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| Fermentation
New Brunswick Scientifi c
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Edison, NJ 08818-4005 USA
New Brunswick Scientific 877-723-3318 (North America)
732-287-1200 (Rest of World)
bioinfo@nbsc.com with cc to suzyk@nbsc.com
www.nbsc.com/btaa.htm
Basics of Benchtop Fermentor Operation for Growth of E. coli
Introduction
Basics of Benchtop Fer
Medium
mentor Operation
While use of shake fl asks, T-fl asks, or roller bottles
for
ha
G
ve become
row
the
lab standard in most fermentation and cell culture research labs, lack
th o
The initial fermentor medium composition inc
f E. coli
luded:
Potassium phosphate monobasic, KH2PO4 3.5 g/L
of controls for pH, dissolved oxygen and other parameters cause an Potassium phosphate dibasic, K2HPO4 5.0 g/L
inherent limitation in cell yields and protein expression. When higher Ammonium Sulfate, (NH4)2SO4 5.0 g/L
yields are needed, rather than inoculate, feed and maintain cultures Yeast Extract
using dozens or even hundreds of bottles or fl asks, a benchtop (note that we use Tastone 900AG from Sensient) 5.0 g/L
I
fermentor
ntrodu
or
ct
cell-culture
ion
bioreactor can be used to increase yields Antifoam, (Breox Foam Control Agent FMT 30) 1.00 ml/L
by 10-fold or more.
While use of shake flasks, T-flasks, or roller bottles
h
This
ave
A
b
pplica
ecom
tion
e t
Report
he lab
describes
standar
a
d
step-by-step
in most fer
procedure
mentatio
for
n
a
gro
nd
wing
cel
E.
l
coli
cul
in
tu
New
re r
Bruns
esear
wick
ch
Scientifi
labs, l
c’
ac
s
k
BioFlo
®
of c

o
310
ntr
benchtop
fermentor. The BioFlo 310 is a versatile, powerful system, designed
ols for
p
for
H,
high-yield
dissolve
gro
d
wth
oxy
of
ge
a
n
wide
and
variety
other
of
pa
organisms,
rameter

s
ranging
cause
from
an
i
yeast
nher
and
ent
bacteria
limita
to
tio
mammalian,
n in cell

y
plant,
ield
and
s an
insect
d pr
cells.
otei

n
The
ex
system
pres-
s
is
io
a
n
vailable
. Wh
with
en
interchangeable
higher yields
vessels,
are n
sized
eede
2.2–14.0
d, rathe
liters,
r th
and
an
icomes noculas ata e,pre-packafeed anged d mkit afor inordering tain cusimplicityltures u, or sincan g dbe oztailorens-
o
confi
re

v
gured by selecting from dozens of standard options.
enhundredsofbottlesorflasks,abenchtopfer-
m

entor or cell-culture bioreactor can be used to
i
Materials and Methods
ncrease yields by 10-fold or more.
The Fermentor
T
A
h
pre-confi
is App
gured
licati
BioFlo
on R
310
epo
F
r
ermentor
t descr
Kit
ibe
with
s a
7.5
ste
L
p
vessel
-by-s
with
Rushton blades (NBS Catalog Number M1287-1172) was used for
te
an
p
p
E.
r

o
coli
ce
fed-ba
dure
tch
fo
fermenta
r grow
tion.
ing
This
E.c
system
oli in
inc
N
ludes
ew
a
B
15”
ru
industrial
nswick
Scolor cientouchscreen tific’s Biocontroller Flo
®
31to 0regulabente chatgitaoption,fe rtemperamentoture,r. T hpH,e
B
DO,
io

F
F
l
oam/Level,
o 310 is
three
a ver
pumps,
satile,
gases
powe
and
rful
much
syste
more.
m, d
A
es
Thermal
igned
f
Mass
or hi
Flo
gh
w
-y
Controller
ield grow
(TMFC)
th of
for
a w
regula
ide
ting
vari
gas
ety

o
ow
f o
in
rg
a
a
range
nisms
of
,
r
0.4–20
anging
Standard
from
Liters
yeast
per
an
Minute
d ba
(SLPM)
cteria
is
to
also
m
standard,
ammali

a
and
n,
p
options
lant, a
for
nd
multiple
insect
mass
cells

.
o
T
w
h
controllers
e system
and/or
is ava
other
ilabl
gas
e w

i
o
th
w
settings are available as standard options. In this run, an optional
i
BioCommand
nterchange®a
Plus
ble v
super
esse
visor
ls, s
y
iz
software
ed 2.2
packa
– 14
ge
.0
(NBS
Liters
Ca
, a
talog
nd
cNumber omes aM1291-0000) s a pre-pacwas kagused ed kto itcontrol for orthe derfeed ingschedule simpliciand ty,
The benchtop BioFlo 310 fermentor is a versatile research
o
log
r c
da
an
ta.
b
To
e
optimize
tailor-c
your
onfi
results,
gured
you
by
can
sele
also
ctin
connect
g from
and
do
control
zens
toolforoptimizingcellgrowthandproductionfromawide
o
numerous
f standa
additional
rd option
options
s.
directly to the BioFlo 310, including
varietyofcelltypes,includingmicrobes,yeastandfungi,as
scales, analyzers and sensors for optical density (OD), CO2, Redox
well as mammalian, insect and plant cell lines.
M
and
a
more.
teria
This
ls a
pa
n
per
d M
presents
etho
a
d
fermenta
s
tion run that was not fully
optimized to obtain the highest yields possible–rather it is meant to
T
ser
he
ve as a guide for basic procedures and ma
Fermentor
terials.
AInoculumpre-configured BioFlo 310 Fermentor Kit with 7.5L M1291-0000), was used to control the feed schedule and
v
The
ess
inoculum
elwithR
was
ush
prepared
tonblad
using
es(N
LB
BS
broth
Cata
a
l
t
o
a
g
25-g/L
Numb
concentra
erM128
tion
7- log data. To optimize your results, you can also connect
1
as
17
the
2)
shake
was u

s
ask
ed f
medium.
or an E
The
.coli
inoculum
fed-batc
was
h fe
cultiva
rment
ted
atio
overnight
n. This and control numerous additional options directly to the
s
on
yst
an
em
NBS
inc
orbital
ludes
shaker
a 15”
a
i
t
n
240
dust
rpm.
rial
OD600
color t
was
ouch
between
screen
2
co
and
n-
4 at the time of inoculation. Inoculum volume was 2-5% of the 5 L
BioFlo 310, including scales, analyzers and sensors for
t
working volume,
roller to reg
or 250 ml.
ulate ag

itation, temperature, pH, DO, optical density (OD), CO2, Redox and more. This paper
Foam/Level, three pumps, gases and much more. A presentsafermentationrunthatwasnotfullyoptimizedto
Th
Protocol
ermal
Guide
Mas

s
ı 200
Flo
8
w

C

o

n

t

r

o

l

l

e

r

(

T

M

F

C

)

f

o

r

r

e

g

u

l

a

t

i

n
V
g
iew
ga
entire
s
protocol
obtain
online
the
at
hi
www
ghe
.biotechniques.com/protocol
st yields possible – rat
ı
h
BioT
er i
echniques
t is mea
ı
n
29
t to
flow in a range of 0.4 – 20 Standard Liters per Minute serve as a guide for basic procedures and materials.
(SLPM)isalsostandard,andoptionsformultiplemassflow
controllers and/or other gas flow settings are available as
NewBrunswick Protocol.indd 29 10/26/07 11:20:33 AM
standard options. In this run, an optional BioCommand
®
Plus supervisory software package, (NBS Catalog Number
New Brunswick Scientific
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Edison, NJ USA
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