Probe:	pCR-mRGS4
Probe preparation:	Antisense labelled with digoxigenin RNA

Structure	Level	Pattern	Image	Note
TS17: rhombomere 4	Present	Regionally restricted	1A	Expressed in the ventral column.
TS17: rhombomere 1	Present	Regionally restricted	1A	Expressed in the anlage of the trochlear nucleus, located in rhombomere 1. But not the nearby oculomotor nucleus, located in the caudal mesencephalon (data not shown).
TS17: midbrain-hindbrain junction	Present		1A
TS17: rhombomere 2	Present	Regionally restricted	1A	Expressed in the ventral column, that is, in the precursors of the trigeminal motor nucleus.
TS17: rhombomere 3	Present	Regionally restricted	1A	Expressed in the ventral column, that is, in the precursors of the trigeminal motor nucleus.

Structure	Level	Pattern	Image	Note
TS17: rhombomere 4	Present	Regionally restricted	1B	Expressed in the ventral region, in a subpopulation of Phox2b+ precursors localized at the border between the ventricular zone, which they had probably just exited, and the mantle layer.

Structure	Level	Pattern	Image	Note
TS17: rhombomere 4	Present	Regionally restricted	1C	Expressed in the ventral region, which had already undergone their last S phase. The more superficially located, thus presumably older postmitotic precursors, had already lost Rgs4 expression.

Structure	Level	Pattern	Image	Note
TS20: hindbrain	Present	Regionally restricted	1D	Expressed in rhombomere 4 (r4). Expression was extinguished before motoneuronal precursors have started to migrate through r5, or in migratory inner ear efferent precursors (data not shown).

Structure	Level	Pattern	Image	Note
TS21: facial nucleus	Present		1E	Expressed in facial motoneuronal precursors as they settle in the facial nucleus anlage.

Structure	Level	Pattern	Image	Note
TS28: facial nucleus	Strong		1F

Structure	Level	Pattern	Image	Note
TS28: trigeminal nucleus	Strong		1G

Structure	Level	Pattern	Image	Note
TS17: hindbrain	Present	Regionally restricted	2A	Expressed in the locus coeruleus and in a neighboring population of interneurons in the rostral hindbrain.

Structure	Level	Pattern	Image	Note
TS18: rhombomere 1	Present	Regionally restricted	2B	Expressed in locus coeruleus precursors in the rostral hindbrain, simultaneously with Phox2b (data not shown), that is soon after Phox2a expression.

Structure	Level	Pattern	Image	Note
TS21: brainstem nucleus	Present	Regionally restricted	2C	Expression in the locus coeruleus was extinguished at this stage. Expressed in facial motoneuronal precursors as they settle in the facial nucleus anlage (see 1E).

Structure	Level	Pattern	Image	Note
TS28: locus coeruleus	Present		2D	Expressed in the mature locus coeruleus neurons.

Structure	Level	Pattern	Image	Note
TS17: sympathetic nervous system	Present	Regionally restricted	3A	Expressed as soon as E10.5 in the aggregating sympathoblasts on each side of the dorsal aorta.

Structure	Level	Pattern	Image	Note
TS21: superior cervical ganglion	Strong		3B	Expressed in a majority of cells.
TS21: glossopharyngeal ganglion	Absent		3B	At this stage expression was lost in the nodose-petrosal complex (gIX-X).
TS21: vagus ganglion	Absent		3B	At this stage expression was lost in the nodose-petrosal complex (gIX-X).

Structure	Level	Pattern	Image	Note
TS21: sympathetic ganglion	Present	Regionally restricted	3C	Expressed in a prevertebral Phox2a+ cell mass below and lateral to the celiac ganglion.

Structure	Level	Pattern	Image	Note
TS21: sympathetic ganglion	Present	Regionally restricted	3D	Expressed in a prevertebral Phox2b+ cell mass below and lateral to the celiac ganglion.

Structure	Level	Pattern	Image	Note
TS21: sublingual ganglion	Strong		3E

Structure	Cell Type	Level	Pattern	Image	Note
TS21: enteric nervous system	enteric neuron	Present	Single cells	3F	Expressed in a small subset of enteric neurons.

Structure	Level	Pattern	Image	Note
TS16: facio-acoustic ganglion complex	Present		4A
TS16: glossopharyngeal ganglion	Present		4A
TS16: trigeminal ganglion	Present		4A
TS16: vagus ganglion	Present		4A
TS16: future hindbrain	Present	Regionally restricted	4A	Expression in the locus coeruleus had just started.
TS16: dorsal root ganglion	Strong	Scattered	4A	Expressed in a salt and pepper pattern.

Structure	Level	Pattern	Image	Note
TS16: facio-acoustic ganglion complex	Present		4B

Structure	Level	Pattern	Image	Note
TS17: glossopharyngeal ganglion	Present	Regionally restricted	4C	Phox2a+ cells delaminated from the glossopharyngeal placode (pIX) and switched on Rgs4 as they aggregated to form the ganglion anlage (gIX).

Structure	Level	Pattern	Image	Note
TS21: dorsal root ganglion	Present	Widespread	4D	A widespread expression, albeit at variable levels, in a salt and pepper pattern.

Structure	Level	Pattern	Image	Note
TS28: dorsal root ganglion	Strong	Regionally restricted	4E	Strong expression in a subset of small-diameter neurons.

Structure	Level	Pattern	Image	Note
TS17: rhombomere 4	Present	Regionally restricted	5A	Expressed in facial motoneuronal precursors in the ventral region.

Structure	Level	Pattern	Image	Note
TS17: rhombomere 4	Absent		5B

Structure	Level	Pattern	Image	Note
TS17: rhombomere 4	Present	Regionally restricted	5C	Expressed in facial motoneuronal precursors in the ventral region.
TS17: midbrain	Present	Regionally restricted	5C	Expressed in precursors of the trochlear and trigeminal motor nuclei and the locus coeruleus.
TS17: rhombomere 2	Present		5C
TS17: rhombomere 3	Present		5C

Structure	Level	Pattern	Image	Note
TS17: rhombomere 2	Absent		5D
TS17: rhombomere 3	Absent		5D
TS17: rhombomere 4	Absent		5D
TS17: midbrain	Present	Regionally restricted	5D	No expressed in precursors of the trochlear and trigeminal motor nuclei and the locus coeruleus.

Structure	Level	Pattern	Image	Note
TS17: rhombomere 1	Present	Regionally restricted	5E	Expressed in the locus coeruleus.

Structure	Level	Pattern	Image	Note
TS17: rhombomere 1	Absent		5F	No expression in locus coeruleus (Phox2a+).

Structure	Level	Pattern	Image	Note
TS17: sympathetic nervous system	Present	Regionally restricted	5G	Expressed in the aggregating sympathoblasts on each side of the dorsal aorta.

Structure	Level	Pattern	Image	Note
TS17: sympathetic nervous system	Ambiguous		5H	No expression in the aggregating sympathoblasts on each side of the dorsal aorta.

Structure	Level	Pattern	Image	Note
TS17: acoustic ganglion	Present		6A
TS17: dorsal root ganglion	Present		6A
TS17: facial ganglion	Present		6A
TS17: glossopharyngeal ganglion	Present		6A
TS17: vagus ganglion	Present		6A
TS17: sympathetic nerve trunk	Present		6A	sympathetic chain

Structure	Level	Pattern	Image	Note
TS17: facial ganglion	Present		6B	Expression was downregulated but not extinguished.
TS17: glossopharyngeal ganglion	Present		6B	Expression was downregulated but not extinguished.
TS17: vagus ganglion	Present		6B	Expression was downregulated but not extinguished.
TS17: acoustic ganglion	Present		6B	Similar as in the heterozygous.
TS17: dorsal root ganglion	Present		6B	Similar as in the heterozygous.
TS17: sympathetic nerve trunk	Absent		6B
TS17: midbrain	Present	Regionally restricted	6B	No expressed in precursors of the locus coeruleus.

Structure	Level	Pattern	Image	Note
TS17: acoustic ganglion	Present		6C
TS17: dorsal root ganglion	Present		6C
TS17: facial ganglion	Present		6C
TS17: glossopharyngeal ganglion	Present		6C
TS17: vagus ganglion	Present		6C
TS17: sympathetic nerve trunk	Present		6C	sympathetic chain

Structure	Level	Pattern	Image	Note
TS17: facial ganglion	Present		6D	Expression was downregulated but not extinguished.
TS17: glossopharyngeal ganglion	Present		6D	Expression was downregulated but not extinguished.
TS17: vagus ganglion	Present		6D	Expression was downregulated but not extinguished.
TS17: acoustic ganglion	Present		6D	Similar as in the heterozygous.
TS17: dorsal root ganglion	Present		6D	Similar as in the heterozygous.
TS17: sympathetic nerve trunk	Present		6D	Similar as in the heterozygous.
TS17: midbrain	Present	Regionally restricted	6D	No expression in the locus coeruleus, which does not form in Phox2a mutants.

Structure	Level	Pattern	Image	Note
TS17: enteric nervous system	Present	Single cells		Very transiently detected in a sparse subset of Phox2b+ migrating precursors.

Structure	Level	Pattern	Image	Note
TS19: myelencephalon	Present	Regionally restricted		Similar expression pattern in rhombomere 4 as in that of E10.5.
TS19: enteric nervous system	Absent

Structure	Level	Pattern	Image	Note
TS21: cervico-thoracic ganglion	Present			Expressed in a majority of cells in stellate ganglion.
TS21: thoracic ganglion	Present	Regionally restricted		Expressed in fewer cells in the trunk sympathetic chain, tended to be clustered.
TS21: paracardiac ganglion	Strong
TS21: submandibular ganglion	Strong
TS21: parasympathetic ganglion	Strong	Regionally restricted		Expressed strongly in pulmonary parasympathetic ganglion. Very few positive cells in ciliary ganglion.
TS21: otic ganglion	Present	Single cells		Very few positive cells.
TS21: sphenopalatine ganglion	Present	Single cells		Very few positive cells.

Structure	Cell Type	Level	Pattern	Image	Note
TS22: hindbrain		Present	Regionally restricted		Expressed in the nVII region.
TS22: adrenal medulla		Present
TS22: dorsal root ganglion		Present
TS22: parasympathetic ganglion		Present
TS22: sympathetic ganglion		Present
TS22: vestibulo-cochlear ganglion		Present
TS22: enteric nervous system	enteric neuron	Trace			Rare.
TS22: facial ganglion		Absent
TS22: glossopharyngeal ganglion		Absent
TS22: trigeminal ganglion		Absent
TS22: vagus ganglion		Absent

Structure	Level	Pattern	Image	Note
TS23: facial nucleus	Absent			Expression was virtually extinguished again by E15.5.
TS23: spinal cord white matter	Absent			No expression was detected in embryonic spinal motoneuronal precursors, whether somatic or visceral, at least until E15.5.
TS23: cerebral cortex	Absent			Undetectable embryonically, at least until E15.5.
TS23: dorsal root ganglion	Trace			Expression had faded out in the vast majority of cells.