Student projects

We welcome students from a variety of backgrounds to participate in our research. Our group currently has the following opportunities for carrying out a bachelors or masters research project.

1. Influence of antibiotics on bacterial chromosomes.

Diseases caused by various bacteria constitute one of the biggest public health problems worldwide, causing ~15 million deaths per year [1]. Therefore improving our understanding of the antibiotic function as well as bacterial resistance mechanism is of paramount importance.

In our lab, we developed a novel method to manipulate bacterial cell shapes and by doing so we investigate the finer structure of E.coli chromosome. In the current framework we plan to study the influence of antibiotics on the chromosomes of live bacteria, by using fluorescence microscopy and quantitative data analysis (Fig.1).

We are looking for enthusiastic and hardworking students who are interested in multidisciplinary research. During this project you will learn how to work with bacterial cultures, how various types of antibiotics function and work with fluorescence microscopy to study the effects of drugs on bacterial chromosomes.

For more information pass by the office (F 0.170) or drop an e-mail : a.japaridze@[TUD]

cell

Fig1. Time-lapse images of an E.coli cell lysis. DNA inside the cell is labelled with fellow fluorescent protein.

[1] World Health Organization. 2013. Mortality and global health estimates. Geneva, Switzerland: World Health Organization

2. BEP/MEP: Genome-in-a-Box: transplanting the genome from a live cell into a synthetic cell.

The overarching goal of the Genome-in-a-Box project is to create a simple model system of the genome (yellow strands in the image) and its interaction with the cell wall (green boundary) and the cytoplasm (blue/purple dots). This is not a computer model, but a “real” experimental synthetic cell containing cytoplasm and a transplanted genome. We hope to re-create inside the synthetic cell the structure and dynamics of the genome observed in real live cells: essentially Feynman’s mantra of “what I cannot create, I do not understand”.
For this BEP/MEP project, we are looking for a creative student to help establish this model system, which would be a world-first! You will need to come up with a way to re-create the crowded conditions of the live-cell cytoplasm inside a synthetic cell. Secondly, the genome of the live-cell needs to be transplanted into the synthetic cell. Finally, we aim to let the synthetic cell expand and contract inside microfabricated structures, to see how the cell size and shape might influence the genome and the cytoplasm. These are certainly non-trivial questions: what molecules do we use to make the cytoplasm, what synthetic cell system do we use, how can we change the shape and size of a synthetic cell?
For this project, you will work independently and you will also be closely involved in the design of experiments. You will learn about microfluidics, cleanroom fabrication, synthetic cells, polymer physics, and many aspects of cell culture and DNA labeling.
If you have any ideas or questions, feel free to contact Anthony Birnie at a.t.f.birnie@[TUD] or find me in office F0.150 of building 58.

pic

3. BEP/MEP: Biomimetic Nuclear Pore Complexes

Life is a dynamic process composed of various complex and regulated sub-processes. One of the key questions is to understand the nuclear pore complexes (NPC) that regulate the selective exchange of RNA and proteins across the nuclear envelope in eukaryotic cells. To tackle this, we developed a minimalistic artificial version of the NPC (see Figure), by combining solid-state nanopores and purified nuclear pore proteins (FG-Nups). We can nicely detect single-molecule transport events of proteins through the Nup-coated pore and test its selective behaviour under different conditions. Ultimately, we aim to unravel the underlying fundamental mechanism of transport through the NPC. Besides, we take advantage of Transmission Electron Microscope (TEM) and Quartz Crystal Microbalance (QCM-D) as complementary techniques.

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We are looking for a motivated and creative Master / Bachelor student with a multidisciplinary background. Depending on the duration of your project, you will get trained on engineering nanopores with FG-Nups, performing nanopore experiments and data processing. Besides, you will get hands-on experience in using a TEM machine and detecting protein binding with a QCM-D.
If interested, please feel free contact Alessio at A.fragasso@[TUD] or drop by office (F0.170).
Further suggested Reading:
*Stefan W. Kowalczyk et al., TRENDS BIOTECHNOL.,29, 607–614 (2011).
** Stefan W. Kowalczyk et al., Nat. Nanotechnol., 6, 433–438 (2011).

4. BEP/MEP: Plasmonics assisted dielectric breakdown for nanopore production

Current nanopore fabrication techniques through TEM drilling are expensive, low-throughput and hence unsuitable for mass production. Fortunately, an alternative method for nanopore fabrication through laser assisted dielectric breakdown promises to circumvent all these problems. Nanopore fabrication through dielectric breakdown offers a cheap, clean and scalable method for production of nanopores. By designing gold antennas that resonate with our laser setup, we are able to pin and produce nanopores (5-10 nm). This method can be generalised over a large range of 2D materials (graphene, boron nitride, etc) for applications in nanopore sequencing. 2D materials exhibit interesting properties that offers new avenues for DNA sensing around a nanopore. The project will be very hands on. The only requirements are a strong interest in learning and in doing experiments. For more information, contact Wayne at w.w.w.yang@[TUD] .
Experimental goals

  1. Plasmonics nanopore breakdown protocol in ultra-thin silicon nitride.
  2. Nanopore breakdown in graphene.
  3. Characterisation of nanopores with DNA translocation experiments.
  4. Extend method to other 2D materials (Boron Nitride, MoS2, Graphene).

Learning outcomes

  1. Fabrication and handling techniques of 2D materials
  2. Experimental skills (DNA translocation experiments, laser assisted breakdown)

bowtienanoporegraphene v3

5. MEP: Atomic force microscope imaging on condensin-mediated dna condensation.

It is still a mystery exactly how the 2 meters long DNA in each of our cells is packaged into the micron-sized structures that are chromosomes. A key component of the molecular machinery that performs this feat is condensin. Condensin is a protein complex with 5 subunits, that form a ring structure, and uses ATP as its energy source. Many studies on condensin have been reported recently, but still the exact structure and the mechanism of condensation are under debate.
Our goal is to shed light on the molecular mechanism of condensin using a new, state of the art, single-molecule technique called high speed atomic force microscopy (HS AFM). HS AFM is an excellent tool for the understanding of protein’s structure and function because we can directly visualize a protein’s structure with high spatial and temporal resolution (< 1nm, < 100 ms). Moreover, this can be done in a liquid phase (in nearly physiological condition), allowing us to see the protein in action. If we can directly visualize both the conformational changes of condensin via ATP hydrolysis and DNA topological changes, we will be able to answer some fundamental questions about the molecular mechanism of condensin.
We are looking for enthusiastic and diligent master students to study the mechanism of condensing-mediated DNA compaction. The student will learn valuable skills in the operation of HS-liquid AFM as well as other biochemical experience.

For more information, please contact Je-Kuyung Ryu (J.Ryu@[TUD]) or drop by office (F0170).

Real-time HS AFM images on two of the five subunits (the SMC dimers) of condensin. The dynamical conformational changes show that the coiled-coils of SMC dimers are flexible and show extensive fluctuations in time (Eeftens et al., 2016, Cell Reports 14, 1813–1818, 2016).

6. BEP/MEP: Single-molecule investigation of RNA-polymerase interaction with supercoiled DNA

Protein-DNA interactions are typically studied using relaxed DNA in contrast to cellular world where proteins only interact with supercoiled DNA. This is mainly due to lack of experimental techniques that allow to visualize supercoiled DNA. We recently developed a novel fluorescence microscopy technique to directly visualize individual supercoiled DNA molecules (see Ganji et al, Nano Lett. 2016). This technique opens myriad of new possibilities to study interactions of proteins with supercoiled DNA. We want to use this technique to study many aspects of RNA-polymerase interaction with supercoiled DNA at single-molecule level. A bachelor/master student with background in physics and/or biology will explore this interdisciplinary project while gaining knowledge on TIRF microscopy, DNA biophysics, and biophysics of protein-DNA interactions.

For more information, please contact Mahipal Ganji (m.ganji@[TUD]) or drop by office (F0190).

DNA stretch

7. MEP: Atomic force microscopy on DNA isolated from bacteria

DNA is the information carrier and master regulator of every living organism. As recent in vivo experiments have pointed out, many aspects of the function of DNA are linked to its structure and dynamics. However, the main drawback of in vivo measurements is the difficulty to determine the influence of condensing proteins, crowding agents, supercoiling and spatial confinement on the structure and dynamics of the DNA. In a living cell all of above mentioned factors act on the DNA, making it near impossible to figure out what cause is responsible for what effect.

In order to overcome these limitations, this project aims to extract the DNA from the bacteria and study it with Atomic Force Microscopy (AFM). This high-resolution approach will enable us to separately asses the effects of condensing proteins, crowding agents and supercoiling on the DNA structure.

For this project we are looking for an enthusiastic and creative student, who is willing to learn and contribute actively to the ongoing research. You will learn various techniques such as (liquid) AFM, cell culturing and full genome extraction.

For more information, contact Anthony Birnie (015 2789299, a.t.f.birnie@tudelft.nl) or Aleksandre Japaridze (a.japaridze@tudelft.nl). For a similar project using fluorescence microscopy instead of AFM, see “MEP: Dynamics of bacterial DNA inside artificial cells”.

ecoli genome

8. Bacterial Nucleoid Project: Study of nucleoid dynamics inside live E.coli.

It is becoming clear that the spatial organisation of DNA is crucially important for its biological function. In living cells, DNA is highly confined in space with the help of condensing agents, high levels of supercoiling and numerous DNA binding proteins. Surprisingly, little is known about the impact of cell boundary on the nucleoid compaction and dynamics. By using fluorescent microscopy, we study the spatial and dynamic organisation of nucleoids in living E.coli cells. We use drugs and molecular genetics tricks to disrupt the cytoskeleton of bacteria, leading to cell sizes far larger than the rod-like wildtype cells. By controllably modifying the cell boundary we can observe a gradual expansion of the otherwise strongly compact nucleoids into donut-like structures.
We are looking for enthusiastic and hardworking students, who are interested in multidisciplinary research. During this project, you will work with various techniques from fluorescence microscopy to genetics manipulations and programming to study bacterial chromosomes.

For more information, feel free to pass by the office (F0.170) or drop an e-mail: a.japaridze@[TUD]

9. MEP project: Dynamics of chromosomal DNA inside artificial cells.

The physical properties of chromosomal DNA have been extensively studied in live cells. Due to the complexity of living organisms, it becomes difficult to find a causal relationship between a certain behaviour of the DNA and a specific parameter of the living system.
A physicist’s approach to biology could help disentangle this convoluted parameter space. Concretely, this means using microfluidics to create a minimal model of a cell: a lipid-coated water-in-oil droplet containing an isolated chromosome.
Starting from this basis, we can perturb the structural and dynamical properties of DNA inside the artificial cell. We will add DNA-condensing proteins to alter the structure of the DNA or vary the size and shape of the artificial cell to change the DNA-dynamics. This enables us to answer not only biologically relevant questions but also longstanding hypotheses from the fascinating world of theoretical polymer physics.
During this interdisciplinary project, you will work with techniques from biology (multi-colour DNA labeling, cell culture, chromosome isolation), cleanroom fabrication, microfluidics, fluorescence microscopy and some funky polymer physics.
We are looking for an enthusiastic and creative student, who is eager to work independently and enjoys to be closely involved in the design of experiments. The exact content of the project is not set in stone and can be tailored to your interests, input and/or previous experience.

For more information, please feel free to contact Antony Birnie (a.t.f.birnie@[TUD])

10. MEP project: Microfluidic engineering of synthetic cell division using biological proteins.

One of the most marvelous and organized cellular processes is division, i.e the formation of new biological cells from a paternal one. Thanks to recent improvements in biological engineering, we are now in a position to use bottom-up methodologies in order to synthetically recreate the process of cellular division: the goal is the controlled splitting of a soft lipid membrane by using proteins that naturally participate in this process.
In our lab we have recently developed a new and exciting microfluidic platform in order to produce liposomes, minimal models of the cell’s membrane (see movie 1). We are now actively applying this platform in order to engineer liposomes fission using proteins machinery (see figure 1).
We are looking for enthusiastic and ambitious students to perform a biophysical characterization of the protein machineries and the study of their activity when confined inside liposomes. During the project, the student will acquire fundamental biophysical skills that will boost forward his future career.
For more information, please feel free to contact Federico Fanalista (f.fanalista@[TUD])ff

Movie 1: production of liposomes (light gray objects) in a microfluidic device.

Movie 1: production of liposomes (light gray objects) in a microfluidic device.

11. Using Nano-Scissors to Cut Liposomes

Dividing a liposome is one of the fascinating tasks in the bottom-up synthetic-biology field. Apart from putting the relevant biological machinery inside the liposome to do the job, we are also thinking of cutting them from outside to achieve their division! Nature already has evolved a fascinating nano-scissor that exactly does this: a protein called dynamin.
We are going to combine this bio-scissor to efficiently and symmetrically divide the primitive synthetic cells, made with our recently-developed microfluidic method .We will be using a lot of microfluidic tricks (e.g. deforming the liposome using electric fields) along the way. We will learn a lot about membrane manipulation and also about the functionality of dynamin itself. You will be involved in a highly creative and fun project, and learn a lot of nice techniques (clean room, microfluidics, fluorescence microscopy etc.). If you are interested, please contact me or just drop by my office (F0.190).

For further information please contact Siddharth Deshpande (S.R.Deshpande@[TUD])
dynamin division

12. Splitting vesicles: manipulation and physical division of liposomes on chip

Recently, we have developed a robust method of making liposomes on chip that produces unilamellar, cell-sized (5−20 µm) liposomes with a high monodispersity and excellent encapsulation efficiency. Now we want to do much more with them: we want to divide them! Apart from using biological machinery to constrict and divide the liposomes, it is equally interesting to try to split them using external physical forces. In this way, we can learn a lot about the efficiency of such a process, the intermediate steps involved and a way to control it better. We wish to use well-defined microstructures to accomplish this task. An example of such a splitting geometry, resulting in a symmetric division of droplets, is shown in the video. You will be involved in a highly creative and interdisciplinary science, and get hands-on experience with clean room, microfluidics, fluorescence microscopy, high-speed imaging, etc. If you are interested, please contact me or just drop by my office (F078).

Droplets splitting in a microfluidic chip

droplet-splitting

Siddharth Deshpande (S.R.Deshpande@[TUD])

13. Pushing the Limits of DNA Imaging Using TEM

There has been always a great need and scientific curiosity to see how life works at atomic scale. Of all developed microscopy techniques, the only type of microscope capable of sub-Angstrom spatial resolution is Transmission Electron Microscope (TEM). However, working with biological molecules has always been a challenge for TEM imaging due to lack of crystallinity as well as low atomic weight of their constituent atoms i.e. Carbon and Nitrogen. In this project, we aim to study the structure, dynamics and interaction of different DNA structures using high-tech TEMs. Sophisticated nanostructures are being developed in our lab to withstand ultra high vacuum condition inside TEM on one hand and to produce lowest background contrast for TEM imaging on the other hand. Moreover, we are developing new TEM imaging techniques that allow us to increase the detection limit.

We are looking for highly motivated and enthusiastic Master or Bachelor student in the field of molecular biology, life science, biochemistry or relevant fields to take part in our exciting and interdisciplinary research project. Your task will be incorporating different electron scattering agents into nucleic acid structures such as dsDNA or DNA origami. You will be working with world experts in the field of molecular biophysics and transmission electron microscopy. Furthermore, you will be learning great deal of techniques such as nanofabrication, Graphene transfer and TEM imaging. Should you have any further questions please do not hesitate to contact me. I can show you atoms!

Yoones Kabiri: y.kabiri@[TUD]

(a) Titan microscope, world’s most advanced microscope with sub-Angstrom resolution (b) AFM image of dsDNA (c) AFM image of our special built-in-house DNA origami

(a) Titan microscope, world’s most advanced microscope with sub-Angstrom resolution (b) AFM image of dsDNA (c) AFM image of our special built-in-house DNA origami

14. Graphene nanopores for DNA sensing

Graphene – a single layer of carbon atoms – is a spectacular material with numerous applications that can be of great significance to mankind. One of those promising applications is a graphene-based biosensor that can directly sense single molecules or sequence DNA. We combine creativity and state-of-the-art nanotechnology to develop different graphene nanopore devices. Two major advantages of graphene are its single-atom thickness that coincides with the size of a single DNA nucleotide, and its electrical conductivity. We exploit these characteristics to maximize the sensing resolution.

Studentproject_StephanieHeerema

In this exciting project different fields meet: physics, chemistry, biology, nanotechnology and electronics. Depending on the duration of the research, student projects can involve one or more of the following: DNA translocation measurements through graphene nanopores, graphene and boron nitride handling, e-beam lithography, AFM, Raman spectroscopy, data analysis and modeling. We are looking for motivated and curious students that would like to contribute to tomorrows next biosensing technology. Further reading: G.F. Schneider, Q. Xu, S. Hage, S. Luik, J.N.H. Spoor, S. Malladi, H. Zandbergen C. Dekker. Tailoring the hydrophobicity of graphene nanopores. Nature Communications 4, (2013) G. Schneider and C. Dekker. DNA sequencing with nanopores. Nature Biotechn. 30, 326(2012) G.F. Schneider, S.W. Kowalczyk, V. E. Calado, G. Pandraud, H. Zandbergen, L.M.K. Vandersypen, C. Dekker. DNA translocation through graphene nanopores Nano Lett., 10, (2010) For more information contact: Stephanie Heerema s.j.heerema@tudelft.nl Office (F066) Begeleider: Stephanie Heerema

14. Imaging DNA and proteins at high resolution and high speed with the Atomic Force Microscope

‘Seeing is believing’ is a saying that applies as much in bionanoscience as it does in the day-to-day world. This is why we use cutting edge microscopy techniques to visualize single proteins and their interaction with DNA. The Atomic Force Microscope (AFM for short) offers by far the highest resolution of the techniques that can be used under near-physiological conditions. Its ability to see individual molecules in their entirety without any labelling makes it a powerful tool for answering biophysical and biological questions. Until recently, the AFM was a very slow technique, and only static or very slowly changing conformations could be imaged. However, the Cees Dekker lab is one of the few in the world to have acquired a new type of AFM, that can make nano-videos up to 20 frames/s. You can have the chance to work with this unique instrument and see molecules yourself, while contributing to one of our current research projects: • Visualizing how NAP1 builds a nucleosome, the packing unit of DNA in our cells • Using DNA origami to create a universal imaging platform for DNA-protein interactions in AFM • Imaging the shape and conformation of cohesin, the molecule that holds our chromosomes together.

Figure 1. A histone tetramer is attached to a DNA molecule and hops between two stable positions 3.6 nm apart. AFM image series taken at 1 frame/s, in physiological buffe.r solution

Figure 2. A DNA origami nanoplate, imaged in liquid. The individual strands and crossovers are clearly visible. Interested? Want to know more? Contact me anytime by email or drop by my office. Begeleider: Allard Katan

15. Plasmonic nanopores

Biomolecules, the little molecular machines that make up life’s complexity, are at the center of disease and treatment. Understanding their properties on the single molecule level at high-throughput is essential for drug development.

Towards this end, we are developing a universal single-molecule biosensor based on a plasmonic gold nanoantenna; a nanoaperture in a freely suspended gold film. When light is impinged on this antenna, electrons in the antenna help focusing the light down to size of the metal nanostructure. This tight electromagnetic field concentration allows us to optically trap biomolecules and provides us read-out from the light scattered by the nanoantenna, providing us information about the physical properties of the biomolecules. Moreover, the thin film allows us to actively attract molecules towards the sensor and act electrokinetic forces on them to calibrate our nanoplasmonic trap.

We encourage highly motivated bachelor and master students to join us promptly

for a bachelor (8-12 weeks) or master (6-12 months) project. This challenging project will have a largely experimental character and a decent background i­­­­­n optics, electronics and nanofluidics is welcomed. You will be involved in biomolecular trapping experiments and data analysis (MATLAB), and can be, depending on your interests, involved in device fabrication, modeling, data analysis (MATLAB), simulations (FDTD), building device electronics and control software.

Please contact Daniel Verschueren (d.v.verschueren@[TUD])

Begeleider:Daniel Verschueren