wild type cd1 embryos were harvested , staged and stained by wholemount in situ hybridization using established methods , .
measurement of palates before dissection and after staining confirmed that no significant shrinkage occurred ( data not shown ) .
palatal explants were cultured ( 37c , 5% co2 ) using the trowell technique   in dmem ( sigma ) , 20 u / ml pen - strep ( gibcobrl ) , 10% fbs ( gibcobrl ) , 50 mm transferrin ( sigma ) and 150 g / ml ascorbic acid ( sigma ) . for cutting and inhibitor experiments ,
serum - free advanced d - mem / f12 ( gibcobrl ) supplemented as above , was used .
su5402 ( calbiochem ) was diluted in medium from 10 mm in dmso stock ; cyclopamine ( sigma ) from 20 mg / ml - ethanol stock .
explants placed in a minimum volume of pbs in wells cut into 1% agarose were digitally imaged under a stereo dissecting microscope and measurements made using the ruler in imagej ( from the nih imagej website ) calibrated with a micrometer slide .
dimensions of fixed material were within 8% of those of fresh , unfixed material ( data not shown ) .
simulations of reaction - diffusion patterning were performed using the javascript in ref   with the parameters du=0.03 , d u=0.02 , a u=0.1 , b u=0.06 , c u=0 , fmax=0.2 , dv= 0.06 , d v=0.5 , a v=0.1 , b v=+0 , c v=0.2 , gmax=0.5 .
wild type cd1 embryos were harvested , staged and stained by wholemount in situ hybridization using established methods , .
measurement of palates before dissection and after staining confirmed that no significant shrinkage occurred ( data not shown ) .
palatal explants were cultured ( 37c , 5% co2 ) using the trowell technique   in dmem ( sigma ) , 20 u / ml pen - strep ( gibcobrl ) , 10% fbs ( gibcobrl ) , 50 mm transferrin ( sigma ) and 150 g / ml ascorbic acid ( sigma ) . for cutting and inhibitor experiments , serum - free advanced d - mem / f12 ( gibcobrl ) supplemented as above ,
su5402 ( calbiochem ) was diluted in medium from 10 mm in dmso stock ; cyclopamine ( sigma ) from 20 mg / ml - ethanol stock .
explants placed in a minimum volume of pbs in wells cut into 1% agarose were digitally imaged under a stereo dissecting microscope and measurements made using the ruler in imagej ( from the nih imagej website ) calibrated with a micrometer slide .
dimensions of fixed material were within 8% of those of fresh , unfixed material ( data not shown ) .
simulations of reaction - diffusion patterning were performed using the javascript in ref   with the parameters du=0.03 , d u=0.02 , a u=0.1 , b u=0.06 , c u=0 , fmax=0.2 , dv= 0.06 , d v=0.5 , a v=0.1 , b v=+0 , c v=0.2 , gmax=0.5 .
 we present direct evidence of an activator - inhibitor system in the generation of the regularly spaced transverse ridges of the palate . 
 we show that new ridges , or rugae , marked by stripes of sonic hedgehog ( shh ) expression , appear at two growth zones where the space between previously laid - down rugae increases . 
 however , inter - rugal growth is not absolutely required : new stripes still appear when growth is inhibited . 
 furthermore , when a ruga is excised new shh expression appears , not at the cut edge but as bifurcating stripes branching from the neighbouring shh stripe , diagnostic of a turing - type reaction - diffusion mechanism . 
 genetic and inhibitor experiments identify fibroblast growth factor ( fgf ) and shh as an activator - inhibitor pair in this system . 
 these findings demonstrate a reaction - diffusion mechanism likely to be widely relevant in vertebrate development . 
Supplementary Material
Methods
Generation of embryos
Explant culture
Imaging, measuring and simulations