Recruitment: (November 2024 - December 2024)

November 2024 - December 2024

Hung recruitment fliers in many buildings across campus
Reached out to various biology classes along campus
Read applications, conducted interviews, and sent out acceptance/rejection letters

Spring Semester 2025

Week 1: January 27 - February 2

Conducted preliminary literature reviews about SynBio applications in water and finalized project topic
Began trainings in wetlab techniques
Researched literature data to understand water synbio software needs and relevant mathematical models

Week 2: February 3 - February 9

Developed subproject ideas and experiments
Continued trainings in wetlab techniques, understanding safety guidelines
Looked into metagenomics data and modeling literature pertaining to subprojects

Week 3: February 10 - February 16

Developed case study topics for wet-lab proof of concept, while simultaneously conducting wetlab trainings
Conducted literature reviews on detection method, phage dynamics, and reviewed structure of metagenomic data

Week 4: February 17 - February 23

Researched cyanobacterial strains from ATCC, UTEX, and Carolina, and reviewed existing bacterial and species databases.
Expanded literature review on phage–bacteria dynamics and modeling, while refining earlier searches for detection methods.

Week 5: February 24 - March 2

Continued literature reviews for current methods
Decided on creating a predictive model
Began writing code for simulations of nonlinear systems of differential equations.

Week 6: March 3 - March 9

Reviewing literature for cyanophage engineering techniques
Reviewing machine learning models pertaining to desired applications
Focused on mcy gene expression and HABs

Week 7: March 10 - March 16

Ordered M. aeruginosa strain LE3 (UTEX 3037), began planning setup for growth conditions and inoculation
Conducted literature reviews on a biosensor and B. subtilis MICI biofilms
Confirmed data availability for machine learning model training. Decided on creating database to train a model on
Conducted simulations of phage-bacteria system of differential equations

Week 8: March 17 - March 23

Refining method for sensor implementation and application in real world
Refined initial software idea, decided on lat/long as essential inputs

Week 9: March 24 - March 30

Ordered reagents for making BG-11 media for growing cyanobacteria while continuing wetlab training
Looked more in depth as to how the parameters affect the phage-bacteria dynamics
Reviewed essential python packages for software development

Week 10: March 31 - April 6

Wrote up protocols for sensor implementation and conducted microscopy training
Looked at the sra database for what it offers.

Week 11: April 7 - April 13

Received initial orders of M. aeruginosa. Inoculated subcultures and determined a preliminary setup for culture growth, additionally made BG-11 media.
Began collection of studies that perform metaT and do DE gene analysis in HABs
Looked at optimal conditions datasets as features.
Started working on phage diffusion in biofilm model

Week 12: April 14 - April 20

Looked into a toehold riboswitch literature database
Inoculated new M. aeruginosa cultures
Decided on using real-world environmental data over lab data
Designed circuits for B. subtilis for corrosion prevention

Week 13: April 21 - April 27

Reanalyzed the geometry of the biofilm-phage mode
Started M. aeruginosa subcultures, tested growth in BG-11 media from different sources (premade by Carolina vs. UTEX vs. in house)
Continued database/dataset collection

Week 14: April 28 - May 4

Continued investigation of toehold riboswitch literature database
Found four useful databases
Looked into numerical methods for simulating partial differential equations

Week 15: May 5 - May 11

NASBA primers creation, targeted regions of mcy genome, and ordered plasmids
Determined procedures for creating BG-11 plates and growing M. aeruginosa lawns
Started deciding essential features for the software

May Break 2025

Week 16: May 12 - May 18

Designed circuits and ordered primers and plasmids
Designed experiments and determined assay techniques
Verified partial differential equations numerical simulations aligned with analytical solutions
Created BG-11 agar plates
Attempted to streak and spread M. aeruginosa on BG-11 agar plates and to grow lawns of M. aeruginosa

Week 17: May 19 - May 25

Designed circuits and ordered primers and plasmids
Designed experiments and determined assay techniques
Preparing media for culturing Bacillus subtilis
Shortlisting database/website extraction points.

Summer Session 2025

Week 18: May 26 - June 1

Began design of an in silico riboswitch for target region
Decided on prioritizing true metagenomics over 16s
Engineered plasmid encoding for a functional cellulase for mycobacterial biofilm degradation
Constructed diffusion model of phage infection of a biofilm
Brought up freeze-dried bacteria received from ATCC
Miniprepped plasmids received from Addgene
Determined procedure for taking M. aeruginosa OD measurements, began testing culture growth under different culturing conditions
Collected environmental samples, started cyanophage enrichments

Week 19: June 2 - June 8

Inoculated plasmids for riboswitch
Finalized databases to extract from
Attempted and failed to transform plasmid with cellulase into mycobacteria
Extracted genomic DNA for B. subtilis 168 and B. subtilis 3610
Successfully began growing M. aeruginosa on agar plates
Validated identity of M. aeruginosa cultures via culture PCR
Began first cyanophage plating assays
Re-analyzed geometry of system for biofilm-phage infection system of PDEs to a 1D cartesian model

Week 20: June 9 - June 15

Assembly of mcy trigger plasmid
Began growing phage to a high titer
Created web scraping scripts for each database
Amplify coding regions from B. subtilis genomic DNA
Mutated out BsaI restriction cut sites in a coding region and assembled them into an empty plasmid
Began first batch of liquid cyanophage assays
Constructed 1D advection simulations that demonstrated cyanophage is limited in treatment ability
Collected lakewater samples, attempted culture PCR on samples to search for presence of M. aeruginosa

Week 21: June 16 - June 22

Collecting trigger plasmid, confirming success
Finalized extraction
Confirmed success of the coding region plasmid
Ordered A. baylyi from Addgene (ADP1-ISx - Bacterial strain 216551)
Engineered B. subtilis integration vector to make it compatible with golden gate assembly
Attempted 16S culture PCR from environmental samples
Attempted environmental DNA extraction from lakewater samples
Analyzed mathematical model for phage-biofilm effectiveness

Week 22: June 23 - June 29

Assembling switch plasmid
Cleaning extracted data for merging.
Confirmed success of integration vector.
Conducted a session of BLAST science outreach program
Identified potential cyanophage plaque on an agar assay
Collected more environmental samples, started more enrichments and assays
Adapted Acinetobacter and parameters into mathematical model for simulations
Constructed novel mathematical model of Acinetobacter-M. aeruginosa dynamics

Week 23: June 30 - July 6

Confirming success of switch plasmid assembly
Looked into how to process our extracted samples for quantitative data
Assembled the promoter, RBS, CDS, terminator into the integration vector for B. subtilis
Made B. subtilis competent cells
Sequenced the assembly for B. subtilis and found a problem for the RBS
Visualized portion of potential cyanophage plaque on SEM, while conducting more cyanophage assays
Tested DNA and RNA extraction procedures on M. aeruginosa
Conducted parameter sweeps on Acinetobacter-M. aeruginosa dynamics

Week 24: July 7 - July 13

Redo-ing switch plasmid assembly, DE gene search
Chose Kraken and Bracken as our method of taxonomic processing
Ordered RBS as DNA oligos and redo the assembly into the integration vector
Ran existing M. aeruginosa sequences through Phastest software, compared existing M. aeruginosa cyanophage sequences with BLAST
Advection-reaction PDE models on Acinetobacter-HAB dynamics

Week 25: July 14 - July 20

New reporter addition to switch plasmid
Created metagenomic pipeline
Confirmed success of the integration vector assembly
Tested TRIzol procedure for M. aeruginosa RNA extraction
Observed substantial plaques on cyanophage assay plate, attempted to propagate plaques
Reaction-diffusion equations for phage transport in biofilms expanded for analysis

Week 26: July 21 - July 27

Cotransforming mcy switch and trigger
Ran samples through metagenomic pipeline
Transformed the constructed integration into B. subtilis
Conducted a small-scale RNA extraction M. aeruginosa and A. baylyi coculture experiment
Observed conditions for proper deployment of phage in pipes
Planned lakewater microcosm experiment

Week 27: July 28 - August 3

Beginning inducing of mcy co-transformation
Merged metagenomic pipeline sample output into a single matrix
Confirmed the success of the integration vector integrated into B. subtilis genome
Made specific media for B. subtilis biofilm assays
Inoculated preliminary lakewater microcosms and began preliminary RNA sequencing experiment
Wrote up models of phage pipe transport and HAB suppression

August Break 2025

Week 28: August 4 - August 10

Grew sterile biofilms on PVC pipes
Mutated single nucleotide mutations in switches
Merged taxonomic abundance result with previously extracted metadata
Literature reviews for B. subtilis biofilm assays
Sample collection for preliminary lakewater microcosm experiment
Created a differential equation for B. subtilis deployment & detachment

Week 29: August 11 - August 17

Began construction of pipe microcosm and 3D printing prototypes of the microcosm parts
Grew nonsterile biofilms on PVC pipes for microcosm innoculation
Redid mcy switch co-transformations
Created initial aquery database
Designed microcosms for B. subtilis corrosion protection ability

Week 30: August 18 - August 24

Set up final household pipe microcosms
Began inducing second mcy co-transformation
Reviewed machine learning models and started predictor script creation
Attempted RNA extractions from preliminary lakewater microcosm samples
Compiled the python files of the mathematical modeling

Fall Semester 2025

Week 31: August 25 - August 31

Began pipe microcosm experiment with flushings and bacterial titer sampling
Did a DE gene search in order to prepare to design DE gene (gdhA and cpcB switches)
Updated predictor script to utilize three classification models
Tested growth curve and biofilm morphology for B. subtilis
Attempted more RNA extractions from preliminary lakewater microcosm experiments
Compiled information and writeups for mathematical models

Week 32: September 1 - September 7

Further prepping to design DE gene (gdhA and cpcB switches)
Continued pipe microcosm experiment with titer measurements
Flushed pipe microcosm with phage
Further analyzed model for B. subtilis biofilm detachmen
Trained model on matrix containing environmental features and species abundance
Measured the motility for engineered B. subtilis strains
Inoculated new lakewater microcosms, began new microcosm experiment

Week 33: September 8 - September 14

Finished pipe microcosm experiment
Compiling info for wiki
Created database SQL format, along with streamlit application and api
Made microcosms for B. subtilis and took SEM pictures for the steel plates treated in microcosms
Collected samples for lakewater microcosm experiment
Extracted DNA and ran 16S PCRs for microcosm experiment samples
Tested new RNA extraction procedure for microcosm samples
Incorporated fitness score from the software into mathematical model framework

Week 34: September 15 - September 21

Flushed biofilms on pipes with phage and placed under HIROX microscope
Worked on RNA seq database, began running Transcriptomic pipeline on Galaxy
Created predictive model frontend scripts
Extracted RNA for RNA seq for all 3 case studies
Conducted more DNA extractions and PCRs for lakewater microcosm experiment
Compiled simulations and graphs from the mathematical model

Week 35: September 22 - September 28

Continuing work on RNA seq database, continued running Transcriptomic pipeline on Galaxy, functional analyses w/DAVID
Looked into metatranscriptomic and what it offers. Created metagenomic and metatranscriptomic comparison scripts
Extract 16S DNA for all 3 case studies, sent 16S for sequencing
Presented mathematical models on synthetic biology in aquatic systems

Week 36: September 29 - October 5

Analyzing 16S data for all 3 case studies
Writing up wiki pages
Utilizing David & Galaxy for RNA-seq analysis
Began AI-based T and metaT analysis
Saving models publicly available repository
Compiled graphs and heatmaps and simulations of mathematical models

Week 37: October 6 - Wiki Freeze!

Analyzed RNA-Seq data for all three case studies
Completing wiki
Continued AI-based T and metaT analysis
Completing wiki and pushing software tools