Department Bionanoscience
Principal investigator Chirlmin Joo
E-mail address C.Joo@tudelft.nl
Website https://www.chirlmin.org
Sequence Dependence of DNA Looping
Supervisor: Koushik Sreenivasa, skoushik333@gmail.com
It’s been known that short double-stranded DNA can form loops. This is seen during transcription factor binding and nucleosome formation. DNA sequence for the most part plays a role in the formation of these loops. While there have been studies on bulk level on how sequence influences the bending of DNA, the rate of the looping process as a function of sequence has not been studied so far.
Using SPARXS, the prospective BEP/MEP student will use a looping assay to measure the DNA looping process of a library of sequences. They would also have the opportunity to analyse and interpret how the curvature and flexibility of the sequences influence the rates of looping and unlooping take place.
Techniques
- Single-molecule FRET
- Molecular Biology
- Python Programming
Further reading
Severins, Ivo, et al. “Single-molecule structural and kinetic studies across sequence space.” Science 385.6711 (2024): 898-904.
Single-molecule structure probing of RNA using RNA-Binding Protein
Supervisor: Archana Sivaraman, archanasivaraman.oct28@gmail.com
Cellular processes adhere to the sequence-structure-function paradigm which states that the sequence of nucleic acids determines the structure which ultimately governs its function. RNA fold into intricate structures, providing an additional layer of control over gene expression. Specifically, the structures in the 5’ untranslated region (5’UTR) of mRNA have been shown to affect translational efficiency thereby regulating cellular proteome dynamics. We employ a ssRNA binding protein to probe the structures in 5’UTRs at single-molecule precision. Through this approach, we aim to correlate the binding frequency of the protein-probe on 5’UTR sequences.
The prospective BEP student will learn to design constructs, produce synthetic RNA through in-vitro transcription, chemical labelling, perform smFRET assays using TIRF microscopy and analyse the data with Python.
Techniques
- Single-molecule FRET
- Molecular Biology
- Protein biochemistry
Single-Molecule Detection of Glycans in Full-Length Proteins
Supervisor: Moon-Hyeok Choi, moon2nick@gmail.com
Post-translational modifications (PTMs) regulate protein function and are closely connected to cellular processes and disease mechanisms. Single-molecule detection allows for the precise identification of rare PTM variations and detailed mapping of their locations, though it faces challenges due to protein complexity.
Our research group developed the FRET exchange (FRET X) technique (iScience, 2021), designed for single-molecule protein analysis focused on specific amino acids. In this project, we will apply FRET X to analyze glycan, a type of PTM in full-length protein by developing a glycan-specific labeling method using enzymes and modified sugars. The prospective MEP student will gain experience in chemical biology techniques and single-molecule experimental methods.
Techniques
- Chemical labelling
- Protein biochemistry
- Single-molecule FRET
Further reading
Filius, Mike, et al. “Full-length single-molecule protein fingerprinting.” Nature Nanotechnology (2024): 1-8.
DNA encoded chemical library
Supervisor: Koushik Sreenivasa, skoushik333@gmail.com
In medical research, a constant challenge is screening and characterising a drug compound against a certain target. Current methods offer high-throughput but do not give a full picture of the interaction of the chemical with its designated target. Recently, our lab has developed a single-molecule, high-throughput technique to measure hundreds of thousands of DNA sequences in one experiment, named SPARXS. The idea then is to attach chemicals to DNA barcodes which would later make it possible to use SPARXS for measurement.
The prospective MEP student will design and use DNA-conjugated chemical libraries to get interaction kinetics at the single-molecule level using SPARXS. They would learn chemical labelling, single-molecule FRET and data analysis for extracting useful information.
Techniques
- Conjugation techniques
- Single-molecule fluorescence
- Data analysis with Python