• What did you learn from surfing FlyBase?

• Why do the inversions in Balancer chromosomes greatly reduce the frequency of crossing over in meiosis?

• Autonomous vs non autonomous

Nonautonomous

• Domineering - mutant cells disrupt the development of neighboring wild type cells.

• Submissive - wild type neighbors rescue mutant cells.

• FLP/FRT can also be used to remodel chromosomes.

RNAi in flies

ds RNA in cultured cells.

http://flyrnai.org/RNAi_goals.html

RNAi in flies

1. Express a fold back RNA from a transgene.

2. This results in tissue/temporal specific gene knockdowns.

3. Collections of transgenic flies are available that contain transgenes to knockdown the expression of most fly genes.

Gene targeting

• Until recently not available in flies.

• One difficulty is that one cannot select for a targeted cell.

• A technique has now been developed that has been shown to work on a number of genes.

• Technique requires first getting a germ line transformant for the knock out construct.

• The DNA for targeting is excised by the activity of the FLP/FRT site specific recombination system.

• The excised circle is then cut by the induced expression of a very rare cutting RE.

• High rate of homologous targeting.

In Vivo Imaging

• A number of developmental stages/tissues provide opportunities for in vivo imaging.

Genetic Screens

• Recessive lethal mutations

The iso la tion of recessive v isib le m uta tions

m utagenize X th st fz in ri/TM 3

screen for flies that are phenotypica lly or fz in .

these flies would be or fz*th st/fz in ri th st in*/fz in ri

X

The iso la tion of recessive le tha l m utations

cn bw treat w ith m utagen (e.g. em s)

cn bw X G la/CyO

ind ividual cn bw /C yO X G la/CyO

* *

***

*

*

cross s ib lings X cn bw /C yO cn bw /CyO

cn bw / cn bwcn bw /C yO2

CyO /CyO

Look for v ia ls w ith no stra ight w inged flies

Flp/FRT

• High efficiency of FLP/FRT clone induction has allowed new types of mutant screens.