June
Week 1 [17/6/24 - 21/6/24]
Monday 17th of June
-
Lab induction [everyone]
- Writing project description [Ruby, Ellin]
- Allocating team roles [everyone]
- Planned Rosetta experiment [George, Oli, Becky, Yara]
- Began developing the wiki [Anouska, Nigella, Ben] using HTML - pages and links were created, and images were added.
Tuesday 18th of June
- Rosetta experiment planning continued [George, Oli, Becky, Yara]
- Continued writing project description [Ruby, Ellin]
- Protein purification practice experiments took place [Ruby, Yara]
- Culturing and incubation for Rosetta experiment [George, Oli, Becky]
- Initial draft of ethics form produced [Ellin]
- Began work on project branding - developing a colour scheme [Anouska, Nigella], thinking up some names [Anouska, Nigella] and creating some logo ideas [Ben].
- Edited the CSS style sheets for the wiki, allowing colours and fonts to be changed.[Anouska, Nigella, Ben]
- Created a 'wiki cheat sheet' PowerPoint containing the basics of HTML [Ben] and CSS [Anouska] to help the rest of the team.
Wednesday 19th of June
- Rosetta cultures failed to grow! Too many heat shocks were performed... protocol from yesterday was repeated [George, Oli, Becky].
- Project description was further developed, following a meeting with John [Ruby].
- Researches mycobacterium tuberculosis bovis quorum sensing, via academic papers [Ellin].
- Added the notebook page to the wiki, with attributions [Ben].
- BL21 transformation was attempted, with Cas 12 and Cas 13a plasmids [Louise, Tom]
Thursday 20th of June
- Western blot practice experiments were performed [Ruby, Yara]
- BL21 experiment for Cas 13a was performed successfully, but not for the Cas 12. The Cas 13a is now being grown in a liquid culture for a gel, ready for an experiment of Friday [Louise], while the Cas 12 is being transformed again [Tom].
- Project description was finalised [Ruby]
- Western blot lab protocol was refined [Yara]
- mCherry was inoculated into liquid culture [George, Becky]
- Agar plates were produced, with varying concentrations of ampicillin and chloramphenicol [Oli, George, Becky]
- More HTML tweaks were performed to better organise certain pages [Ellin, Ben]
- The notebook page was reworked to contain a photo diary and collapsible sections, and updated with additional attributions [Ben]
- A new template for wiki text boxes was developed, with shadow effects and curved edges [Anouska]
- Comprehensive aesthetic changes (colours, container shapes, fonts, and menu bar appearance) were effected [Anouska, Nigella]
- More gRNA literature was researched [Ellin]
Friday 21th of June
- Group meeting to discuss the week and plans for week 2 [everyone]
- A tweak to the CSS left all the pictures on the notebook wiki page random sizes - this was fixed, and dividers were created [Ben]
- General wiki edits, tidying up some of our previous HTML experiments [Ben, Nigella, Talia, Anouska]
- mCherry Rosetta culture was plated onto agar-antibiotic plates [Oli, Becky, George]
- mCherry Rosetta culture was placed into a checkerboard 64 well plate. Fluorescent intensity was then measured for photon wavelengths 587-610 nm. The OD (optical density) at 600 nm is being measured as well [George, Becky, Oli].
- Rosetta experiment was written up, as part of our contribution section of the wiki [Becky]
- The Western blot experiment didn’t work [Ruby, Yara]
- Ran a practice Qubit protein concentration experiment [Ruby, Yara]
- Some of the non-wiki team began learning HTML [Ruby, Yara]
Wiki development is underway!
Growing our first Rosetta culture - the mCherry batch (left) has come out a lot better.
Ruby hard at work.
Very exciting when the results come back!
Week 2 [24/6/24 - 28/6/24]
Monday 24th of June
- Integrated human practices were researched [Ruby, Talia]
- Contact was made with farmers, via John [Ruby]
- Past wikis were scoured in search of inspiration [Ruby]
- The team page was reformatted and filled out [Ben], using a new layout
- The logo was given tweaks, The new name was added (BoviTect) and the menu bar was tweaked [Nigella, Anouska]
- Notebook sections were expanded to include week 2, and the images were properly resized (trust me it's harder than it sounds) [Ben]
- Rosetta has a lower chloramphenicol resistance than expected as the cultures didn't grow on the agar, either with cam present or in the wells. Agar plates with just amp or lower concentrations of cam have now been produced, and the mCherry rosetta cultures will plated on them tomorrow. [Oli, Becky, Ellin]
- Learning HTML and editing the wiki [Yara, Ruby]
- Research into presentation and promotional video requirements [Yara]
Tuesday 25th of June
- Ethical approval form & rewrite info sheet and consent form for 2024. [Ellin, Anouska]
- Produced overnight mCherry Rosetta culture, which will then be plated on the individual antibiotic plates tomorrow, and again left to grow overnight. We should have results from this experiment on Thursday. [Oli, Becky]
- IHP Research and learning to code the wiki. [Ruby, Yara]
- Re-nickling columns ready for BL21 protein purification tomorrow. [Ruby, Yara]
- General wiki tweaks, mostly editing the teams page. [Ben]
Wednesday 26th of June
- Plated up cultures on the antibiotic agar plates… and now we wait. [Becky, Oli]
- Meeting with Dr John Love. [et al]
- Final ethics form completed [Anouska]
- Installed flutter, to begin app development. Researched modelling possibilities. Drafted hardware concepts. [Oli, Ben, Talia]
- Writing the lab protocol for transformation for publication on the Wiki [Becky]
- Wiki tweaking [Nigella]
- Making up buffers for protein purification [Yara, Ruby]
- Lysed cells for protein purification [Louise, Tom]
- Made liquid culture for mCherry and blank plasmid [Louise]
Thursday 27th of June
- Rosetta grew on some of the agar plates. Our previous question of Rosetta not being very chloramphenicol resistant, is confirmed to be true, as no cultures grew on any of the plates with just chloramphenicol, besides the 1.1μg/mL conc plate; This yielded a similar result to the control, implying that it had no effect on growth. [Oli, Becky, George]
- Inoculated mCherry and blank plasmid into autoinduction media [Louise, Anouska]
- Miniprep of the mCherry and blank [Tom]
- COSHH form done for autoinduction media [Louise]
- Attempting to put in animations using JSON and Lottie [Anouska]
- Redesigning logo for letter headings, etc. [Anouska, Nigella]
- crRNA attempting to attach the cr and tracrRNA [Tom]
- The checkerboard test will need to be repeated 2 more times. [Oli, George, Becky]
- Protein purification of Cas 12a and Cas 13, renickeling column and running 2x Qubits [Yara, Ruby]
Friday 28th of June
- Optimized the process for producing graphs for our checkerboard test, by using matplotlib in conjunction with for loops in python. Produced dose response curves for ampicillin. [Oli]
- Lysed cells for protein purification mCherry and pX1900 [Louise]
- Working on the Wiki: Back to top button, designing animations [Anouska]
- Drawing graphics for wiki using Procreate + wiki tweaks [Nigella]
- Lab photography for wiki [Yara]
- Purification of the blank and mCherry controls [Yara, Anouska, Nigella]
- Writing questions for interviews (Integrated HP) [Becky, Talia]
- Learning how to use dartpad for app development[Talia]
- Completing attaching tracrRNA and spacer sequence together (with added buffer region) and calculating crRNA shapes [Tom]
Wiki development continues!
Anouska, Nigella and Yara carrying out the purification of the blank and mCherry controls through the nickel columns!.
Actual (left) and simulated (right) results of a test to check for the presence of our plasmids. Left-to-right, these are Cas13a, Cas12, mCherry and a blank control plasmid.
To keep morale high, the occasional team-building activity has taken place as a brief respite from iGEMing - such as this impromptu exercise session...
July
Week 3 [1/07/24 - 5/07/24]
Monday 1st of July
- Re-designed non-biological hardware concept (using suggestions from the Dundee 2013 team), so that the challenge of implementing a phone camera into the design can be neglected all together; instead focusing on light-sensitive photodiodes connected to a raspberry pi, which can send signals to a mobile device through an app. This should be much easier to implement from both a hardware and software perspective [Oli]
- Transformed Ampicillin-resistant, mCherry Rosetta again for our 8x8 well plate. Also plated Rosetta onto 2 separate agar plates containing Chloramphenicol from different stocks, to determine if there is a contamination issue. [Oli, George]
- After facing issues with the desktop Android Studio, app development was resumed on the online version [Ben]
- Writing more interview questions and short explanation of our project for each person we interview [Becky]
- Qubit concentrations for Cas12a, Cas13, mCherry and blank plasmid [Yara, Nigella]
- Wiki tweaking [Nigella]
- Stash design, setting up team page [Anouska]
- Redid a final agarose gel for all the plasmids, looks pretty [Louise]
- Writing up the protocols for plasmid to protein pipeline [Louise]
Tuesday 2nd of July
- Starting on CAD design for hardware [Anouska]
- Meeting with Becky for help with coding to fix parts of Wiki (sticky contents bar, header background) [Nigella, Anouska]
- Drew final version of wiki banner [Nigella]
- Fixed code for DNA sequences. Sequences can now be printed with their complementary strain and their inverse. [Oli]
- Produced overnight liquid cultures for 8x8 well plate tomorrow. Also verifying that ampicillin-resistant mCherry plasmid cannot be expressed in chloramphenicol environment. [Oli, George]
- Ran a western blot on our lovely 34 samples [Ruby, Yara]
- Had a meeting with Mark and Chloe on how to optimise our protein expression, cell lysis and western blot [Ruby, Yara]
- Became a lab assistant for the morning and was Very Helpful ⭐ (except when I forgot to change the pipette end) [Ben]
- Continued Flutter development - now working on accessing the camera through the app [Ben]
- BLAST sequencing of genes [Becky]
- Set up the team email, added a signature and started drafting emails to people we would like to interview [Becky, Louise]
- Lab photography [Yara, Ruby]
Wednesday 3rd of July
- Update from Chloe on the lab work required over the next few weeks and allocating dates for everyone to be in the lab [everyone]
- Working on new version of hardware on CAD [Anouska]
- Stash orders and design [Anouska]
- Image hover on the Wiki - modifying teams page [Nigella]
- Western blot results - WE GOT CAS!!! [Yara, Ruby]
- Restriction digest with linear DNA that is maltose binding protein [Ruby]
- Running agarose gel [Ruby, Becky]
- Running an SDS-PAGE on the pellet to see if more Cas proteins were there [Louise, Ben]
- Uploaded some photos [Louise]
- Lab protocol write ups from last week's lab work [Yara]
- Showed Louise and Ben how to do SDS-Page [Yara]
- Produced liquid mCherry Rosetta culture from plates yesterday, and started to run our 8x8 well plate protocol overnight. [George,Oli]
Thursday 4th of July
- E.coli transformation into BL21 and Rosetta. Writing up protocols for restriction digest, gel excision, DNA extraction + ligation [Ruby, Becky]
- Ethics research and finishing protocol write up [Louise]
- Drafted emails for human practice interviews. Used flutter to continue with software development [Talia]
- Made a python script for detection of cas12a spacer sequences. Tested spacer sequences for potential DIVA opportunities and for false positives with the cow genome [George]
- Finished finding the cas12a spacer sequences [Tom, George]
- Modifying wiki page layouts [Nigella]
- Adding image transition to team page on wiki [Anouska]
- Lab protocol write ups from last week's lab work. Wrote an explanation about Cas spacer sequences [Yara]
- Found the surrounding sequences for the spacer sequences (50bp up and downstream of spacer sequence complement) [Yara, Nigella]
- Made the wet lab timeline colour-coded by project (see under lab protocols tab) [Ruby]
- 8x8 well test again. Performed Rosetta transformations with 5 different plasmids and a negative control. Plated onto 3 separate types of agar plates: Ampicillin, Chloramphenicol, and ampicillin + Chloramphenicol. [Oli, George]
Friday 5th of July
- Observed that the promoter seems to affect mCherry expression on ampillin + chloramphenicol plates. pX1800_PJ23100-mCherry and pX1900_PJ23100-mCherry didn’t produce any purple cultures on the plates, whilst pX1900_T7-mCherry did. This implies high copy numbers affect the mCherry expression. 8x8 well plate again as well… [Oli, George]
- crRNA using introns only is completed, there is still some concern about where these sequences sit within the cow genome (cas13a). The bTB (cas12a) sequences have worked really well [Tom]
- MBP plates didn't grow except for 1- the DH5alpha cas-13 plate [Ruby, Becky]
- Made complete protein purification to analysis method (including nickel regeneration, making buffer, protein purification, qubit and western blot). Updated all protocols for the Wet lab section of the notebook [Ruby]
- Wrote up protocol and results for Rosetta experiments [George]
- Emailed companies about potential interviews [Becky]
- Set up social media for bovitect and created posts/content [Talia]
- Lab protocol write ups and making results tables for last week's experiments [Yara]
- Looking at safety forms and information on what is allowed [Louise]
Becky readying plates for over-night Rosetta growth.
Some wiki work with the rest of the group. Anouska is working, don't worry.
Thermocycling Rosetta and BL21 derived cas protein samples in preperation for a western blot!
Western blots indicating CAS presence
Week 4 [8/07/24 - 12/07/24]
Monday 8th of July
- Analyzed data from last week. Data is inclusive, however we have shown, on Agar plates, that Rosetta transformed with high copy number promoters results in mCherry protein not being expressed when in the presence of ampicillin and chloramphenicol. [Oli, George]
- Rewrote all of the IHP documents and made the comprehensive guide to IHP [Ruby]
- Prepared sterile LB broth for induction of Cas later this week, and prepared Ampicillin LB-agar plates for transformation. Troubleshot crRNA sequences, hopefully no longer inverting weirdly [Tom]
- Transformed Cas13a, Cas12a, T7-mCherry, T7-Px1900 into Rosetta and BL21 E.Coli, aiming to produce a greater yield of Cas' [Becky, Yara, Tom]
- Finalising results tables of Qubit and Western Blots from Week 2 and 3 [Yara]
- Communicating with University of Warwick about the UK iGEM meet-up [Becky]
- Made liquid cultures for the maltose binding practical [George]
Tuesday 9th of July
- Plasmid Miniprep of cells that contain plasmids with MBP and Cas13 proteins (TBC using sequencing). Qubit of plasmids containing MBP and Cas13 proteins [Yara]
- Prepared Amp/Clam antibiotic aliquots for induction cultures tomorrow [Becky, Tom]
- Prepared IPEG solution for induction of T7 tomorrow, then started methods for probe testing. [Tom]
- Completed Fluorescein concentration curve and created graphs [Oli, George]
Wednesday 10th of July
- Made publishable graphs for fluorescein concentration curves from yesterday at different gains. Drafted COSSH form for RNAse and Dnase Alert kits [Oli]
- Inoculated induction culture media's (250 mL) with BL21 and Rosetta overnights (Cas12a, Cas13a, mCherry, PX1900 - note no PX1900 for Rosetta), monitored growth for 4 hrs and induced T7 expression at 0.6 OD. [Becky, Tom]
- Prepared for cell harvest on Thursday [Becky, Tom]
- Lab photography [Yara]
- Qubit of plasmids containing MBP and Cas13 proteins [Yara]
- Preparation of competent Rosetta cells
- Plasmid mini prep [Anouska, Nigella]
- Ethics resubmission [Anouska]
Thursday 11th of July
- Catching up and writing a section of 'All about bTB', basically consist of current knowledge, impact, statics, current test about bTB [Ellin]
- Harvested induced cells and freezed the pellets for weekend storage [Becky, Tom]
- Measured the fluorescence of simulated blood and PBS mixed with fluorescein. We have shown that the blood significantly blocks the signal [Oli, Tom]
- Preparation of competent Rosetta cells and storage in freezer – Day 2 [Yara]
- Editing photos from the lab [Yara]
- Made aliquots of Ampicillin for future use [Becky]
- Various tweaks to the Wiki, mostly involving the Teams page photo optimisation [Ben]
Friday 12th of July
- Found method of pairing flutter apps with Arduino via Bluetooth, for our hardware [Oli]
- Made publishable graphs from the blood vs PBS sensitivity tests [George]
- Planned out experiments for testing probes [George]
- More miscellaneous wiki tweaks [Ben]
Cultivating and harvesting Rosetta and BL21!
mCherry is a Roestta culture showing up strongly in the dark.
Tom pipetting simulated blood into flouroscein and PBS to measure amount of flourescence!
Week 5 [15/07/24 - 19/07/24]
Monday 15th of July
- Set up a Planner page to keep track of progress [Oli], and tasks were added to monitor progress [everyone]
- More photos of the team at work were taken for the wiki, and the notebook page was brought up to date [Ben]
- Meeting [everyone]
- Finished off old protocols from last week, organised stuff for this week on Planer, prepared column buffers for tomorrow [Tom]
- Hardware design [Anouska, Oli]
- Transformation of pX1900 P_J23100-mCherry E Coli [Oli]
- IHP question brainstorming [Anouska, Oli, George]
- Email and research about the use of blood [Anouska, George]
- Refining the protocol for testing the probes and carrying out the first part of the practical [George]
- Made up new nickel columns for protein purification [Louise, Yara]
- Planning engineering section of wiki [Ellin]
- Transforming Cas12a and Rosetta [Ellin, Nigella]
- Editing footer of wiki (linking wiki to Instagram and Email) [Nigella]
Tuesday 16th of July
- Protein purification of Cas12a, Cas13, PX1900 and mCherry expressed in both Rosetta and BL21 [Yara, Ellin, Ben]
- Testing 'cas' concentration needed for final experiment for both DNA and RNA [George, Anouska, Oli]
- Graphing 'cas' concentration experiment [George]
- More interview questions and wiki tweaking [Nigella]
- Overnight for our new competent Rosetta E coli [Oli]
- Hardware iteration design adding more epindorf openings (from 2 to 4) [Anouska]
- Lysed Rosetta and BL21 cells ready for protein purification [Tom, Louise]
- Remade contaminated Tris-HCl buffer solutions and researched methods for detecting RNA. [Tom]
- Resumed Flutter work, making a Doctor Who app (a test to work on my programming skills before tackling the cow app proper) [Ben]
Wednesday 17th of July
- Writing up a check in form for potentially bringing Cas out of the lab [Louise]
- Made amp plates [George]
- Changed the 'cas' concentration experiment graph into fluorescein concentration equivalents and performed statistical analysis on it [George]
- Group meeting catching everyone up with the different areas of the project [Everyone]
- Western blot of Cas12a, Cas13, PX1900 and mCherry expressed in both Rosetta and BL21 – due to use of incorrect antibody, need to repeat tomorrow [Yara, Ellin]
- Poster for Warwick meet-up [Oli, Anouska, Nigella, Talia]
- Completed the Doctor Who app and started laying the foundations for the Cow App in Flutter, which farmers will use to get test results [Ben]
Thursday 18th of July
- Finishing Warwick poster [Anouska, Oli, Nigella, Ruby]
- Writing up detailed notes of our lab progress so far [Anouska]
- Updated wet lab timeline [Ruby]
- Description of Chloramphenicol-Ampicillin experiments for the Wiki, along writing the next steps for the experiment going forward. [Oli]
- Engineering cycle [Ruby]
- Making competent Rosetta cells day 1 [George, Oli]
- Research into ethics of using blood further and research of making our own model blood [George]
- Second western blot [Ellin, Yara]
- Flutter work: making multiple (blank at this stage) pages of the Cow App, and decorating the home page [Ben]
Friday 19th of July
- Fixing menu bar on wiki [Anouska, Nigella]
- Protocol draft for photodiodes and fluorophore quencher system [Oli]
- Western blot of proteins in other BL21 fractions [Ellin, Yara]
- Finished Check in form for potentially taking the Cas proteins outside the lab [Louise]
- Started attributions page [Louise]
- Competent Rosetta E. coli prep [George, Oli]
- Carried out gel extraction, qubit quantification and started write up [Tom]
- Set up the camera page on the Cow App in Flutter [Ben]
Trying to figure out how hinges function in order to draw then in CAD for our hardware design!
Our weekly meeting catching everybody up to date on the progress of different parts of the project!
Ben keeping a close eye on his protein purification.
More protein purification pictures
Week 6 [22/07/24 - 26/07/24]
Monday 22nd of July
- Concentration curves for Sulforhodamine 101 made and more concentrations of probes tested with optimal Rnase concentration found [George]
- Repeated PCR for Cas13a DNA test sequences and ran a 1.5% agarose gel, with blanks, to see if dimer primers where forming. Then gel extracted/PCR cleaned all samples. [Tom]
- Transformed Rosetta with Cas plasmids, mCherry, blank plasmid and nothing, to check for new rosetta contamination and for protein expression this week [Louise, Ellin]
- Lab photography [Yara]
- Lab protocol write up and presenting work in results tables [Yara]
Tuesday 23rd of July
- Team meeting with John, Mark, Chloe and Yik Ting
- Produced circuit diagrams for the photodiode detection system part of our hardware. Code for the voltage readings has been produced as well. [Oli]
- Graphed and completed the experiment of differing DNase concentration effects on the probes and completed the effects of varying probe concentration on RNase [George]
- Drawing up new hardware iteration of Fusion [Anouska]
- Research for Dr Rees interview [Anouska, Talia]
Wednesday 24th of July
- 3D printer Test [Anouska, Oli]
- Editing hardware on CAD [Anouska]
- Preparing for interview with Dr Rees [Talia, Anouska, George, Tom]
- Made week 6 update powerpoint for interview with david andrews [Ruby]
- Waiting on approval to use a physics lab to test parts of our hardware [Oli]
- Further work making the graphs better and creating a model using the graphs [George]
- Made up and sterilised LB media for our large induction cultures [Louise]
- Built a puzzle for team building purposes [Becky, Ben]
- Finally got Bluetooth page to the stage where the app could compile - however there were still errors present [Ben]
- Made up ITPG solution [Ellin,Tom]
- Carried out PCR clean up and ran another Tapestation gel column, then put results into a figure for wiki. [Tom]
Thursday 25th of July
- Team interview with David Andrews [Everyone]
- Updating wiki notebook [Anouska]
- Warwick meet-up [Ruby, Becky]
- Made up induction media and started incubating [Louise, Tom]
- Wrote summary of hardware concepts as well as what we want to achieve with our Arduino. Physics lab induction, just need to wait for components to arrive [Oli]
- Planned out my experiments for cas, probes and crRNA all working together [George, Tom]
- Making diagrams for how the test functions [George]
- Prepared T7 transcription reaction mixtures for tomorrow
- Induced expression of BL21 with Cas12a with IPEG [Ellin]
Friday 26th of July
- Warwick meet-up [Ruby, Becky]
- 3D Printer troubleshooting and ordered components for our hardware [Oli]
- Interview with Actiopohage (Cath Rees) [Talia, George, Tom]
- Triallled the full experiment with cas, probe, crRNA and target [Tom and George]
- Carried out transcription and Tapestation RNA gel, to confirm RNA was made [Tom]
- Making up buffer for nickel columns [Yara]
- Harvested cells for lysis to freeze over weekend [Louise]
- Extract, harvest, and freeze pellet [Ellin]
A few members of the team are climbing fans. Here Yik Ting (one of our supervisors) is photographed during a session.
First time 3D printing one of our hardware prototypes!
Ruby shows off our work so far at the iGEM meetup in Warwick!
Week 7 [29/07/24 - 2/08/24]
Monday 29th of July
- Fixed circuit diagram (components now actually have a power supply 🤦♂️), 3D printer cleaning and troubleshooting again… [Oli]
- Emailed farmers and CHeCS about setting up interviews [Becky]
- Lysis of Cas 12a/13a BL21 large expressions [Louise, Ellin]
- Nickel column protein purification of Cas12a and Cas13a in BL21, couldnt continue purification as the columns were very clogged up, due to the use of large volumes of cell lysate [Yara]
- Continued the protein purification until 8 pm!! [Chloe]
- Lab and 3D printing photography [Yara]
- Emailed about the properties of Thermo Fisher's probes [George]
- Added protocols onto one note (8x8 well plate, ampicillin plates, making stock solutions of antibiotics) [George]
- Worked on diagrams to show how the project works [Ruby, George]
Tuesday 30th of July
- After small tweaks to the printer calibration, the phone is finally being printed [Oli]
- Update wet lab timeline [Yara]
- Western blot of Cas12a and Cas13a, cultured using IPTG and autoinduction respectively [Yara, Ruby]
- Finished graphics and description for the project, cas12a & cas13a [Ruby]
- Responded to check in form and did another for blood, plus emailing [Louise]
- Draft COSHH form for blood [Oli]
- Organised and graphed results of previous experiments [George]
- Research for the human practices section of the wiki [Becky]
- Transformation of BL21 with Cas 12a, 13a, and PX-1900 [Ellin]
- Ran RNAase contamination experiments on RNAase free water and elution buffer [Ruby]
Wednesday 31th of July
- Meeting deciding project direction after evil western blot with Chloe and Mark [Ruby, Becky, Lousie, Yara, Talia, Ellin, George]
- Update wet lab timeline [Yara]
- Physics lab photography [Yara]
- Wrote protocols for DNA/RNA contamination experiment [Ruby]
- Second protein purification of Cas12 using nickel column and purifying again with PD10 column [Ellin]
- Spoke to Dan and wrote up the ideas about amplifying signal or removing background noise [George]
- Wrote up protocols onto one note (Fluorescein concentration curve, Suflorhodamine concentration curve, making buffers, testing different enzyme concentrations for Rnase as well as the developments) [George]
- Moved 3D printer to cooler area and started test print again [Oli]
- Fluorescein photodiode and LED experiment: testing arrangements of LED to photodiode, as well as signal response from varying concentration of fluorescein [Oli]
- Transcription of Cas12a crRNA hsp20 sequence [Yara]
- Ran DNAase contamination experiments on 'RNAase free water', elution buffer and milliq [Ruby]
- Ran Sulphurholomine blood experiments- red blood is a no go [Ruby]
Thursday 1st of August
- Cleaned up successful 3D print. Added updates to Hardware section in the notebook. Learning AutoCAD to produce a frame for a gel-based BP-Filter [Oli]
- Did a tapestation of Cas12a crRNA hsp20 transcript, as well as transcription and tapestation of Cas13a crRNA BCL2_a and Cas13a target BCL2_a [Yara]
- Brainstormed ideas for our promotional video [Becky]
- Ran DNAase contamination experiments on milliq and autoclaved milliq [Ruby]
- Ran DNAase contamination experiments on milliq from downstairs, equilibriation buffer and highland mineral water [Ruby]
- Western blot [Ellin]
Friday 2nd of August
- Hardware experiments with perpendicular arrangement and bandpass filter [Oli]
- Ran RNAase contamination experiments on equilibriation buffer and milliq [Ruby]
- Organised team visit to farm with david andrews [Ruby]
- Team meeting/lab update [Ruby, Becky, Louise, Yara, Ellin, Oli]
- Finishing off western blot and clear out nickel columns [Ellin]
- Ran an agarose gel of the transcribed Cas13a crRNA BCL2_a and Cas13a target BCL2_a samples [Yara]
- Making buffers for next weeks experiments [Lousie, Becky]
Testing our hardware circuits!
Results from Cas12a crRNA transcription
To work with our app, we had a go at 3D printing a box to contain our samples to cut out external light and allow maximum contrast for photos. Here is the first prototype.
August
Week 8 [5/08/24 - 9/08/24]
Monday 5th of August
- Lysis of BL21 with Cas13a [Louise]
- Protein purification [Nigella, Louise]
- Wrote up protocols onto one note (Testing different enzyme concentrations for Dnase, Testing different probe concentrations for Dnase and Rnase and trial of whole test put together as well as the developments) [George]
- Completed the blood COSHH form [George]
- Researched experiments for how to make plasma from blood [George]
- Remaking nickel columns [Becky, Yara]
- Transcription, tapestation and agarose gel write up [Yara]
- Went on the hunt for video equipment and sent out some emails. Drafted some initial ideas for the intro video. [Ben]
- Prep for farm trip tomorrow [Ellin]
- Going over and processing data of the 96 well plate contamination experiments [Ruby]
Tuesday 6th of August
- Wiki tweaking: fixing formatting issues [Nigella]
- Western blot [Yara, Talia, Ben, Nigella]
- Scouting locations for team/portrait photos and taking practise shots [Yara, Nigella]
- Updated wet lab timeline [George]
- Research on how to separate blood, SHERLOCK system and fluorophores [George]
- Went and interviewed David and the other staff at Warson beef [Louise, Ruby, Becky, Ellin]
- Collected dung and sputum samples [Louise, Ruby, Becky, Ellin]
Wednesday 7th of August
- Drafting email to local MPs requesting for interviews [Nigella]
- Had a gander around the campus to construct a storyboard for our promotional video [Ben, Becky]
- Helping out with promo video structure and script, looking through videos from previous years [Nigella]
- Filtered smaller chunks out of our purified proteins with size exclusion centrifuge columns [Louise, Ellin]
- SDS page and western blot [Ellin]
- Writing up protocol improvements [Louise]
- Talked to Dan Barber and Mark about how to improve tapestation results [Yara]
- Transcription of Cas13a crRNA/target HBEGF_a, using nuclease free water. Tapestation performed using improved protocol, including running RNA control and original DNA template [Yara]
- Planning experiments with blood along with further research [George]
Thursday 8th of August
- Writing up results from yesterday's tapestation [Yara]
- Tweaking wet lab protocols, in preparation for their upload to the wiki [Yara]
- Completed warson beef interview page [Ruby]
- Emailed MPs [Ruby]
- Continued to write the script for the promotional video and learning to edit videos [Becky]
Friday 9th of August
- Emailed MP as actually in Honiton and Sidmouth constituency [Louise]
- Updating journals/wet lab timeline [Louise]
- Taking portrait photos of team members (Ben and Louise) for the wiki teams page [Yara, Nigella]
- Editing team members' portrait photos [Yara]
Ben's hunt for a recording studio to record our narration for the video was not entirely successful
Yara searching for a good spot for team photos.
Ellin's western blot of Cas13a before, during, and after the purification.
Week 9 [12/08/24 - 16/08/24]
Monday 12th of August
- Interview with NFU [Ruby, Anouska, Nigella]
- Taking portrait photos of Ruby and Anouska for the wiki teams page [Yara, Nigella]
- Editing team members' portrait photos [Yara]
- Updating notebook [Anouska]
- DNase treatment and tapestation for Cas13a crRNA/target HBEGF_a [Yara]
- Began work on 2-minute intro video [Ben]
Tuesday 13th of August
- Emailing potential interviewees [Nigella]
- Taking portrait photos of John, George, Ben and Nigella for the wiki teams page [Yara, Nigella]
- Taking photos of John in the lab [Yara]
- Lab photography of Yara [Anouska]
- DNA probe experiments [Ruby]
- Troubleshooting contamination with DNase probes [Rube, George]
- Finding methods for transporting pigs blood [George]
- Preliminary intro video work continues [Ben]
Wednesday 14th of August
- Protein purification with the AKTA machine [Louise, Ellin]
- Completing safety form [Louise]
- Finalising the transcription, tapestation and DNase write ups [Yara]
- Lab photography of George, Ruby, Ellin and Louise [Yara]
- Organised meeting with Ian Roome [Ruby]
- Meeting with Penn State iGEM team [Anouska, Nigella, Yara, Ben, Ruby]
- Promo video script writing [Nigella, Anouska, Ben]
- Emailed IDT about Dnase Alert Kit [George, Yara]
- Adapting and completing testing blood using a centrifuge/3D-Fuge to be able to show fluorescence through the blood more easily [Ruby, George]
Thursday 15th of August
- Pig blood collection from abattoir [Anouska, Ruby, Nigella]
- Even more video script writing [Ben]
- Did RNA probe re-runs with amended protocol [Ruby]
- Ran blanks for probe experiments [Ruby]
- Protein purification with the AKTA machine [Louise]
- SDS-page and western blot [Ellin]
- Testing absorbance of filtered horse blood [Anouska, Nigella, George]
- Running absorbance spectra for different dilutions of blood to find peaks to be able to measure the decrease in colour of the blood in other experiments [George]
- Further research and amendments to all blood protocols [George]
Friday 16th of August
- More promo video script writing [Nigella, Anouska, Louise, Ben]
- Emailing interviewees [Ellin, Nigella]
- Completed experiments involving heating and centrifuging blood [George]
Ben and Yara looking through the photos for the teams page!
Pig blood successfully acquired from the abattoir. Next mission: flouroscien experiments.
Getting ready to meet the Penn State iGEM team by video call!
Week 10 [19/08/24 - 23/08/24]
Monday 19th of August
- Caught up on transcription stuff, writing part registry description and Cas12/13a transformation. [Tom]
- All parts added to the registry! [Louise]
- Prep for auto induction and wet lab write up [Ellin]
- Test shots for video [Anouska, Nigella]
- Writing up proper description page for wiki [Nigella, Anouska]
Tuesday 20th of August
- Collated all RNA probe experimental data [Ruby]
- Wrote interview questions for Ian Roomes [Ruby, Becky, Anoushka]
- Research into re-design for a low-cost fluorometer [Oli]
- Produced LB for auto induction and finishing write up [Ellin]
- Prepared overnights, repeated transcription and prepared for lysis protocol for tomorrow. [Tom]
- Added features onto parts [Louise]
- Promo video filming [Nigella, Anouska, Becky, Ruby, Ben]
- Making graphics and animations for promo video [Anouska, Nigella]
Wednesday 21th of August
- Editing promo video, continuing making animations and graphics [Anouska, Nigella]
- Making logo animation for wiki home page [Nigella]
- Sending off more emails for interviews (DWT, WWF) [Nigella]
- Contacting IDT for DNA probes [Ruby]
- Made powerpoint for ian roomes interview [Ruby]
- Wiki tweaks and Notebook page updates, particularly to images [Ben]
- Writing the bronze contribution section of the wiki for RNA probe [Becky, Ruby]
- Made up autoinduction media and inoculated with overnight Cas12a cultures. [Tom]
- Prepared protocol and carried out plasmid cleave/blunting and then T7-transcription [Tom]
- Added blurb text onto some of the registry pages [Louise]
- Finalised Safety form [Louise]
- Research into novel low-cost lock-in amplification method, after a meeting with Bio-photonics expert Julian Moger [Oli]
Thursday 22th of August
- Harvesting BL21 Cas 12 and started auto-induction of BL21 Cas 13 [Ellin]
- Interview with Ian Roome [Anouska, Becky, Ruby]
- Starting on IHP wiki page [Anouska]
- Finishing Promo Video [Nigella, Anouska]
- Writing the bronze contribution page of the wiki [Ruby, Becky]
- Began hardware write-up for the wiki. Further research into lock-in amplification. Reached to engineering department for Circuit board printing, which is also of interest to another non-iGEM physics researcher [Oli]
- RNA probe experiment reruns [Ruby]
- Carried out Dnase digest on T-7 transcription products, than ran digest on Tapestation [Tom]
- Restriction digest of crRNA and target plasmids, and agarose gel. [Tom]
- Finished experiments testing reducing bloods opacity including hand centrifuge and filtering [George]
- Graphed previous results from experiments testing reducing bloods opacity [George]
Friday 23th of August
- Tecan results & data processing [Ruby]
- Cell harvesting [Ruby, Becky]
- Hardware wiki writing. Waiting on response from engineering regarding PCB. Looking into other cheap optical detection methods. [Oli]
- Made in silica gel to compare to restriction digest. And repeated Dnase digest, and tape station. [Tom]
- Continued writing for crRNA registry entries [Louise]
- Research into how to make HEX (Hexochlorofluorescein) [George]
- Beginning to write up results for wiki [George]
Nigella making the opening logo animation for the wiki home page!
Anouska making the animated explanation of Cas proteins for our promo video
Yara and Ruby taking pictures of Nigella for the team wiki page
Week 11 [26/08/24 - 30/08/24]
Monday 26th of August
- Bank Holiday!
Tuesday 27th of August
- Creating English captions for promo video [Anouska]
- Submitting promo video [Nigella]
- Making sponsor card for wiki footer and making animated graphics for description page [Nigella]
- Writing up IHP page for wiki [Anouska, Ruby]
- Brainstorming adapted area framework [Ruby, Anouska]
- 3D printing components for our hardware. Got a quote for the PCB manufacturing [Oli]
- Using Compton scattering to determine what proportion of energy needs to be removed from measurements [Oli]
- Graphed results for reducing blood opacity and Rnase probes in different environments [George]
- Wrote up the protocol for the reduction of blood opacity [George]
- Produced new batches of buffer [Ellin, Louise]
- Tapestation results troubleshooting - Dnase digestion, metal bead extraction. Also conducted qubit quantification of DNA contents of samples. [Tom]
Wednesday 28th of August
- 3D printing components. PIC research as an alternative to our circuit. Decided on Fourier analysis to workout noise [Oli]
- Re-ran Dnase and metal bead extraction and ran a Tapestation results again [Tom]
- Graphed results for the excitation and emission spectra for fluorescein and DNase Alert probe [George]
- Writing results up for concentration curves [George]
- Ran fluorescence spectra [George]
- ICCD framework writing [Ruby]
- DNA probe initial experimentation with new probes [Ruby]
- Writing up interviews into IDCC framework [Anouska]
- Adding more to registry pages [Louise]
Thursday 29th of August
- Wrote protocol for full scale T7 transcription and clean up for tomorrow [Tom]
- Started getting images for registry ready [Tom]
- Formatting Home page on Wiki [Nigella]
- PCB Gerbers sent to manufacturer for printing, other relevant information regarding hardware component substitutes researched as well to reduce the cost of production [Oli]
- ICCD framework writing [Ruby]
- Interview presentation prep [Ruby]
- Harvesting BL21 cells containing cas [Becky, Ruby]
- Writing results for probe validity [George]
- Designing and coding results page for wiki [Anouska]
- Writing up the RNAse experiment for bronze contribution [Becky]
Friday 30th of August
- Curated fluorescein concentration equivalents graphs for the results section of the Wiki. Producing diagrams for our hardware to go on the wiki [Oli]
- Michelsen-Menten enzyme kinetic analysis for our RNA Probes [Oli, George]
- Writing more ICCD framework entries for IHP [Ruby]
- Meeting with David Cotton, dairy farmer [Ruby, Becky, Anouska, Tom]
- Carried out cleaving/blunting, T7 transcription. [Tom]
- Creating a rough flowchart for the wet lab [George]
- Writing results for sample opacity [George]
- Tweaks on whole of wiki [Nigella]
- Writing up silver HP [Anouska]
Nigella creating her trademark animated infographics for the description page
Diluted blood, following heating protocol. This was tested for opacity by George, in tandem with the fluorescein detection experiment.
Oli's wonderful 3D-printed casing, which will hold all electronic and optical components, plus cow sample
September
Week 12 [2/09/24 - 6/09/24]
Monday 2nd of September
- Completed scaled up transcription protocol and conducted a Tapestation test. [Tom]
- Writing up silver HP for wiki [Anouska]
- Lysis of BL21 Cas 12 [Ellin and Louise]
- Fixing audio issues with promo video + wiki hardware and home page designing [Nigella]
- Wiki updates [Ben]
Tuesday 3rdth of September
- Attomarker interview [Anouska, Tom, George, Oli]
- Spoke with Ben Gardner with regards to lock-in amplification and other low cost methods of fluorescent detection. Researched the paper Ben gave me. Created order list for the circuits and obtained relevant existing materials to reduce strain on the budget. [Oli]
- Writing up Attomarker interview in IDCC framework [Anouska]
- Loading Cas12 onto His-Trap column for AKTA [Louise]
- AKTApure protein purification of Cas 12 [Ellin and Louise]
- Writing up the RNAse experiment for the bronze contribution [Becky]
- crRNA clean up experments and troubleshooting [Tom]
- Continuing wiki home + hardware page [Nigella, Ben]
Wednesday 4th of September
- Adding to description page, silver HP and IHP on wiki [Anouska]
- Hardware write for the description page of the wiki and the hardware page [Oli]
- Lysis of Cas13 [Louise]
- Loading Cas13 onto His-Trap column for AKTA [Louise]
- Western blot and qubit [Ellin]
- Writing up the RNAse experiment for the bronze contribution [Becky]
- crRNA clean up experments and troubleshooting [Tom]
- Added Flutter section to Hardware page, and populated it with images [Ben]
- Re-ran DNase experiments for differing enzyme concentrations [George]
- Created buffers for upcoming cas12a experiments [George and Louise]
Thursday 5th of September
- Hardware write-up for the wiki [Oli]
- Scripting/story boarding the presentation video [Becky, Talia]
- crRNA clean up, quantification and troubleshooting [George, Tom]
- 1st attempt doing a full test of Cas12a [George (mostly), Tom (helped)]
- Collecting fractions of Cas13 [Louise]
- Size exclusion AKTA purification of Cas 13a [Louise]
- Interview with Piers Pepperell [Anouska, Louise]
- Interview write up [Anouska]
- Fixed formatting problems with hardware page, and added more sections [Ben]
- Oxford meeting [Anouska, George, Tom and Becky]
- Re-ran DNase experiments for differing probe concentrations [George]
Friday 6th of September
- Western blot [Ellin, Becky]
- Began circuit assembly, waiting on a surface-mount capacitor but then it should soon be finished [Oli]
- Wrote up text for Hardware page [Oli, Ben]
- Completed maths for upcoming cas12a and cas13a [George]
Our hardware, disconnected from any power supply.
The readout of background noise from our lock-in amplifier, as measured by Oli
Western blot after purification of Cas12a on AKTA pure by Ellin
Week 13 [9/09/24 -13/09/24]
Monday 9th of September
- Presentation video planning, wiki description editing [Nigella]
- Hardware assembly, wiki writing [Oli]
- HP page write-ups [Anouska]
- Filling in team page [Anouska]
- Presentation video script writing [Becky, Talia]
- More wiki tweaks [Ben]
- Further planning of Cas13a and Cas12a experiments [George and Tom]
- Run of a cas13a fluorophore cleavage experiment [George]
Tuesday 10th of September
- Full hardware assembly as well as mock-up for Paris [Oli]
- Visited David Andrews' farm to observe first day of bTB testing + interview [Nigella, Anouska, Becky, Ben]
- Adding new info from interview to wiki [Nigella]
- Presentation video script writing [Becky, Talia, Nigella]
- Transformations of cas samples into DH5 [Becky]
- Transcription of Cas13a (crRNA and target) and Cas12a (crRNA) sequences, then cleaned up and quantified. [Tom]
- Made ampicillin plates [George]
Wednesday 11th of September
- Got the hardware to pair with the firmware and software, was able to take measurements with it. Need to optimize the process for better readings. [Oli]
- HP wiki page [anouska]
- Ran experiments to see if ribolock is needed in cas13a reactions [George]
- Planning of controls experiment for cas13a [George, Tom]
- Graphing of DNase contamination data [George]
Thursday 12th of September
- More tables for the wiki [Oli]
- Safety page for the wiki [Louise]
- Minipreps and quantification of cas12a targets [Tom]
- Initial tests Cas12a [Tom]
- Hardware wiki page [anouska]
- Taking rest of the team pictures and editing [anouska]
- Making parts page for wiki and presentation video script writing [Nigella]
- Serial dilution of blood experiment [George]
Friday 13th of September
- Cas expression meeting [Everyone]
- Team page [anouska]
- Writing presentation video script + general wiki clean up [Nigella]
George trialling components used in cas13a system
Nigella, Anouska, Ben and Becky visiting David Andrews's farm to observe bTB testing
Picture of the hardware in action. The LED flashes at a fixed frequency, exciting the sample, which can then be measured using the lock-in amplifier
