Vol.
49,
No.
1
APPLIED
AND
ENVIRONMENTAL
MICROBIOLOGY,
Jan.
1985,
p.
187-191
0099-2240/851010187-05$02.00/0
Copy igh
C
1985,
Ame ican
Socie y
o
Mic obiology
Gibbe ella
ujiku oi
Mu an s
Ob ained
wi h
UV
Radia ion
and
N-
Me hyl-N'
-Ni o-N-Ni osoguanidine
J.
AVALOS,
J.
CASADESUS,
AND
E.
CERDA
OLMEDO*
Depa amen o
de
Gene ica,
Facul ad
de
Biologia,
Uni e sidad
de
Se illa,
Se illa,
Spain
Recei ed
9
Ma ch
1984/Accep ed
16
Oc obe
1984
N-me hyl-N'-ni o-N-ni osoguanidine
(ni osoguanidine)
and
o
a
lesse
ex en
UV
adia ion
a e
e y
mu agenic
o
Gibbe ella
mic oconidia.
The
ecommended
ni osoguanidine
doses
lead
o
much
highe
equencies
o
mu an s
han
a e
ound
in
o he
mic oo ganisms.
The
equency
o
mu an s
among
he
su i o s
inc eases
linea ly
wi h
he
ni osoguanidine
dose
(mola
concen a ion
x
ime);
he
absolu e
numbe
o
iable
mu an s
in
a
gi en
popula ion
eaches
a
maximum
o
a
dose
o
ca.
0.7
M
*
s.
The
mic oconidia
a e
uninuclea e.
The
onse
o
ge mina ion
b ings
abou
inc eased
le hali y
o
ni osoguanidine,
bu
i
does
no
modi y
he
ac ion
o
UV
adia ion.
Mycelia
a e
mo e
esis an
han
spo es
o
bo h
agen s.
Visible
illumina ion
e ec i ely
p e en s
le hali y
when
gi en
immedia ely
a e
UV
i adia ion.
Auxo ophs
and
colo
mu an s
a e
e y
easily
ob ained.
Pink
adenine
auxo ophs
and
se e al
classes
o
colo
mu an s
a e
a ec ed
in
he
biosyn hesis
o
he
ca o enoid
pigmen ,
neu ospo axan hin.
The
indus ial
in e es
in
he
ungus
Gibbe ella
ujiku oi
(as
an
impe ec
ungus,
known
also
as
Fusa ium
monili-
o me)
s ems
om
i s
abundan
p oduc ion
o
gibbe ellins,
plan
ho mones
widely
used
in
ho icul u e
and
b ewing
(15).
The
o ganism
may
also
ha e
a
u u e
in
he
indus ial
p oduc ion
o
p o ein
and
o
ca o enoids.
Mo phological
and
biochemical
mu an s
o
G.
ujiku oi
ha e
been
isola ed
a e
ea men
wi h
di e en
chemical
and
physical
mu agens
(1,
10,
11, 18,
24).
No
su p isingly,
hese
e o s
ha e
been
concen a ed
la gely
on
mu an s
a ec ing
gibbe ellin
p oduc ion.
N-me hyl-N'-ni o-N-ni osoguanidine
(ni osoguanidine)
has
been
ex ensi ely
used
o
he
induc ion
o
mu a ions
in
many
o ganisms
(12).
The
e ec s
o
he
d ug,
including
he
equency
o
mu an s
and
hei
dis ibu ion
o e
he
genome,
a y
om
one
o ganism
o
ano he .
In
yeas s
(8),
as
in
bac e ia
(5),
he
mu a ions
a e
p e e en ially
induced
in
DNA
egions
being
eplica ed
a
he
ime o
mu agen
expo-
su e.
In
bac e ia
(13),
bu
no
in
yeas s
(2),
his
esul s
in
ela i ely
high
a es
o
closely
linked
double
mu an s.
De ailed
s udies
o
he
applica ion
o
ni osoguanidine
o
he
ungi
Saccha omyces
ce e isiae
and
Phycomyces
spp.
ha e
ecen ly
been
published
by
Calde on
and
Ce da'-
Olmedo
(2)
and
by
Ronce o
e
al.
(21),
espec i ely.
We
in es iga ed
he
e ec s
o
ni osoguanidine
on
G.
ujiku oi
and
compa ed
hem
wi h
hose
o
UV
adia ion.
MATERIALS
AND
METHODS
S ain,
media,
and
cul u e
condi ions.
G.
ujiku oi
IM158289
was
ob ained
om
he
Commonweal h
Mycological
Ins i-
u e,
Kew,
Su ey,
Englapd.
Minimal
medium
con ained
(pe
li e )
glucose,
30
g;
NaNO3,
3
g;
KH2PO4,
1
g;
MgSO4
*
7H20,
0.5
g;
KCl,
0.5
g;
HBO4,
10
,ug;
CUSO4,
100
,ug;
FeC13,
200
,ug;
MnCl2,
20
,ug;
MoO4Na,
20
Rxg;
and
ZnSO4,
2
mg.
Nu ien
b o h
was
p epa ed
by
adding
4
g
o
yeas
ex ac
(Di co
Labo a o ies,
De oi ,
Mich.)
and
8
g
o
pep one
(Bac o-T yp one;
Di co)
o
he
minimal
medium.
Solid
medium
con ained
addi ion-
*
Co esponding
au ho .
P esen
add ess:
Depa men
o
Biology,
Uni e si y
o
U ah,
Sal
Lake
Ci y,
UT
84112.
ally
16
g
o
aga
(Analema,
Vo quimica,
Vigo,
Spain).
Cul u es
we e
incuba ed
a
30°C
in
he
da k.
To
ha es
mic oconidia,
cul u es
we e
g own
on
a
special,
ca bon-lim-
i ed
medium
con aining
(pe
li e )
yeas
ex ac
(Di co),
1
g;
NH4NO3,
1
g;
KH2PO4,
1
g;
MgSO4
*
7H20,
0.5
g;
and
aga ,
16
g.
The
pla es
we e
g own
o
5
days
a
30°C
10
cm
away
om
a
ba e y
o
40-W
luo escen
lamps
(Syl ania
Li eline
Dayligh
F40
V/D),
yielding ca.
1.25
W
o
whi e
ligh
m
Mic oconidia
we e
ha es ed
by
washing
he
spo ula ed
mycelia
wi h
s e ile
dis illed
wa e
and
cleaned
by
passage
h ough
il e
pape
and
low-speed
cen i uga ion.
Conidial
suspensions
we e
s o ed
a
4°C
and
i a ed
by
mic oscopic
obse a ion
wi h
a
Pe o -Hause
chambe .
Viable
coun s
we e
de i ed
om
colony- o ming
abili y
on
nu ien
aga .
Fo
nuclea
s aining,
mi h amycin
(a
kind
gi
o
P ize ,
Inc.,
G o on,
Conn.)
was
dissol ed
a
10
,ug
ml-'
in
a
mix u e
o
equal
olumes
o
dime hyl
sul oxide
and
wa e .
Ni osoguanidine
mu agenesis.
Ni osoguanidine
(Sigma
Chemical
Co.,
S .
Louis,
Mo.)
was
dissol ed
in
dis illed
wa e
a
1
mg
ml-',
kep
ozen
un il
use,
and
ne e
e ozen.
The
sa e y
ules
p oposed
by
Eh enbe g
and
Wach meis e
(9)
we e
adop ed.
F eshly
collec ed
suspensions
o
ca.
106
conidia
pe
ml
o
dis illed
wa e
we e
exposed
o
he
d ug.
Se e al
doses
o
ni osoguanidine
we e
used.
We
de ine
he
dose
as
he
p oduc
o
he
mola
concen a ion
o
he
d ug
and
he
exposu e
ime
in
seconds.
Any
dose
can
he e o e
be
ob-
ained
by
using
di e en
combina ions
o
concen a ion
and
exposu e.
T ea men s
we e
s opped
by
dilu ing
and
wash-
ing.
UV
mu agenesis.
F eshly
collec ed
mic oconidia
we e
in-
ocula ed
on o
nu ien -aga
pla es,
placed
49
cm
away
om
a
Syl ania
G15T8
lamp
(UV
lux,
0.35
W
m-2),
and
hen
incuba ed
in
he
da k.
Fo
pho o eac i a ion,
UV
exposu e
was
ollowed
by
a
5-h
incuba ion
in
he
ligh ,
as
desc ibed
abo e,
be o e
incuba ion
in
he
da k.
Cha ac e iza ion
o
mu an s.
G ow h
equi emen s
o
auxo-
ophic
mu an s
we e
de e mined
ollowing
he
me hod
o
Holliday
(14).
Ca o enoids
we e
ex ac ed
wi h
ace one
om
lyophilized,
weighed,
and
g ound
mycelial
samples
and
de e mined
wi h
a
Bausch
&
Lomb
Spec onic
2000
eco d-
ing
spec opho ome e .
187
on Feb ua y 23, 2016 by USE/BCTA.GEN UNIVERSITARIAh p://aem.asm.o g/Downloaded om on Feb ua y 23, 2016 by USE/BCTA.GEN UNIVERSITARIAh p://aem.asm.o g/Downloaded om on Feb ua y 23, 2016 by USE/BCTA.GEN UNIVERSITARIAh p://aem.asm.o g/Downloaded om
188
AVALOS,
CASADESUS,
AND
CERDA-OLMEDO
'I
FIG.
1.
Spo es
o
G.
ujiku oi
IM158289
s ained
wi h
mi h-
amycin
and
pho og aphed
wi h
a
luo oscence
mic oscope.
Ba ,
10
,um.
RESULTS
Mic oconidia.
Ou
Gibbe ella
s ain
g ows
igo ously
bu
conidia es
poo ly
unde
usual
labo a o y
condi ions.
Less
han
103
conidia
pe
pla e
we e
ound
a e
5
days
o
g ow h
in
he
da k
o
on
nu ien
medium
in
he
ligh .
Simila
cul u es
g own
on
minimal
aga
in
he
ligh
p oduced
ca.
2.4
x
104
conidia
pe
pla e.
Abundan
spo es
(5.1
x
106
pe
pla e)
we e
p oduced
in
cul u es
g own
o
5
days
on
an
DOSE
(
M
x
s)
FIG.
2.
Su i al
o
Gibbe ella
spo es
exposed
o
ni osoguani-
dine.
Open
symbols,
es ing
spo es
(kep
in
wa e );
closed
symbols,
spo es
p eincuba ed
o
2
h
in
nu ien
b o h.
The
doses
indica ed
in
he
abscissa
we e
ob ained
by
exposu es
o
15
( iangles),
30
(squa es),
45
(ci cles),
60
(diamonds),
o
75
(s a s)
min
o
he
app op ia e
ni osoguanidine
concen a ions.
o
I
I
I
I
I
I
I
I
I
*1I
.01
1
10
8
16
GERMINATION
TIME
(
h )
FIG.
3.
E ec
o
ge mina ion
on
he
su i al
o
Gibbe ella
spo es
mu agenized
wi h
ni osoguanidine.
The
spo es
we e
p ein-
cuba ed
o
a ious
pe iods
in
nu ien
b o h
and
hen
exposed
o
ni osoguanidine
a
0.234
M
*
s
(100
,ug
ml-'
o
30
min;
iangles)
o
0.351
M
-
s
(100
,ug
ml-'
o
45
min;
ci cles).
especially
poo
aga
(conidia ion
aga )
unde
he
same
b igh
illumina ion.
The
spo es
a e
mic oconidia,
ha
is,
unicellula ,
ellipsoid-
al
s uc u es,
4
o
6
pum
long
and
2
o
3
,um
wide,
each
con aining
a
single
nucleus
(Fig.
1).
Ni osoguanidine
le hali y
and
mu agenesis.
The
conidia
a e
qui e
sensi i e
o
he
le hal
e ec s
o
ni osoguanidine.
Su i al
and
mu agenesis
depended
only
on
he
dose
o
ni osoguanidine
and
no
on
he
pa icula
concen a ions
and
imes
used
in
each
expe imen
(Fig.
2).
Ni osoguani-
dine
le hali y
was
enhanced
by
p eincuba ion
o
he
mic o-
conidia
in
nu ien
b o h,
i.e.,
by
he
onse
o
ge mina ion
(Fig.
2
and
3).
The
highes
sensi i i y
was
eached
a e
a
2-h
p eincuba ion,
be o e
he
appea ance
o
he
i s
g oss
mo phological
changes
and
was
main ained
h ough
he
p o usion
o
ge m
ubes.
A e
6
h
o
p eincuba ion,
80%
o
he
spo es
exhibi ed
ge m
ubes;
a e
12
h,
99%
exhibi ed
ge m
ubes.
Mu agenesis,
as
judged
by
he
p opo ion
o
auxo ophs
among
he
su i o s,
inc eased
linea ly
wi h
he
dose.
The
numbe
o
iable
mu an s
induced
by
ni osoguanidine
in
a
gi en
ini ial
cell
popula ion
eached
a
maximum
a
a
dose
o
abou
0.07
M
*
s
(Fig.
4).
Res ing
spo es
su e ed
ewe
dea hs
and
mu a ions
han
p eincuba ed
spo es,
i
he
com-
pa ison
was
made
a
he
same
dose.
I
bo h
kinds
o
spo es
we e
killed
o
he
same
su i al
le el,
hey
showed
he
same
mu a ion
equencies
(Table
1).
UV
le hali y
and
mu agenesis.
The
Gibbe ella
spo es
we e
eadily
killed
by
UV
i adia ion
a
a
wa eleng h
o
254
nm.
The
le hal
e ec
o
UV
i adia ion
was
e e sed
by
immedia e
exposu e
o
whi e
ligh
(Fig.
5).
The
le hali y
app oxima ely
ollowed
i s
o de
kine ics
wi h
a
small
shoulde .
APPL.
ENVIRON.
MICROBIOL.
on Feb ua y 23, 2016 by USE/BCTA.GEN UNIVERSITARIAh p://aem.asm.o g/Downloaded om
GIBBERELLA
MUTANTS
0-%
C,,
:No
I--
0
X
C
-4
CD
a
--4
-4
C-,
'm
I-e
%,,
<
.1
cn,
.01
DOSE
(M
*
s)
FIG.
4.
Pe cen age
o
auxo ophs
among
he
su i o s
o
ni oso-
guanidine
exposu e
(0).
The
spo es
we e
p eincuba ed
o
2
h
in
nu ien
b o h,
exposed
o
ni osoguanidine,
and
pla ed
on
nu ien
aga ;
he
esul ing
colonies
we e
es ed
o
auxo ophy.
The
same
da a
a e
eplo ed
as
he
pe cen age
o
iable
auxo ophs
ela i e
o
he
o al
numbe
o
cells,
li ing
o
dead
(A).
UV
le hali y
was
no
a ec ed
by
ge mina ion
up
o
he
massi e
p oduc ion
o
ge m
ubes;
mycelia
we e
mo e
esis an
o
UV
i adia ion
han
we e
conidia
(Fig.
6).
UV
exposu e
was
mu agenic,
bu
less
so
han
was
ni oso-
guanidine
ea men ,
leading
o
compa able
su i al
le els
(Table
1).
Gibbe ella
mu an s.
The
su i o s
o
ni osoguanidine
ea men
we e
no
always
iden ical
o
each
o he
and
o
he
wild
ype.
Colony
size,
su ace
ex u e,
colo ,
and
o e all
mo phology
we e
o en
abe an .
The
same
was
ue,
o
a
lesse
ex en ,
o
he
su i o s
o
UV
i adia ion.
Ni osoguanidine-induced
auxo ophs
showed
a
wide
a-
ie y
o
nu i ional
equi emen s:
we
iden i ied
se en
a gi-
nine,
i e
me hionine,
ou
leucine,
h ee
lysine,
wo
bio ine,
wo
nico inic
acid,
one
yp ophan,
and
one
phenylalanine
auxo oph.
We
s udied
wo
pink
mu an s,
which
con ained
he
same
ca o enoids
as
he
wild
ype
and
an
addi ional
wa e -soluble
TABLE
1.
Mu agenic
e ec s
o
ni osoguanidine
and
UV
adia ion
on
G.
ujiku oi"
Auxo ophs
P eincuba ion
Ax oh
P icb ion
Su i al
among
he
Mu agen
Dose
in
nu ien
b o h
(h)
)
su i o s
Ni oso-
0.5
M
*
s
0
15
2.5
guanidine
0.1
M
*
s
2
14
3.1
0.05
M
*
s2
33
1.4
UV
adia ion
0.7
J
m-2
0
20
0.8
A
o al
o
1.040
colonies
we e
es ed
o
auxo ophy
in
each
case.
The
auxo oph
equency
o
UV
adia ion
is
signi ican ly
lowe
han
hose
o
ni osoguanidine,
as
judged
by
he
es
a e
he
a csin
ans o ma ion
(23).
Fo
example.
i
he
mu agenici ies
a e
assumed
o
be
equal.
he
p obabili y
o
a
di e ence
as
la ge
as
o
la ge
han
ha
be ween
he
lowe
wo
lines
is
P
<
0.04.
0
1
2
3
DOSE
(J*m-2)
FIG.
5.
Killing
o
Gibbe ella
spo es
by
UV
adia ion
and
pho o-
eac i a ion
by
isible
ligh .
A e
exposu e
o
he
dose
gi en
in
he
abscissa,
samples
we e
exposed
(0)
o
no
(0)
o
isible
ligh
and
hen
incuba ed
in
he
da k.
pink
pigmen .
These
mu an s
we e
auxo ophic
o
adenine.
The
syn hesis
o
he
pink
pigmen
was
ep essed
by
adenine:
mycelia
g own
wi h
20
mg
o
adenine
li e -'
in
he
da k
we e
deeply
colo ed;
wi h
200
mg
li e -',
a he
aded;
and
wi h
2
g
li e -',
whi e.
All
o
hese
ea u es
a e
ypical
o
some
adenine
auxo ophs
in
o he
ungi,
such
as
he
mu an s
o
Cn.1
.02
000.0
I
I
0
0
0
0
0
~~~~~0
0
0
1
1
1
8
GERMINATION
TIME
(h )
16
FIG.
6.
E ec
o
ge mina ion
on
he
su i al
o
Gibbe ella
spo es
exposed
o
UV
adia ion.
The
spo es
we e
p eincuba ed
in
nu ien
b o h
o
he
pe iod
gi en
in
he
abscissa,
exposed
o
1
J
m-2
UV
adia ion,
exposed
(0)
o
no
(0)
o
isible
ligh ,
and
incuba ed
in
he
da k.
0
0
0
I
I
I
I
I
I
i
VOL.
49,
1985
189
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190
AVALOS,
CASADESOS,
AND
CERDA-OLMEDO
genes
ad]
and
ad2
in
S.
ce e isiae
(22)
and
adel
and
ade2
in
Candida
albicans
(19).
O he
mu an s
we e
pu ple
when
g own
in
ei he
he
ligh
o
he
da k.
They
we e
pho o ophic,
and
hei
ca o enoid
con en
coincided
wi h
ha
o
he
wild
ype.
The
na u e
o
he
pigmen
emains
unknown.
The
es
o
ou
colo
mu an s
di e ed
om
he
wild
ype
in
neu ospo axan hin
con en .
The
wild
ype
was
o ange
when
g own
in
he
ligh ,
con aining
100
o
150
,ug
o
neu ospo a-
xan hin
g-'
(d y
weigh )
(ppm).
In he
da k,
he
syn hesis
o
neu ospo axan hin
was
ep essed,
and
he
mycelium
was
whi e
(less
han
10
ppm
o
neu ospo axan hin).
The
ca o -
enoid
mu an s
can
be
classi ied
in o
h ee
pheno ypic
classes.
One
o
hem
is
made
up
o
mu an s
wi h
li le
o
no
pigmen
in
ei he
he
ligh
o
he
da k.
Ano he
class
is
indis inguish-
able
om
he
wild
ype
in
he
ligh ,
bu
is
o ange
in
he
da k,
due
o
accumula ion
o
30
o
100
ppm
o
neu ospo axan hin.
The
emaining
ca o enoid
mu an s
a e
deep
o ange
when
g own
in
ei he
he
ligh
o
he
da k,
and
hei
ca o enoid
con en
inc eases
linea ly
wi h
ime,
up
o
ca.
2,000
ppm
o
neu ospo axan hin.
DISCUSSION
G.
ujiku oi
is
a
highly
sui able
o ganism
o
he
induc ion
and
isola ion
o
mu an s.
The
uninuclea e
mic oconidia
eadily
allow
he
exp ession
o
ecessi e
mu a ions.
To
app ecia e
he
con enience
o
Gibbe ella
mic oconidia,
con-
side
he
di icul ies
encoun e ed
in
he
isola ion
o
mu an s
in
mul inuclea e
cells,
such
as
Phycomyces
spo es
(21).
No
all
s ains
o
G.
ujiku oi
a e
equally
a o o able
o
hese
s udies,
because
many
s ains
p oduce
mul icellula
mac o-
conidia
(16).
Ni osoguanidine
and
UV
adia ion
kill
Gibbe ella
mic o-
conidia
abou
as
e ec i ely
as
hey
kill
he
mic oo ganisms
mos
o en
used
in
gene ics.
The
onse
o
ge mina ion
inc eases
he
sensi i i y
o
mic oconidia
o
ni osoguanidine.
In
a
simila
way,
ac i ely
eplica ing
bac e ia
and
yeas
and
ge mina ing
Phycomyces
spo es
a e
mo e
eadily
killed
by
ni osoguanidine
han
he
co esponding
es ing
cells
(3,
4,
21).
Inc eased
cell
pe meabili y
and
changes
in
nuclea
s uc u e
and
unc ion
a e
p esumably
esponsible
o
his
di e ence.
Pe meabili y
changes
may
be
he
mos
impo an ,
since
UV
adia ion
makes
no
dis inc ion
be ween
es ing
and
ge mina ing
conidia.
The
mycelia,
a e
he
p o usion
o
he
ge m
ubes,
a e
much
mo e
esis an
o
bo h
le hal
agen s
han
a e
spo es.
G.
ujiku oi
shows
a
mos
ac i e
pho o eac-
i a ion
o
UV
damage,
compa able
o
ha
o
Phycomyces
(6)
and
Saccha omyces
spp.
(17).
As
wi h
o he
o ganisms,
ni osoguanidine
is
a
mo e
e ec i e
mu agen
han
UV
adia ion
o
G.
ujiku oi.
The
auxo oph
equencies
epo ed
he e
a e
among
he
highes
epo ed
o
any
o ganisms.
Auxo oph
equencies
depend
o
a
ce ain
ex en
on
subjec i e
assessmen ,
depending
on
he
c i e ia
o
inclusion
o
leaky
mu an s.
E en
so,
ou
equencies
a e
much
highe
han
hose
usual
o
Phycomyces
spp.
(less
han
0.1%)
o
yeas
cells
(ca.
1%)
unde
simila
condi ions.
The
ecommended
ni osoguanidine
dose
o
p ac ical
applica ions
depends
on
he
na u e
o
he
desi ed
mu an s.
Fo
a e,
nonselec able
mu an s,
he
dose
should
be
high,
o
imp o e
he
chances
o
success
a
he
isk
o
accumula ing
mul iple
mu a ions
in
he
same
genome.
Fo
selec able
mu an s,
pa icula ly
hose
in ended
o
physiological
s ud-
ies,
he
dose
should
be
as
low
as
possible,
o
a oid
mul iple
mu a ions.
The
s a us
o
Gibbe ella
gene ics
does
no
ye
allow
a
di ec
es
o
whe he a
pheno ype
is
due
o
one
o
mo e
mu a ions.
Ni osoguanidine
mu agenesis
in
Esche ichia
coli
quickly
eaches
sa u a ion:
u he
exposu e
inc eases
le hali y
bu
no
he
equency
o
mu an s
(4).
In
G.
ujiku oi,
as
in
Phycomyces
spp.
(21),
he
equency
o
mu an s
inc eases
linea ly
wi h
he
dose.
This
sugges s
a
undamen al
di e -
ence
in
he
way
ni osoguanidine
ac s
in
he
wo
ungi
and
he
bac e ium.
The
whi e
and
deep-colo ed
Gibbe ella
mu an s
esemble
simila
mu an s
in
Phycomyces
spp.
(7).
Mos
a ac i e
a e
he
mu an s
ha
syn hesize
neu ospo axa hin
in
he
da k
bu
ha
a e
simila
o
he
wild
ype
in
he
ligh .
These
no el
mu an s
should
con ibu e
o
he
s udies
o
pho oinduc ion
o
ca o enoids,
qui e
de eloped
in
a
ela ed
ungus,
Fusa -
ium
aquaeduc uum
(20).
Ou
esul s
encou age
he
sea ch
o
all
kinds
o
Gibbe -
ella
mu an s,
including
hose
o
po en ial
indus ial
in e es .
ACKNOWLEDGMENTS
We
hank
A.
Fe nandez
Es e ane
and
D.
Sud ez
o
hei
assis -
ance
and
he
Comisi6n
Aseso a
pa a
In es igaci6n
Cien i ica
y
Tdcnica
o
inancial
suppo .
LITERATURE
CITED
1.
Bea de ,
J.
R.,
J.
MacMillan,
C.
M.
Wels,
M.
B.
Cha ey,
and
B.
0.
Phinney.
1974.
Posi ion
o
he
me abolic
block
o
gibbe -
ellin
biosn hesis
in
mu an
B1-41a
o
Gibbe ella ujiku oi.
Phy o-
chemis y
13:911-917.
2.
Calde 6n,
I.
L.,
and
E.
Ce da-Olmedo.
1982.
Simul aneous
induc ion
o
mul iple
mu a ions
by
N-me hyl-N'-ni o-N-
ni osoguanidine
in
he
yeas
Saccha omyces
ce e isiae.
Cu .
Gene .
6:237-243.
3.
Calde 6n,
I.
L.,
and
E.
Ce da-Olmedo.
1983.
Induc ion
by
N-me hyl-N'-ni o-N-ni osoguanidine
o
nuclea
and
cy oplas-
mic
mu a ions
in
Sacha omyces
ce e isiae.
Mu a .
Res.
108:133-146.
4.
Ce dk-Olmedo,
E.,
and
P.
C.
Hanawal .
1968.
The
eplica ion
o
he
Esche ichia
coli
ch omosome
s udied
by
sequen ial
ni oso-
guanidine
mu agenesis.
Cold
Sp ing
Ha bo
Symp.
Quan .
Biol.
33:599-607.
5.
Ce a-Olmedo,
E.,
P.
C.
Hanawal ,
and
N.
Gue ola.
1968.
Mu agenesis
o
he
eplica ion
poin
by
ni osoguanidine:
map
and
pa e n
o
eplica ion
o
he
Esche ichia
coli
ch omosome.
J.
Mol.
Biol.
33:705-719.
6.
Ce da-Olmedo,
E.,
and
P.
Reau.
1970.
Gene ic
classi ica ion
o
he
le hal
e ec s
o
a ious
agen s
o
he e oka yo ic
spo es
o
Phycomyces.
Mu a .
Res.
9:369-384.
7.
Ce da-Olmedo,
and
E.
S.
To es-Ma inez.
1979.
Gene ics
and
egula ion
o
ca o ene
biosyn hesis.
Pu e
Appl.
Chem.
51:631-637.
8.
Dawes,
I.
W.,
and
L.
A.
Ca e .
1974.
Ni osoguanidine
mu a-
genesis
du ing
nuclea
and
mi ochond ial
gene
eplica ion.
Na u e
(London)
250:709-712.
9.
Eh embe g,
L.,
and
C.
A.
Wach meis e .
1977.
Sa e y
p ecau-
ions
in
wo k
wi h
mu agenic
and
ca cinogenic
chemicals,
p.
401-410.
In
B.
J.
Kilbey,
M.
S.
Lega o ,
W.
Nichols,
and
C.
Ramel
(ed.),
Handbook
o
mu agenici y
es
p ocedu es.
Else ie /No h-Holland
Biomedical
P ess,
Ams e dam.
10.
E okhina,
L.
I.
1969.
Ce ain
cha ac e is ics
o
Fusa ium
mo-
nili o me
mu an s.
Gene ika
5:143-147.
11.
E okhina
L.
I.,
and
B.
D.
E emo .
1970.
Biochemical
mu an s
o
Fusa ium
monili o me
(Sheld.).
Gene ika
6:170-172.
12.
Gichne ,
T.,
and
J.
Veleminsky.
1982.
Gene ic
e ec s
on
N-
me hyl-N'-ni o-N-ni osoguanidine
and
i s
homologs.
Mu a .
Res.
99:129-242.
13.
Gue ola,
N.,
J.
L.
Ing aham,
and
E.
Ce d.i-Olmedo.
1971.
Induc ion
o
closely
linked
mul iple
mu a ions
by
ni osoguan-
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Holliday,
R.
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new
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iden i ica ion
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Je eys,
E.
G.
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Kuhlman,
E.
G.
1983.
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o
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ujiku oi
wi h
anamo phs
in
Fusa ium
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Pa y,
J.
M.,
and
B.
S.
Cox.
1968.
The
e ec s
o
da k
holding
and
pho o eac i a ion
on
ul a iole
ligh -induced
mi o ic
ecom-
bina ion
and
su i al
in
yeas .
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Res.
12:187-198.
18.
Phinney,
B.
O.,
and
C.
Spec o .
1967.
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Poul e ,
R.
T.
M.,
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E.
H.
A.
Rikke ink.
1983.
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analysis
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albicans.
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Bac e iol.
156:1066-1077.
20.
Rau,
W.
1980.
Blue
ligh -induced
ca o enoid
biosyn hesis
in
mic oo ganisms,
p.
283-298.
In
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(ed.),
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synd ome.
Sp inge -Ve lag,
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Ronce o,
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1983.
Mu agenesis
in
mul inuclea e
cells:
he
e ec s
o
N-me hyl-N'-
ni o-N-ni osoguanidine
on
Ph co n ces
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Mu a .
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22.
Sil e ,
J.
M.,
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N.
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Ea on.
1969.
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blocks
o
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adl
and ad2
mu an s
o
Saccha omi ces
ce e 'isiae.
Biochem.
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34:301-305.
23.
Sokal,
R.
R.,
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F.
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1969.
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24.
Spec o ,
C.,
and
B.
0.
Phinney.
1966.
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Science
153:1397-1398.
VOL.
49,
1985
on Feb ua y 23, 2016 by USE/BCTA.GEN UNIVERSITARIAh p://aem.asm.o g/Downloaded om

ERRATUM
Gibbe ella
ujiku oi
Mu an s
Ob ained
wi h
UV
Radia ion
and
N-Me hyl-N'
-Ni o-N-Ni osoguanidine
J.
AVALOS,
J.
CASADESUS,
AND
E.
CERDA-6LMEDO
Depa amen o
de
Gene ica,
Facul ad
de
Biologia,
Uni e sidad
de
Se illa,
Se illa,
Spain
Volume
49,
no.
1,
p.
187,
abs ac ,
line
5:
".
.
.
eaches
a
maximum
o
a
dose
o
ca.
0.7
M
s"
should
ead
".
.
.
eaches
a
maximum
o
a
dose
o
ca.
0.1
M.
s."
Page
187,
abs ac ,
line
9:
"Pink
adenine
auxo ophs
and
se e al
classes
o
colo
mu an s
a e
a ec ed
.
.
."
should
ead
"Some
adenine
auxo ophs
a e
pink,
and
se e al
classes
o
colo
mu an s
a e
a ec ed...."
Page
187,
column
2,
lines
2-6:
An
ing edien
should
be
added
o
he
desc ip ion
o
he
ca bon-limi ed
medium:
"glucose,
1
g."
735