초파리 배아, 유충의 준비
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Q1: How do you set up an egg-laying chamber to collect Drosophila embryos?
Create an egg-laying cup by poking holes in a container and covering it with porous material for ventilation. Prepare an apple or grape juice agar plate, streak it with yeast paste, and scratch the center. Add flies to the chamber, invert it so the plate is at the bottom, and incubate. Replace the plate every 1-3 hours to obtain best-aged embryos. Twenty flies of each sex produce 100-200 embryos per hour.
Q2: What is the purpose of dechorionation in Drosophila embryo preparation?
Dechorionation removes the hard outer membrane, or chorion, from embryos, which is essential for techniques like microinjection and live cell imaging. This can be done manually by dissecting the embryo from its chorionic sheath or chemically using 50% bleach for 2-10 minutes. Thorough rinsing with distilled water ensures the naked embryo remains undamaged.
Q3: How are Drosophila larvae collected using a sucrose solution?
Add sucrose solution to the vial containing larvae; since it is denser than larvae, they float to the top. Place the vial on a rotator to dislodge food from the medium. Remove larvae with a brush or pipette for harvesting and experimentation. This method is more efficient than manual collection when gathering large numbers of early-stage larvae.
Q4: What are imaginal discs and why are they important in Drosophila research?
Imaginal discs are partially developed organs in larvae that become whole adult structures. An imaginal eye disc becomes an adult eye, antennal discs become antennae, and wing discs become wings. Study of imaginal discs has revealed important discoveries about development and reproduction drosophila melanogaster, including the role of homeobox genes in pattern formation.
Q5: What are the three larval instar stages in Drosophila development?
Drosophila larvae progress through three instar, or molting, stages. The first instar lasts one day, the second instar lasts one day, and the third instar lasts two days. First and second instar larvae are found in food 1-3 days after setting up a cross. On day four, third instar larvae migrate up the container sides to form pupae.
Q6: How is microinjection used to create transgenic Drosophila mutants?
Microinjection inserts customized genetic material in the form of circular DNA plasmids into dechorionated embryos. Embryos are dechorionated by physically rolling them on double-sided tape to avoid chemical damage. Plasmids encoding proteins like tubulin fused with fluorescent reporter proteins such as GFP can be injected to create transgenic fly lines for visualizing cellular processes.
Q7: What does a crawling assay measure in Drosophila larvae?
A crawling assay evaluates Drosophila locomotor behavior by measuring the distance a larva travels under specific conditions. Collected larvae are immersed in a drugged sucrose solution predicted to interfere with motility, allowing researchers to observe the effects of drugs on motor function and behavioral responses.