khushi singh.in

Research · Model system

Model system: the fruit fly

Above all the fruit fly, Drosophila melanogaster: its egg chambers in the ovary and its earliest embryo, at the moment the embryo’s own genes take over from its mother’s. Here is why biologists use it, how it develops, the tools that make it powerful, and the other organisms and proteins I have worked with.

Main model organism

Drosophila melanogaster, the fruit fly

Most of my research has followed the fly from egg chamber to early embryo: oogenesis in the ovary, and embryogenesis up to stage 5, when the maternal-to-zygotic transition hands control from the mother’s RNAs and proteins to the embryo’s own genes.

Projects: Microbes, lactate and inheritance · Behaviour: activity and sleep · oskar RNA condensates · ER shape and autophagy · NCBI Taxonomy 7227

  1. 1 Egg chamber, stage 9: oskar marks the germ plasm
  2. 2 The mature egg
  3. 3 Embryo, stage 5: cells form as the embryo’s genes switch on
  • ≈ 10 daysfrom egg to adult at 25 °C
  • ≈ 14,000protein-coding genes on four chromosome pairs
  • 14stages of egg-chamber development in the ovary
  • 1 + 15oocyte and nurse cells in every germline cyst
  • 13nuclear divisions before the embryo forms cells
  • ≈ 6,000cells form at once in the stage 5 embryo
  • 130–170minutes after egg laying: stage 5, at 25 °C

From FlyBase (Öztürk-Çolak et al. 2024; stage definitions FBdv), Adams et al. 2000, Jennings 2011, Bastock & St Johnston 2008, Foe & Alberts 1983 and Mazumdar & Mazumdar 2002.

Why biologists use it

Flies breed fast, about ten days from egg to adult at 25 °C, and are cheap to keep in large numbers. Their genome was sequenced in 2000, with about 14,000 protein-coding genes on four pairs of chromosomes, and most human disease-associated genes have a recognisable counterpart in the fly. Genetic switches turn a chosen gene on or off in a single tissue. The germline is set aside very early by the germ plasm at the back of the egg, so what a mother puts into her egg can be followed into the embryo.

  • Of 929 human disease genes checked, 77% had a recognisable match in the fly genome (Reiter et al. 2001); a later estimate is about 65% (Ugur et al. 2016). Established
  • The GAL4/UAS system switches a chosen gene on in a chosen tissue (Brand & Perrimon 1993). Established

Life cycle

At 25 °C an egg hatches after about a day; the larva feeds and grows through three stages for about four days, then forms a pupa, in which the adult body is built over about four more days. Adults live for one to two months, and a female can lay up to about 100 eggs a day (Jennings 2011). Established

~10 daysegg → adult at 25 °CEgg~1 dayLarva (3 stages)~4 daysPupa~4 daysAdultlives ~1–2 months

From egg chamber to embryo

Oogenesis. In the ovary, a germline stem cell divides to make a cyst of 16 connected cells: one becomes the oocyte and 15 become nurse cells, which pour their RNAs and proteins into it. Somatic follicle cells wrap the cyst into an egg chamber, which passes through 14 stages. Along the way, mRNAs such as oskar and bicoid are placed at the back and front of the oocyte, laying down the future body axes and the germ plasm (Bastock & St Johnston 2008). Established

Embryogenesis to stage 5. After fertilisation the nuclei divide 13 times in about two hours without forming cells, sharing one cytoplasm (Foe & Alberts 1983). Pole cells, the future eggs and sperm, bud off at the back. In stage 5, membranes grow between about 6,000 nuclei at once, and the embryo strongly switches on its own genes, helped by the pioneer factor Zelda (Liang et al. 2008), while maternal mRNAs are cleared: the maternal-to-zygotic transition (Vastenhouw et al. 2019). Established

Step through oogenesis, the hand-over from egg to embryo, and the first seven embryonic stages. Stage 5, the cellular blastoderm at the maternal-to-zygotic transition, is where most of my work sits.

In the mother’s ovary: oogenesis

Egg to embryo

In the laid egg: embryonic stages

pole cellsmembranes grow between the nucleia layer of about 6,000 cells around the yolkanteriorposterior
Schematic drawing, anterior left and dorsal up. Blue: nuclei · magenta: membranes and follicle cells · green: oocyte · amber: germ plasm and pole cells.

Embryonic stage 5 (cellular blastoderm)

Embryonic stage 5: cellularization and the hand-over

Time (25 °C)
130–170 min after egg laying
Nuclei
Interphase of cycle 14
Mother’s RNAs and proteins
Being cleared (maternal and zygotic pathways)
Embryo’s own genes
Major wave

Stage 5 is the cellular blastoderm, lasting about 40 minutes at 25 °C in the long interphase of cycle 14. It is the heart of the maternal-to-zygotic transition: the major wave of zygotic genome activation, while maternal mRNAs are cleared by a maternal and a zygotic pathway. The pole cells keep their own genes quiet.

In my research. Stage 5 embryos at the maternal-to-zygotic transition are the focus of my current project, which asks how chromatin marks change at this hand-over (MPI-IE, 2026–2027).

FlyBase stage term: FBdv:00005311

13 of 15

Showing Embryonic stage 5: cellularization and the hand-over.

The maternal-to-zygotic transition, stage by stage
1234567StageNuclear divisionsCellsGerm line (pole cells)Embryo’s own genesMother’s mRNAs: maternal decayMother’s mRNAs: zygotic decay1–23–8910–13interphase 14one shared cytoplasm (syncytium)cellularizationformtranscriptionally quietminor wavemajor waveSMAUG-dependent, from egg activatione.g. miR-309 cluster0306090120150180minutes after egg laying (25 °C)▲

Bars show when each process is under way, not how much; arrows mean it continues. Stage times from FlyBase (FBdv). Sources: Foe & Alberts 1983; Tadros et al. 2007; Bushati et al. 2008; Liang et al. 2008; Tadros & Lipshitz 2009; Van Doren et al. 1998.

Timeline as a table
StageMinutes after egg laying (25 °C)NucleiMother’s RNAsEmbryo’s genes
10–25Divisions 1–2In chargeSilent
225–65Divisions 3–8In chargeSilent; the first transcripts appear near the end (about cycle 8)
365–80Division 9In chargeMinor wave
480–130Divisions 10–13In chargeMinor wave (Zelda-dependent)
5130–170Interphase of cycle 14Being cleared (maternal and zygotic pathways)Major wave
6170–180Cycle 14Largely replaced by zygotic productsOn
7180–190Cycle 14Largely replaced by zygotic productsOn

Stage 5 in 3D

About 6,000 nuclei sit just under the surface of the stage 5 embryo. Pick a gene to see where it is switched on, from the maternal bicoid gradient to the seven stripes of even-skipped, made by the embryo’s own genome.

A fly embryo at stage 5, in 3D

About 6,000 nuclei, the moment the embryo’s own genes switch on.

even-skipped: pair-rule (7 stripes). Anterior (head end) is on the left of the long axis. Drag to rotate.

Model generated in code: patterns simplified from textbook knowledge (e.g. 7 eve stripes), not measured data.

The genetic toolkit

  • Switch a gene on in one tissue. GAL4/UAS: a “driver” line decides where, a “responder” line decides which gene. Cross them and the gene is active only where the driver works (Brand & Perrimon 1993).
  • Turn a gene down. Genome-wide collections of RNAi lines let researchers reduce almost any gene, in a chosen tissue, with GAL4 (Dietzl et al. 2007; Perkins et al. 2015).
  • Edit the genome. CRISPR/Cas9 tools for flies make precise mutations, tags and deletions in the germline or in single tissues (Port et al. 2014).
  • Keep complex stocks for years. Balancer chromosomes carry inversions that block recombination and visible markers such as curly wings, so combinations of mutations can be kept and recognised by eye (Kaufman 2017).
  • Shared resources. FlyBase collects genes, expression data and stocks (Öztürk-Çolak et al. 2024); public stock centres send fly lines worldwide.

Try the logic of GAL4/UAS: pick a driver (where) and a responder (what).

1 · The driver (GAL4): where
2 · The reporter (UAS): what
abdomen (gut, ovaries)thoraxhead

The gut driver switches on the reporter only in the gut, which now glow.

Two separate parts must meet in one fly: the driver says where, the reporter says what. Scientists combine them by crossing flies.

Discoveries made in flies

  1. 1933 Genes sit on chromosomes (Thomas Hunt Morgan)
  2. 1946 X-rays cause mutations (Hermann J. Muller)
  3. 1995 Genes that build the embryo’s body plan (Edward B. Lewis, Christiane Nüsslein-Volhard, Eric F. Wieschaus)
  4. 2011 How innate immunity is switched on, first found through Toll in flies (Jules A. Hoffmann, shared prize)
  5. 2017 The molecular clock behind circadian rhythms (Jeffrey C. Hall, Michael Rosbash, Michael W. Young)
  6. 2024 A wiring diagram of a whole adult fly brain: 139,255 neurons (Dorkenwald et al. 2024)

Nobel Prizes in Physiology or Medicine, from nobelprize.org; 2024 from Dorkenwald et al. 2024.

The adult fly in 3D

A fruit fly, much bigger than life

Rotate with a finger or mouse. On a phone you can place it in your room (AR).

  • Brain: about 140,000 neurons, mapped one by one in 2024
  • Gut: home to a few kinds of bacteria
  • Ovaries: where egg chambers develop
  • Haltere: balance organ: 2 wings + 2 halteres

Stylised Drosophila melanogaster built in code (anatomy: 6 legs, 2 wings, halteres, red eyes). Model: khushisingh.in, CC BY 4.0.

A stylised model built in code. Real flies are about 2.5 mm long.

What I did

Worked with: Oogenesis (egg chambers in the ovary) and early embryogenesis, above all stage 5 embryos at the maternal-to-zygotic transition.

  • Reduced an ER-shaping protein by RNAi and studied autophagy, ubiquitination and tissue structure in embryos and ovaries (IISER Pune, 2024–2025).
  • Optimised single-molecule FISH to image oskar RNA condensates in embryos (IIT Kharagpur, 2025).
  • Now asking whether metabolism shapes histone marks in the early embryo, and how it shapes activity and sleep (MPI-IE, 2026–2027, ongoing).

Other organisms and proteins

Insects and plants on the iiser pune campus

Field work

A field project on how insects respond to light and how plants and their pathogens interact across the campus (IISER Pune, 2023). It is where the habit of careful observation began.

Ecology and evolution

Proteins from other species

In these projects I worked with purified proteins in the test tube, not with the organisms themselves. Borrowing a protein from the species where it is easiest to study is common in biochemistry.

  • Escherichia coli

    Gut bacterium and lab workhorse

    PstS, the phosphate-binding protein

    Engineered and labelled with a dye so that it glows brighter when it binds phosphate: a real-time phosphate sensor (Brune et al. 1994).

    What I did: Prepared, labelled and calibrated the sensor (IISER Pune, 2025–2026).

    Phosphate biosensor and FtsZ · NCBI Taxonomy 562

  • Spiroplasma melliferum

    A bacterium without a cell wall

    FtsZ

    The bacterial cousin of tubulin, which forms the ring that divides a bacterium in two (Bi & Lutkenhaus 1991).

    What I did: Followed its use of GTP in real time with the phosphate sensor and fitted the kinetics (IISER Pune, 2025–2026).

    Phosphate biosensor and FtsZ · NCBI Taxonomy 2134

  • Mus musculus

    House mouse

    Prion protein (PrP)

    A normal protein of nerve cells that can misfold and spread in prion diseases (Prusiner 1998).

    What I did: Mapped the salt, pH and protein concentrations at which purified PrP forms liquid droplets or clumps (IISER Pune, 2024).

    Prion protein phase separation · NCBI Taxonomy 10090

← All research · The methods (toolbox) · Policy brief: why model organisms are worth investing in

References

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