In the latest episode of Genetics Unzipped, presenter Kat Arney is squelching through the Californian mud, swimming with platypuses, bearing witness to daylight robbery and even finding time to catch an episode of Star Trek as she looks back on some of the most mind-blowing stories from the world of genetics in 2021.
We meet the Borgs – huge genetic elements in archaea that can assimilate genes from their neighbours – and discover how whitefly pulled off a genetic theft that enabled them to become one of the world’s most destructive agricultural pests.
We hear how researchers are developing mirror-image DNA polymerases that can make mirror-image DNA – perfect for long-term, stable data storage. Then there’s the strange discovery that hundreds of viruses use a DNA base called 2-aminoadenine, known as Z, instead of the usual adenine (A), with big implications for our understanding of the genetic code as we know it.
And finally, we take a dive into the duck-billed platypus genome, to discover what these mysterious monotremes can teach us about mammalian evolution.
If you enjoy the show, please do rate and review on Apple podcasts and help to spread the word on social media. And you can always send feedback and suggestions for future episodes and guests to podcast@geneticsunzipped.com Follow us on Twitter – @geneticsunzip
The 8th Edition of the Annual Portuguese Drosophila Meeting (#DrosTuga2021), aims at bringing together national and international members of the Portuguese Drosophila community. Along with them, Portuguese Drosophila scientists abroad and any participant from other country interested in Drosophila and developmental research are invited. https://igc.idloom.events/drostuga2021
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The purpose of this Drosophilameeting is to promote open sharing of data and ideas, as well as to provide arich forum for discussion of new research findings and conceptual breakthroughs in an informal environment.
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Due to the ongoing uncertainty caused by the COVID-19 pandemic, this edition will be held entirely ONLINEon the afternoons of the 29th and 30th of November 2021.
The event will include selected short and long talks presented at the Plenary Sessions via Zoom. Thanks to our sponsors, the best talks and posters will receive a prize. In addition to the presentations, there will be time for discussion and mixing between researchers at all career stages during the two interactive Poster Sessions (held in the Hopin platform). You can take a look at the programme for more details.
In this DrosTuga 2021 edition, we will have the pleasure of listening to two great Keynote Speakers. Their exciting work spans a broad range of topics of interest to our community: Isabel Palacios and Nicolas Gompel. Besides our speakers research, we will have the opportunity of talking about outreach and the importance of Drosophila studies in science.
Attendance is FREE, but registration is compulsory.
NEW Abstract submission deadline for posters only: 7th November 2021 (23:59h GMT+1). Click here to submit your abstract. The deadline for short and long talks is now closed (24th October 2021).
Registration deadline: 24th November 2021 (23:59h GMT+1). Click here for registration.
Below you’ll find each of the talks, plus a Q&A chaired by Development Editor-in-Chief James Briscoe. The next #DevPres webinar will be held on 10 November 2021, and chaired by James Wells – subscribe to our mailing list for updates.
Daisy Vinter (University of Manchester) – Dynamics of hunchback translation in real time and at single mRNA resolution in the Drosophila embryo
Ping Wu (University of Southern California) – Cyclic growth of dermal papilla and regeneration of follicular mesenchymal components during feather cycling
Yan Gong & Dominique Bergmann (Stanford University) – The Arabidopsis stomatal polarity protein BASL mediates distinct processes before and after cell division to coordinate cell size and fate asymmetries
In April 2020, I should have attended The Company of Biologists Workshop “The Cytoskeletal Road to Neuronal Function”. If only there was not the beginning of the SARS-CoV-2 pandemic. The Workshop was ultimately canceled, however the organizers suggested to us (the enthusiastic and disappointed participants) to initiate a Webinar as a virtual platform for the neuronal cytoskeleton research community. I and four other participants (Satish Bodakuntla, Meng-meng Fu, Oliver Glomb and Lisa Landskron) volunteered as organizers, and this is how “The Cytoskeleton of Neurons and Glia” Webinar was established in April 2021. Since then, we hosted more than twenty diverse and inspiring speakers. On October 7th our efforts reached a milestone – the twelfth Webinar marked six months of its existence!
Actin monomers as cellular cobblestones
I wrote this Node inspired by one of our speakers – Dr. Eric Vitriol. He presented a work from his Lab focusing on how actin dynamics are affected by actin monomers in neuronal cells. But first, let us take one step back and borrow the cytoskeletal road metaphor from The Company of Biologists workshop. Imagine a trail made of cobblestones, assembling underneath your feet and in front of you while you are walking. This trail will fall apart at the back once it is no longer needed. Next, imagine the cobblestones are self-aware of their numbers, and they will never start making the trail unless they “know” there is a sufficient number of them to start the construction (“critical concentration”). Additional factors like a solid ground to build on, stabilizing mortar, continuous supply of high quality cobblestones, all facilitate the trail assembly. While a self-aware and self-building trail is still far from our reality, practically all cells in our bodies have plenty of “cobblestones” called actin monomers. The monomers are present way above the critical concentration and can assemble into trails and many different cellular structures. As a matter of fact, with such a good supply of actin monomers why not building all the time?!? Well, cells have figured a way to protect themselves from an energy-costly unproductive actin assembly. This is achieved by sequestering actin monomers via another molecule called Profilin (PFN), that will release the “cobblestones” once there is a “construction permit”.
Profilin, actin monomers and neurons
In cells with neuronal origins, the team of Dr. Vitriol studied how different actin structures assemble from a common monomer pool and how Profilin 1 (PFN1) is influencing this process. They varied the concentration of PFN1 and induced cellular shortage, normal levels, or excess of PFN1. In those three conditions, they analyzed two established actin assemblies (branched Arp2/3-mediated and linear Mena/VASP based) at a highly dynamic part of the cell called leading edge. The cells responded with downsizing actin assembly throughout the cell when there is a shortage of PFN1. In addition, the lack of PFN1 repositioned the Arp2/3 nucleator complex towards the center of the circular cells, and reduced Mena/VASP function, ultimately disrupting the architecture of the leading edge. At low concentration of PFN1 cells seem to employ a mechanism to be resourceful and favor linear networks constructions. Abundance of PFN1 signals that both linear and dendritic actin networks can be reestablish. You can find more details in the paper published in 2020 in Current Biology, with Dr. Kirsten Skruber as a first author (1). This study provides us with a glimpse on how mammalian cells reshape actin assemblies when they face challenging situations when the concentrations of a major “guardian” and nucleators of the actin building blocks are changed. These disturbances must come at high costs for cell fitness, especially in long-lived, specialized cells like neurons. And while short term each cell has certain capacity to cope with different intra- and extracellular challenges, long-term exposure to the same “stretch” will eventually lead to neuronal dysfunction.
Mutations in PFN1 are direct cause of a late onset, incurable neurological disorder called amyotrophic lateral sclerosis (very recent review from another speaker in the Webinar series Dr. Kai Murk (2)). In addition, decreased levels of PFN2 were detected in cells from patients with Charcot Marie Tooth disease – genetically heterogeneous disorder affecting the peripheral nerves, as found by the team led by one of my PhD supervisors – Dr. Vincent Timmerman (3). Thus, the building blocks of the actin cytoskeleton are getting the closer attention they deserve, as there are plenty missing pieces in the puzzle that costs humans their health.
Skruber et al., 2020, Current Biology 30, 2651-2664;
Murk et al., 2021, Front. Cell Dev. Biol. 9, 681122;
Juneja et al., 2018, J Neurol Neurosurg Psychiatry 89, 870-878.
Our twelfth SciArt profile of the series features Giacomo Moggioli, a PhD student at Queen Mary University of London studying genomics of deep sea worms
Where are you originally from and what do you work on now?
I am from Milan, Italy. I did both my bachelor’s and master’s degrees at University of Milan-Bicocca. During my years as undergraduate student I started to feel fascinated by deep sea environments, so I decided to apply for the Erasmus project and spend one year working on my master’s thesis at Heinrich-Heine University in Düsseldorf. My thesis was focused on the role of iron-sulphur clusters in the origin of life, which, accordingly to the most robust hypothesis, happened in hydrothermal vents environments. My curiosity for this kind of deep-sea environments motivated me to apply for a PhD at Queen Mary University of London. Now, as a PhD student, I work on a fascinating clade of Annelid worms, Siboglinidae, which thrive in hydrothermal vents. I love to be able to learn more and more about these creatures. They possess such unique features such as the lack of a digestive system in favour of a symbiotic lifestyle that allows them to harness the energy contained in sulphur compounds in order to keep their metabolism going.
Darwin of Life I made this for an Art and Science contest organized by the university of Milan-Bicocca and it has been exhibited at the Natural History Museum of Milan. I have tried to shape the tree of life as the portrait of the famous scientist. The origin of life is at the tip of the beard and from there you can follow the evolution organism by organism upward. Plants, animals, fungi, invertebrates, I have simply tried to put as many different organisms as possible. Can you spot the Koala?
Were you always going to be a scientist?
Not at all. I believe that what happens in our lives keep on shaping our desires, aspirations and interests, so I don’t feel I ever had a clear path ahead of me. Nevertheless, when I was a little kid I was fascinated about being a marine biologist, and here I am today! Over the years I had many different part-time jobs, for example I was working in a flower shop for a couple of years during University, I worked at a stand during design week promoting some very nice lamps and I have also worked in a motorbike customisation shop, where I was mainly painting on helmets and bikes. So I definitely had the opportunity to try different paths, but overall the part time job I loved the most during my time in Milan was being a freelance illustrator. As an illustrator I was taking part in art fairs, art markets and exhibitions all over Italy showing my art and selling my prints. I am happy where I am now, working with deep sea animals in London, but I definitely considered making a living out of my illustrations.
Neko-no Kami, the spirit of cats. This is part of a project on Japanese spirits, my idea is to make a sort of an encyclopedia about the many different “Kami” or spirits that are coming from the Shintoism. This ancient animistic religion believes every natural and even artificial entity has its own soul, its own spirit. I have expanded this concept and started to imagine how these spirits might look.
And what about art – have you always enjoyed it?
Yes, as far as I can remember. When I was a kid my mother was buying me awesome illustrated books about sea animals. I was spending hours looking at those beautiful drawings of all the amazing life forms we can find in our oceans! I always tell my friends that those books made me who I am today, having marine biology and illustration as the main passions in my life. I am very grateful to have had the opportunity to study art history at high school. In those classes I learnt a lot about artists and understand how they made their art and what they wanted to say with what they were doing. I really feel there is a world behind every single art piece we see; a world made by the experiences of the artists and their opinions, ideas and way of looking at the world. This feeling really enhanced my curiosity about Art. Now I really enjoy going to art museums and exhibitions and when I am walking around the city, I always love to search for street art, murals and graffiti, trying to imagine the world that might be behind them.
“I really feel there is a world behind every single art piece we see; a world made by the experiences of the artists and their opinions, ideas and way of looking at the world.”
Henshin-no Kami, the spirit of metamorphosis Another artwork from my Kami series.
What or who are your most important artistic influences?
There were many amazing artists that impacted me and there will always be new ones as well. To name just a few of them: Salvador Dalí, who showed me that the only limit in art is our imagination, an Italian street artist named “Blu”, his rich and meaningful works taught me to always keep an eye open for hidden beauty that always surround us, Masashi Kishimoto, the author of the manga “Naruto” from which I understood the artistic potentiality of a black outline, and finally another Italian street artist “Hitnes”, from which I have learnt the artistic potentiality of not using a black outline.
Disassembled tit bird. I am really fascinated by birds and I love to play with their shapes and their colours and see the final result!
How do you make your art?
When I was living in Milan I had a studio and this allowed me to try many different techniques. After these experiments, I managed to find a technique which I felt comfortable with. I was first drawing with pencil on paper, then inking the lines with a black marker, erase the pencil away and finally color the drawing with alcohol-based markers which allowed a very good control of the shades. Then I move to London, and I couldn’t carry my studio with me. Therefore, I decided to switch to digital techniques and now I am mainly working using my tablet and a drawing app. After a steep learning curve, I now feel comfortable drawing with my tablet and I love to be able to draw from my sofa instead of sitting on a desk fully covered in ink and all sort of different drawing tools.
Primate’s face Primates all together are forming a human face. When making this composition I always like to depict different interactions between the characters. Can you spot the monkey trying to steal the baboon’s banana?
Does your art influence your science at all, or are they separate worlds?
I would say that science is influencing my art more than the other way around. I love to make science-inspired drawing, but I have never really done art-inspired science before. Nonetheless, I think that art may indirectly influence my science. While doing my art I have learnt how to transform big, complicated and very detailed subjects into some way simpler yet still very descriptive drawings. I find this “simplification process” I use in my art to be very useful in science as well. As a scientist a big part of my work is handling huge amounts of genomic data and identity the key features in order to be able to simplify the information and make a good description of the organism I am studying.
Common frog This is part of my icon-like animal series, heavily inspired by the British illustrator Owen Davey. I have started to make these icons and share them with my colleagues so that they could include them in their slides and hopefully have a nice simple representation of the organisms there are studying.
What are you thinking of working on next?
At the moment, my priority is completing my PhD and publish my first paper as first author on deep sea worm genomics. After that I would like to keep on working with marine organism genomics in a Postdoc. As a side artistic project, I have started to work on a card game during the long lockdown evenings and I would like to finish the last details and try to release it to the public. Little spoiler: there will not be deep sea creatures but there will be some dinosaurs in my game .
T-rex toy This vintage-like dinosaur toy will be part of my upcoming card game. Stay tuned!
If you want to have a look at my other works and be updated on my new projects you can follow me on Instagram: https://www.instagram.com/kelp_art/
We’re looking for new people to feature in this series – whatever kind of art you do, from sculpture to embroidery to music to drawing, if you want to share it with the community just email thenode@biologists.com (nominations are also welcome!)
We needed an afternoon snack in the Morris Lab today, so I conducted a blind taste test. With a sample size of 7 (everyone loitering around lab this afternoon), I investigated whether people preferred @AldiUSA energy bars or Clif bars. Here are my results: pic.twitter.com/jZkYXJBSaJ
Instead of using the word "interestingly" every five sentences, what if we start incorporating exclamation points in the scientific literature? Why is this taboo?
Our Production Editors at Development have responded that they are happy to accept exclamation marks in the text, but only if they follow the instructions below: Authors must indicate their level of excitement/surprise using a scheme similar to the use of asterisks for significance level: !, interesting finding; !!, surprised by this result; !!!, gobsmacked/couldn’t believe it; !!!!, that can’t be right, we should repeat that experiment
Thanks to the #DevBio community for sharing their thoughts, especially on twitter. If you have some news that you think we should share on our blog, please get in touch at thenode@biologists.com. If you are interested in getting involved with writing preLights you can find out more here.
After a year of lockdowns and virtual classes at Bangor University, the opportunity to do a real lab project this summer at the Francis Crick Institute was definitely not one to miss. Under the patient supervision of Adrien Franchet and Sebastian Sorge in Alex Gould’s lab, I set out to explore the roles of some amino acid transporters during the development of the genetic model organism Drosophila.
This two month summer project was my first opportunity to gain hands-on experience doing hypothesis-driven science and to interact with many talented researchers at the Crick. As an undergraduate, my only previous exposure to fruit flies was from reading published papers but, right from day one, I got stuck in to the nitty gritty of Drosophila developmental biology and larval dissections.
The Gould lab are interested in figuring out how the neural stem cells of the developing CNS are so highly protected against environmental stresses such as nutrient restriction (NR) and hypoxia. This process is a key part of brain sparing, which involves sustaining the growth of the CNS at the expense of other organs such as adipose tissue. In mammals, brain sparing is commonly observed in neonates following intrauterine growth restriction. However, the key signalling and metabolic pathways underlying brain sparing are still unclear.
Amino-acids are key signals for growth and they are also critical for protein synthesis. The uptake of amino acids by tissues involves a large number of different amino-acid transporters and I set out to decipher whether two of these transporters (AAT1 and AAT2) are required in the neural stem cell niche (glia in Drosophila) or in adipose tissue (fat body in Drosophila) for CNS and body growth. My project stemmed from Adrien’s and Sebastian’s recent RNA interference (RNAi) screen of amino acid transporter candidates. I followed up two of their screen hits (AAT1 and AAT2) using UAS-RNAi knockdowns lines crossed with Gal4-driver lines specific for glia (repo-Gal4) or fat body (Cg-Gal4). The goal was to measure the phenotypic effects of these cell-type genetic manipulations during standard fed development and also during severe NR on an agar-only diet. Phenotypes were measured for larval and pupal weights using an accurate microbalance. I also quantified CNS phenotypes from confocal microscopy images by measuring CNS area and also neural stem cell (neuroblast) proliferation via the incorporation of a labelled nucleotide analogue (EdU).
I found that RNAi knockdowns of either AAT1 or AAT2 produced more severe phenotypes in glia compared to fat body (Figure 1). Hence, larval pupal and adult weights were largely normal with the fat body knockdowns (Figure 1A, 1C). However, both glial knockdowns gave modest changes in body weight at the larval stage but, by the pupal stage, these only remained significant for AAT2 (Figure 1B, 1D). I also noticed that glial knockdown of AAT2 eventually resulted in adult lethality, shortly after eclosion, with flies displaying very severe locomotor defects.
Figure 1: Larval and pupal weights of AAT1 or AAT2 knockdowns.Larvae were raised on standard lab diet (Fed) or nutrient restriction (NR). (A,B) Larval weights of fat body (Cg-Gal4) knockdowns (A) or glia (repo-Gal4) knockdowns. (C,D) Pupal weight of fat body (C) or glial (D) knockdowns. mCherryRNAi is a control of RNAi line and times refer to hours after larval hatching (ALH). Statistical significance (asterisks) was determined using Tukey’s Multiple Comparison Test.
In a parallel set of experiments, I investigated the effects of the AAT2 knockdowns on the growth of the developing CNS and on the proliferation of neural stem cells. To do so, I dissected brains from fed larvae and from larvae exposed to one day of NR. I then performed an in vitro EdU incorporation assay as an indicator for neuroblast progression through S-phase of the cell cycle. I found that the fat body manipulations had no significant effect on CNS size or on neuroblast proliferation. In contrast, the glial manipulations revealed that AAT2 is required in glia for proper growth of the larval brain, as the CNSs of repo-GAL4; AAT2RNAi larvae were strongly reduced in size and likewise the EdU incorporation was much lower than genetic controls (Figure 2A, 2B). This glial requirement for AAT2 for neuroblast proliferation occurred in both fed and NR larvae (Figure 2C, 2D). Thus, in conclusion, my project has revealed a constitutive function in glia for the amino acid transporter AAT2 during both normal CNS growth and brain sparing. It will be important in future to explore whether AAT2 is required in the surface glia of the blood-brain barrier or in the internal cortex glia that surround neuroblasts and their daughter cells. Equally importantly, it will be interesting to identify which specific amino acids are transported by AAT2.
Figure 2: Glial AAT2 is required for neuroblast proliferation in the larval CNS Larvae were raised on standard lab diet (Fed) or nutrient restriction (NR). (A,B) Confocal images of the nuclear marker DAPI (cyan) and the proliferation marker EdU (yellow) for third-instar larvae expressing repo-Gal4 driving AAT2 RNAi and their genotype conrols (mCherryRNAi) (C,D) Quantitation of CNS area (C) and average EdU intensity (D) for control and AAT2 RNAi lines. **** Statistical significance determined suing Tukey’s Multiple Comparison Test.
Overall, this fascinating project has given me a first taste of biological research at the bench and has also allowed me to develop critical thinking and data processing skills. I am indebted to Adrien Franchet and Sebastian Sorge for their fantastic direction, and to Alex Gould and all of his lab for their encouragement throughout. I would also like to thank the Francis Crick Institute for hosting me and the Medical Research Foundation Rosa Beddington Fund for supporting my project and allowing me to contribute to this captivating field of research.
This summer, I was given the opportunity to conduct research at the Francis Crick Institute in the Znamenskiy lab. The aim of the Znamenskiy lab is to understand the relationship between connectivity, gene expression and function of cortical neurons.
The neocortex is a region of the brain integral in performing higher cognitive functions. Neocortical projections can be divided into three broad classes. Corticothalamic (CT) neurons are located mostly within layer 6 and send axons to the thalamus. Pyramidal Tract (PT) neurons are nearly exclusively positioned within layer 5 and project to brainstem and spinal cord. Intratelencephalic (IT) neurons are distributed throughout all six layers and project to distant cortical areas (Kast & Levitt, 2019). The expression of transcription factors during development can affect projection patterns. For example, when Fefz2 is deleted, the cortex no longer sends projections to the brain stem and instead sends projections to the thalamus or contralateral hemisphere (Kast & Levitt, 2019). This shows that genes expressed by a neuron during development play an important role in determining its wiring patterns.
Beyond these broad projection classes, the genetic basis underlying patterns of neocortical connectivity is little understood. The primary visual cortex (V1) is a region of the brain that is important for receiving, segmenting, integrating, and processing visual information relayed from the retinas. Subsequently, the processed information is then sent to other regions of the brain. This is a highly specialised process that allows the brain to recognise patterns quickly and with the absence of a conscious effort. The V1 provides a platform for understanding the neocortex due to its serially homologous structure, and therefore can be used as a model for neocortical projections. The V1 sends projections to several higher visual areas as well as many other areas of the brain such as the lateral geniculate and lateral posterior thalamic nuclei, superior colliculus, striatum, and other subcortical structures but little is known about how these connectivity patterns are established (Kast & Levitt, 2019).
To understand which genes are important for specifying long range connectivity patterns from V1, in vivo genetic manipulations using CRISPR/Cas9 can be used to determine what happens to connectivity patterns when the expression of target genes is altered. CRISPR/Cas9 is a simple, rapid method to modify gene expression which can be pooled together to look at many genes in parallel. As well as knocking-out the gene of interest using the prototypical CRISPR/Cas9 gene editing approach (Figure 1A), methods for modulating gene expression using catalytically inactive Cas9 fused to transcriptional modulators have recently been developed (Figure 1B-C). CRISPR activation (CRISPRa) allows functional analysis of redundant genes through overexpression, whereas CRISPR interference (CRISPRi) allows analysis of gene function by knocking-down gene expression at the transcriptional level and is thought to have fewer off-target effects (Gebre et al., 2018). The aim of my project was to perform preliminary experiments validating whether gRNA constructs designed to be used to examine changes in in vivo V1 connectivity patterns, using CRISPR knockout, CRISPRi or CRISPRa, altered gene expression in vitro. The first part of my project was to clone some of the gRNA CRISPR constructs, and the second part was to test constructs in vitro.
FIGURE 1. Mechanism of CRISPR/Cas9 Genetic Modulation. (A) CRISPR knockout involves co-expressing Cas9 and a gRNA in a cell. The Cas9 protein recognises a specific sequence called the scaffold sequence in the gRNA while another sequence within the gRNA called the spacer region determines the target site within the genome to be modified. The Cas9 protein generates double strand breaks in the gene of interest that are repaired through the non-homologous end joining (NHEJ) pathway that is prone to producing indel mutations (red bases here indicate an insertion) that can render genes non-functional when translated. (B) CRISPR activation (CRISPRa) constructs work via transcriptional activators fused to catalytically dead Cas9 (dCas9) which are targeted near transcriptional start sites of the endogenous gene of interest by the gRNA to induce their overexpression. (C) CRISPR interference (CRISPRi) constructs consist of dCas9 fused to transcriptional repressor domains that are recruited in proximity of the transcription start site of an endogenous gene to repress transcription.
gRNA constructs were tested along with corresponding Cas9s (SP-Cas9, dCas9-KRAB-MeCP2, and dCas9-VPR for CRISPR knockout, CRISPRi, and CRISPRa, respectively) to determine whether a change of expression in our genes of interest occurred within Neuro-2A (N2a) cells. Target genes for validation (Frizzled 1 (FZD1), Androgen Receptor (AR), Polycystic Kidney and Hepatic Disease 1 (PKHD1), and Anaplastic Lymphoma Kinase (ALK)) were identified due their established endogenous gene expression in N2a cells. To determine whether the gRNA constructs worked I co-transfected Cas9’s with the gRNA construct into N2a cells and observed whether this altered expression of target genes by looking at endogenous protein levels through immunostaining. Endogenous protein levels in each condition were compared to a control plasmid without a gRNA insert. The results obtained from the quantification of the transfection and subsequent immunostaining are shown in Figure 2A-C. These results did not reveal expected differences in gene expression between gRNA constructs and further experiments need to be performed using alternative antibodies or staining conditions. However, the project has given me an insight into the molecular basis of developmental biology, and I thoroughly enjoyed learning the techniques and protocols required to complete the cloning process. During my research internship I was able to obtain applied, practical experience within the laboratory which due to the COVID-19 pandemic, has been limited during my undergraduate degree. I also was given a level of independence which I did not expect within the laboratory, completing the transfection of gRNA constructs was an engaging, albeit challenging process as my cells became contaminated during the passaging process. However, I was able to overcome this setback and build resilience. Overall, I really enjoyed my project, and it has encouraged me to pursue a career in scientific research.
FIGURE 2. Quantitative Immunofluorescence after transfection of Cas9 plasmids and mCherry expressing gRNA plasmids targeting either ALK or FZD1. (A) ALK antibody fluorescence in +/- mCherry Cells. (B) FZD1 antibody fluorescence in + mCherry; +/- Cas9 Cells. KO/i/a – KO = Knockout; i= interference, a = activation. Each symbol shows the mean normalized grey value of N2A cells which reflects the level of fluorescence from antibodies targeting the endogenous protein-of-interest after immunostaining. The negative control used was a plasmid without a gRNA insert. SP-Cas9 was used for ALK/FZD1 KO & negative control, whereas dCas9-KRAB-MeCP2 was used for ALK/FZD1 CRISPRi, dCas9-VPR used for ALK/FZD1 CRISPRa. For the FZD1 transfection there was an unexpectedly low number of +Cas9 cells. (C) Immunohistochemistry staining against mCherry and ALK, as well as DAPI staining in N2a cells transfected with SP-Cas9a and a gRNA targeting ALK. mCherry is expressed by the gRNA constructs, staining this protein shows which cells were transfected with our construct of interest, whereas DAPI staining marks the nuclei of all cells. The overlap in ALK and mCherry signals suggests further optimisation of immunostaining and imaging conditions is required to avoid bleed-through.
I would like to take this opportunity to thank the Francis Crick Institute, particularly the Znamenskiy lab for allowing me to undertake research at their facility, alongside my supervisor Benita Turner-Bridger for supporting me in my project. Furthermore, I would like to show my appreciation to the Medical Research Foundation and the Rosa Beddington fund which has provided the financial support for my project. It is an honour to have the opportunity to contribute to The Node and the British Society of Developmental Biology, and I would strongly encourage other undergraduate students to pursue a similar research project during their studies. This experience has been unlike any other.
References:
Gebre, M., Nomburg, J.L and Gewurz, B.E (2018). CRISPR-Cas9 Genetic Analysis of Virus-Host Interactions. Viruses, 10(2), 55.
Kast, R.J and Levitt, P (2019). Precision in the development of neocortical architecture: From progenitors to cortical networks. Progress in Neurobiology, 175, 77-95.
Unique functions for Notch4 in murine embryonic lymphangiogenesis Ajit Muley, Minji Kim Uh, Glicella Salazar-De Simone, Bhairavi Swaminathan, Jennifer M James, Aino Murtomaki, Joseph D McCarron, Chris Kitajewski, Maria Gnarra, Gloria Riitano, Yoh-suke Mukouyama, Jan Kitajewski, Carrie J Shawber
Androglobin, a chimeric mammalian globin, is required for male fertility Anna Keppner, Miguel Correia, Sara Santambrogio, Teng Wei Koay, Darko Maric, Carina Osterhof, Denise V Winter, Angèle Clerc, Michael Stumpe, Frédéric Chalmel, Sylvia Dewilde, Alex Odermatt, Dieter Kressler, Thomas Hankeln, Roland H. Wenger, David Hoogewijs
Retrospective analysis of enhancer activity and transcriptome history Ruben Boers, Joachim Boers, Beatrice Tan, Evelyne Wassenaar, Erlantz Gonzalez Sanchez, Esther Sleddens, Yasha Tenhagen, Marieke E. van Leeuwen, Eskeatnaf Mulugeta, Joop Laven, Menno Creyghton, Willy Baarends, Wilfred F. J. van IJcken, Joost Gribnau
Sex differences and risk factors for bleeding in Alagille syndrome Simona Hankeova, Noemi Van Hul, Jakub Laznovsky, Katrin Mangold, Naomi Hensens, Elvira Verhoef, Tomas Zikmund, Feven Dawit, Michaela Kavkova, Jakub Salplachta, Marika Sjöqvist, Bengt R. Johansson, Mohamed Hassan, Linda Fredriksson, Vitezslav Bryja, Urban Lendahl, Andrew Jheon, Florian Alten, Kristina Teär Fahnehjelm, Björn Fischler, Jozef Kaiser, Emma R. Andersson
Dynamic regulation and requirement for ribosomal RNA transcription during mammalian development Karla T. Falcon, Kristin E.N. Watt, Soma Dash, Annita Achilleos, Ruonan Zhao, Daisuke Sakai, Emma L. Moore, Sharien Fitriasari, Melissa Childers, Mihaela E. Sardiu, Selene Swanson, Dai Tsuchiya, Jay Unruh, George Bugarinovic, Lin Li, Rita Shiang, Jill Dixon, Michael J. Dixon, Paul A. Trainor
Tig1 regulates proximo-distal identity during salamander limb regeneration Catarina R. Oliveira, Dunja Knapp, Ahmed Elewa, Tobias Gerber, Sandra G. Gonzalez Malagon, Phillip B. Gates, Hannah E. Walters, Andreas Petzold, Hernan Arce, Rodrigo C. Cordoba, Elaiyaraja Subramanian, Osvaldo Chara, Elly M. Tanaka, András Simon, Maximina H. Yun
High resolution snRNA-seq analysis of Drosophila Malpighian tubules from Xu, et al.
A cell atlas of the fly kidney Jun Xu, Yifang Liu, Hongjie Li, Alexander J. Tarashansky, Colin H. Kalicki, Ruei-Jiun Hung, Yanhui Hu, Aram Comjean, Sai Saroja Kolluru, Bo Wang, Stephen R Quake, Liqun Luo, Andrew P. McMahon, Julian A.T. Dow, Norbert Perrimon
Bringing TrackMate into the era of machine-learning and deep-learning Dmitry Ershov, Minh-Son Phan, Joanna W. Pylvänäinen, Stéphane U. Rigaud, Laure Le Blanc, Arthur Charles-Orszag, James R. W. Conway, Romain F. Laine, Nathan H. Roy, Daria Bonazzi, Guillaume Duménil, Guillaume Jacquemet, Jean-Yves Tinevez
A reference tissue atlas for the human kidney Jens Hansen, Rachel Sealfon, Rajasree Menon, Michael T. Eadon, Blue B. Lake, Becky Steck, Dejan Dobi, Samir Parikh, Tara K. Sigdel, Guanshi Zhang, Dusan Velickovic, Daria Barwinska, Theodore Alexandrov, Priyanka Rashmi, Edgar A. Otto, Michael P. Rose, Christopher R. Anderton, John P. Shapiro, Annapurna Pamreddy, Seth Winfree, Yongqun He, Ian H. de Boer, Jeffrey B. Hodgin, Laura Barisoni, Abhijit S. Naik, Kumar Sharma, Minnie M. Sarwal, Kun Zhang, Jonathan Himmelfarb, Brad Rovin, Tarek M. El-Achkar, Zoltan Laszik, John Cijiang He, Pierre C. Dagher, M. Todd Valerius, Sanjay Jain, Lisa Satlin, Olga G. Troyanskaya, Matthias Kretzler, Ravi Iyengar, Evren U. Azeloglu, for the Kidney Precision Medicine Project
ShareLoc – an open platform for sharing localization microscopy data Jiachuan Bai, Wei Ouyang, Manish Kumar Singh, Christophe Leterrier, Paul Barthelemy, Samuel F.H. Barnett, Teresa Klein, Markus Sauer, Pakorn Kanchanawong, Nicolas Bourg, Mickael M. Cohen, Benoît Lelandais, Christophe Zimmer
Julia for Biologists Elisabeth Roesch, Joe G. Greener, Adam L. MacLean, Huda Nassar, Christopher Rackauckas, Timothy E. Holy, Michael P.H. Stumpf
In the latest episode of Genetics Unzipped, Kat Arney explores the science behind one of the most remarkable but often overlooked organs in the mammalian body: the placenta.
To find out more, Kat chats with Ros John, who leads the Pregnancy Research Epigenetics Group or PREGLab at Cardiff University. Ros’s research focuses on understanding maternal mental health, imprinted genes and the role of the placenta during pregnancy and even beyond, with big implications for the health of both mother and child.
Kat also speaks to Sam Behjati, a group leader at the Wellcome Sanger Institute, who recently made the surprising discovery that the placenta is a genetic ‘dumping ground’. The pattern of genetic alterations in the placenta is different to any other human organ and resembles that of a tumour, harbouring many of the same genetic mutations found in childhood cancers.
If you enjoy the show, please do rate and review on Apple podcasts and help to spread the word on social media. And you can always send feedback and suggestions for future episodes and guests to podcast@geneticsunzipped.com Follow us on Twitter – @geneticsunzip
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