Social media: video editing for iJET collaboration and sending project information to iGEM Aix-Marseille for collaboration (guess the project name from emoticons) Team building: sports day with swimming, beach volleyball and table tennis

August

1st	2nd	3rd	4th	5th	6th	7th
	First meeting for writing of a review for the journal initiative from iGEM Maastricht Cloning level II: assemble AEP (with HA tag, 35S promoter, 35S terminator), the cyclotide precursor (with Myc-tag, 35S promoter, 35S terminator) and GFP (with 35S promoter, 35S terminator) into an LII vector backbone by Golden Gate reaction and transform the LII plasmids into E. coli (Erik, Teresa, Mirjam) HP Meeting: Meeting Team Taiwan I	Cloning level I: clone the synthesized AMP fragments into an LI vector backbone by Golden Gate cloning and transform them into E. coli; pick colonies from the level II vector transformation for overnight cultures (Erik, Teresa, Mirjam) HP Meeting: Maastricht Journal initiative meeting I	Miniprep of the LII cultures, control digest and agarose gel electrophoresis, send the plasmids for sequencing; cloning level III: assemble level II AEP, GFP and cyclotide precursors with promoters, terminators and tags into one level III vector: GFP and precursor for 2in1 constructs and AEP, GFP and precursor for 3in1 constructs; pick colonies from the level I vector transformation for overnight cultures (Erik, Teresa, Mirjam) Drylab Partnership Meeting with Kolkata: Kolkata will help us build our simulations, we will run them using our supercomputer access HP Meeting: Panel discussion meeting, working out deadlines and structur	Drylab Meeting with Dr. Thiel: Questions about supercomputer BinAC Miniprep of the LI cultures, control digest and agarose gel electrophoresis, send the plasmids for sequencing; pick colonies from the level III vector transformation for overnight cultures (Erik, Mirjam) Finance Meeting: Excellence Cluster	Miniprep of the LIII cultures, control digest and agarose gel electrophoresis, control PCR send the plasmids for sequencing (Erik, Anton) Meeting for review writing (journal initiative iGEM Maastricht)	Drylab Meeting: Talking about what exactly our Molecular Dynamics systems should look like
8th	9th	10th	11th	12th	13th	14th
	Set precultures of E. coli containing level III 3in1 and 2in1 constructs for Midiprep (Anton, Erik)	Midiprep of 3 colonies for each 3in1 and 2in1, control digest of the purified plasmids, send plasmids for sequencing with multiple primers (Mirjam, Teresa, Erik)	Golden Gate cloning level IV: clone c_blank, c_CHEN_1 and c_CHEN_6 into level III 3in1 and 2in1 vectors, Transformation of competent E. coli (Teresa, Erik) HP Meeting: General discussion, contacting Experimenta Heilbronn for a possible collaboration in the panel discussion	Pick colonies and set overnight cultures (Mirjam)	HP Meeting: Meeting with Thomas Potthast; Podcast details and explanation	Drylab Meeting: Further learning of Molecular Dynamics simulations, including coarse-grained and all-atom
15th	16th	17th	18th	19th	20th	21st
	Finance Meeting: Organization of meeting with contact of experimenta; Checked bank status	We transformed Agrobacteria with constructs: LIII 3in1 14, LIII 3in1 3, LIII 3in1 16, LIII 2in1 3, LIII 2in1 16. (Mirjam, Erik)	HP Meeting: General discussion of podcast, field trip and panel discussion	From the overnight cultures we picked 2 colonies per construct and inoculated those. (Mirjam, Erik)	We infiltrated 2 plants per construct and a negative control with AS medium. (Teresa, Catia, Mirjam) HP Meeting: first meeting with Dr. Robert Friedrich from the experimenta for a possible collaboration in the panel discussion	Drylab Meeting: Beginning work on supercomputer BinAC; Developing collaboration with iGEM Team CCU Taiwan
22nd	23rd	24th	25th	26th	27th	28th
Finance Meeting: Text for bank company; Ordering for excellence cluster	We checked the GFP expression under the fluorescence microscope.		We prepared media for the Assays and extracted the 3in1 14, 3in13 and the untransformed constructs from the leaves. (Mirjam, Catia) HP Meeting: meeting with experimenta II Subsequent discussion General discussion, trying to win the Dieter Schwarz Stiftung as a sponsor in exchange for working with experimenta	We carried out the Assays, to determine the effect of our crude extract on B. subtilis and E. coli in different media (BHI, MH) and different concentration. (Teresa, Alicia, Catia)	We checked the results of the forementioned assay by checking the absorbance under light of 600nm (Alicia)	Drylab Meeting: Running first simulations from iGEM Team IISER Kolkata Giant Jamboree fee paid
29th	30th	31st
	Erik prepared buffers for Wetlab Received consumables from biotech companies; contacted bank account manager	HP: Interview Thomas Potthast

September

1st	2nd	3rd	4th	5th	6th	7th
We extracted the infiltrated leaves that were prepared on the 20th august (3in1 14, 3in1 16 and untransformed samples). We then used these extracts to purify them via the included His-tag. Unfortunately, we could not measure proteins in the purified extracts. (Mirjam, Teresa, Erik, Alicia)	Since we could not detect proteins the day before, we repeated the his-tag purification with the flow-through from the day before. We then concentrated the elution fractions with dialysis and measured again the protein content of our solution. (Erik, Teresa, Alicia)	We quantified the amount of GFP in the cellular crude extract and flow-through (Teresa, Erik)			Tricine SDS-PAGE and western blot, set overnight cultures of agrobacteria with 3in1 c_blank, c_CHEN_1 and c_CHEN_6 (Mirjam, Teresa, Erik) HP Meeting: Collaboration: Interview with Team CCU Taiwan	Continue with the western blot, set new overnight cultures of agrobacteria with 3in1 c_blank, c_CHEN_1 and c_CHEN_6, protein quantification assay (Mirjam, Teresa, Erik)
8th	9th	10th	11th	12th	13th	14th
HP Meeting: Discussion about experimenta, podcast, collaboration Team Taiwan	HP Meeting: Meeting with experimenta III, getting advice on the panel discussion, working on the collaboration				Drylab Meeting: Discussion of Presentation Video Ideas; Running coarse-grained simulations for force-analysis; Collaboration CCU Taiwan Structure Prediction Protein extraction of constructs 3, 14 and 16 using 12 leaves of each plant was done. Protein extraction of constructs 3, 14 and 16 using 12 leaves of each plant was done. We purified all 3 constructs via His tag affinity purification and dialysis and quantified protein concentrations of all constructs afterwards. (Erik, Inga, Julius) We also carried out cloning of Level III vectors (constructs 9 and 18) with subsequent transformation of E. coli. (Erik, Adrián, Laura, Dennis, Julius, Inga)	Wiki & Design Meeting: Writing of Wiki pages delegated to sub teams, and further planning regarding Wiki We prepared samples for the SDS-PAGE and inoculated 2 of the colonies which have grown over night. An inhibition zone assay using our purified protein suspensions was carried out. (Laura, Dennis, Adrián, Mirjam, Inga)
15th	16th	17th	18th	19th	20th	21st
We extracted and purified plasmid DNA from the overnight cultures using the Monarch Plasmid Miniprep Kit from NEB. We assembled Level III vectors using the purified plasmid DNA of constructs 9 K1, 9 K2, 18 K1 and 18 K2 and transformed them into E. coli. As yesterday's inhibition zone assays did not work, we repeated them with E. coli und B.subtilis. We tested different synthetic peptides as well as crude extracts or purified solutions of our grafted peptides. We also used the purified protein solutions for SDS-PAGE with subsequent instant blue staining or Western Blot. (Laura, Dennis, Dilara, Adrián) HP Meeting: Finishing draft expose and invitation mails for the panel discussion	As the inhibitions zone assay of yesterday yielded no significant results, we repeated them with higher concentrations. Level I plasmids were digested with HindIII and analyzed gel electrophoresis. Overnight cultures of Level III plasmids were inoculated. We continued yesterday's Western Blot and carried out the detection as well. For purification, we carried out an acetone precipitation for several crude extracts as well already purified protein solutions. (Laura, Dennis, Dilara, Adrián, Erik, Julius, Inga) HP Meeting: Meeting concerning the field trip to icon genetics/nomad bioscience in Halle	We extracted and purified plasmid DNA of yesterday's overnight cultures using the Monarch Plasmid Miniprep Kit from NEB. Afterwards, a control digest with subsequent gel electrophoresis was carried out. As yesterday's Western Blot showed no significant signals, the membrane was washed and incubated with primary antibody over the weekend at 4°C. (Erik, Inga) Taking team pictures for Wiki	Drylab Meeting: Further progress on coarse-grained umbrella simulations; Running and Results of Molecular Docking; Further work with IISER Kolkata	Social media meeting: planning for Wiki, other plans for possible interviews	We finished our western blot and purified our constructs via His-tag purification. Replication of gel of control digestion of construct LIII 3in1 9 &18.	Agrobacteria transformation with 3in1 9 & 18. SDS-PAGE and Western blot of constructs 3in1 3 & 14. HP: Field trip to icon genetics/ nomad bioscience in Halle
22nd	23rd	24th	25th	26th	27th	28th
Continuation of yesterday's labwork. HP: Field trip to icon genetics/ nomad bioscience in Halle	Drying of leaves infected expressing 2in1 & 3in1 16 and p19. Overnight culture of transformed Agrobacteria. HP: Field trip to icon genetics/ nomad bioscience in Halle	Extraction of 2in1 & 3in1 16 and p19 constructs (according to Poon et al., 2018). Taking team pictures for Wiki	Drylab Meeting: Finishing of Taiwan Structures, starting simulations		Wetlab: Extraction of proteins from plant material directly with Lämmli buffer (untransfected, p19 control, constructs 2in1 3&16, 3in1 3,14&16) SDS-PAGE with these samples plus from the extraction from 24th September, Coomassie staining of one gel, start of Western Blot with 1. Ab anti-His-Tag Transformation of A. tumefaciens with plasmid with construct 3in1 9	Wetlab: Finishing and detection of Western Blot, repeating of SDS-PAGE with same samples, Coomassie staining of one gel, start of two Western Blots with 1. Ab anti-His-Tag and 1. Ab anti-HA-Tag microdilution assay for antimicrobial activity testing with our synthesized reference peptides, CHEN and KR-12
29th	30th
Wetlab: Finishing and detection of Western Blots, read out of assay from yesterday Wiki Writing Day (the team came together and continued working on the Wiki pages) HP Meeting: General discussion, solving questions concerning Wiki and panel discussion	Wetlab: Inoculation of overnight cultures of A. tumefaciens with colonies of constructs 3in1 9, 3in1 18 and 3in1 16

October

1st	2nd	3rd	4th	5th	6th	7th
Wetlab: Infiltration of N. benthamiana plants with O.N. cultures from the day before and p19 vector as control	Wiki Writing Day 2 (the team came together and continued working on the Wiki pages) Drylab Meeting: Recap of coarse-grained simulations HP Meeting: Meeting with experimenta Heilbronn IV		Wetlab: harvesting of plants from Friday, fluorescence microscopy of whole plants to check infiltration efficiency, brightfield and GFP filter microscopy of single infiltrated leaves, protein extraction from plant tissue with new extraction buffer (Teresa, Erik) Western Blot: re-incubate membranes from last week with freshly prepared primary antibody solutions (anti-His-Tag and anti-HA-Tag) HP Meeting: Contacting the Dieter Schwarz Stiftung	Wetlab: SDS removal via acetone precipitation, microdilution assay for determination of antimicrobial activity of samples from 4th October (Alicia, Erik, Catia)	HP Meeting: General discussion and recap Wetlab: SDS-PAGE and subsequent either Coomassie staining or western blot with samples extracted with the new extraction buffer from the 4th, read out of the antimicrobial assays from the 5th (Catia, Erik, Inga)	Wetlab: washing and detection of the western blots from the 6th (Alicia, Catia) HP Meeting: Meeting with experimenta V
8th	9th	10th	11th	12th	13th	14th
Wetlab: extraction of protein from gel slices cut out of from SDS-PAGE gels for further analysis via MALDI-TOF/MS (Erik, Inga)	Drylab Meeting: Analysis of Membrane Simulations; Recap Meeting Kolkata	Video Meeting: Presentation Video, first day of shooting	Video Meeting: Presentation Video, second day of shooting HP: Interview Heike Brötz-Oesterhelt Wetlab: MALDI-TOF/MS experiments (Erik, Teresa)
15th	16th	17th	18th	19th	20th	21st
			HP: Meeting with experimenta (VI) as preparation for panel discussion		HP: panel discussion in cooperation with experimenta

About Us

We are the iGEM Team Tuebingen, a group of motivated students who are working on creating a fast screening platform for stabilized peptides. We are aiming to provide a system that gives everyone the ability to stabilize peptides such as antimicrobial peptides to create better medical agents.