Document ID: EPA-HQ-OPP-2006-0349-0002
Agency: epa
Document Type: Supporting & Related Material
Title: 
Posted Date: 2006-07-19T04:00Z

UNITED
STATES
ENVIRONMENTAL
PROTECTION
AGENCY
WASHINGTON,
D.
C.
20460
OFFICE
OF
PREVENTION,
PESTICIDES
AND
TOXIC
SUBSTANCES
DEC
05
2005
MEMORANDUM
SUBJECT:
Review
of
materials
submitted
by
Bayer
Crop
Sciences
in
support
of
a
proposed
Section
G
Experimental
Use
Permit
(
EUP
No:
264RUN,
DP#
315394)
for
Cry1Ab
cotton.
Submission
dated
January
17,
2005.

FROM:
Tessa
Milofsky,
M.
S.
Agronomist
Microbial
Pesticides
Branch,
Biopesticides
and
Pollution
Prevention
Division
(
7511C)

TO:
Sharlene
Matten
Biologist
Microbial
Pesticides
Branch,
Biopesticides
and
Pollution
Prevention
Division
(
7511C)

CONCLUSION:
The
requested
EUP
and
corresponding
acreage
are
ACCEPTABLE.
It
is
recommended
that
the
registrant's
full
request
of
370
EUP
acres
should
be
granted.

COMMENTS/
RECOMMENDATIONS:
 
Containment
procedures
described
in
the
submissions
(
data
package
dated
6/
13/
05
and
letter
dated
10/
3/
05)
and
corresponding
emails
(
dated
9/
7/
05
and
12/
5/
05)
are
adequate
for
field
sites
located
in
the
continental
United
States.

 
Cry1Ab
cotton
EUP
plots
located
in
Puerto
Rico
will
be
covered
with
netting/
cages
during
anthesis.
This
is
an
acceptable
alternative
to
the
standard
3
mile
isolation
distance
requirement
for
all
Bt
cotton
EUP
acreage
in
Puerto
Rico.

BACKGROUND:
Bayer
Crop
Sciences
has
submitted
materials
in
support
of
their
request
for
an
Experimental
Use
Permit
(
EUP),
covering
the
period
February
2006
through
February
2007,
to
allow
field
testing
of
Cry1Ab
cotton
(
Event
T303­
3
and
T304­
40
derived
cotton
plants).
The
product
is
said
to
protect
cotton
against
lepidopteran
larvae
such
as
bollworm
(
CBW,
Helicoverpa
zea)
and
tobacco
budworm
(
TBW,
Heliothis
virenscens),
which
are
common
pests
of
cotton.
Planting
proposed
under
the
EUP
will
take
place
in
6
states
and
Puerto
Rico,
and
will
not
exceed
370
total
acres.

PROGRAM
OVERVIEW
Participant/
Collaborator
Information
2
The
names,
telephone
numbers,
and
mailing
addresses
of
cooperators
and
participants
who
will
supervise
experimental
work
covered
under
this
EUP
are
listed
in
the
confidential
appendix.

BPPD
Review
Participant/
cooperator
information
is
sufficient
provided
that
the
names
and
contact
information
of
any
additional
collaborators
are
submitted
prior
to
experiment
initiation.

Field
Trial
Containment
The
proposed
field
trial
containment
program
outlined
in
Bayer's
June
13,
2005
submission
to
EPA
complies
with
the
USDA­
APHIS
Performance
Standards
for
regulated
cotton
trials.
The
USDA's
guidelines
are
general,
leaving
room
for
registrants
to
develop
containment
programs
that
are
appropriate
for
each
management
system.

Bayer's
October
3,
2005
letter
to
EPA
provided
greater
detail
on
the
proposed
containment
program
for
Cry1Ab
cotton.
In
this
document,
the
registrant
committed
to
implementing
at
least
one
containment
measure
for
Bt
EUP
cotton
trials
sown
in
the
continental
United
States.
Further,
to
address
EPA's
concerns
regarding
EUP
cotton
containment
in
Puerto
Rico,
the
registrant
agreed
to
surround
all
EUP
plots
with
24
non­
Bt
cotton
border
rows
and
in
place
of
EPA's
3
mile
isolation
distance
requirement
for
Puerto
Rico,
the
registrant
proposed
to
cover
all
plots
with
netting
or
cages
during
anthesis.

BPPD
Review
Containment
measures
described
in
the
June
13,
2005
and
October
3,
2005
submissions
are
adequate
for
field
sites
located
in
the
continental
United
States.
After
receiving
the
initial
submission,
it
was
recommended
to
the
registrant
that
at
least
one
of
the
listed
containment
measures
should
be
implemented
for
each
experiment.
In
their
September
7,
2005
email
message
and
October
3,
2005
letter
to
EPA,
Bayer
agreed
to
implement
at
least
one
containment
measure
for
each
experiment.

Experimental
field
sites
located
in
Hawaii,
Puerto
Rico,
the
US
Virgin
Islands,
and
southern
Florida
(
south
of
Route
60)
require
a
distance
isolation
of
3
miles
and
24
border
rows
to
serve
as
a
trap
crop
for
cotton
pollinators.
Monitoring
for
volunteers
in
experimental
fields
is
also
required
for
one
year
following
experiment
completion.
These
containment
measures
are
needed
to
mitigate
gene
flow
to
wild
and/
or
feral
relatives
of
cotton.
In
their
September
7,
2005
email
message
and
October
3,
2005
letter
to
EPA,
Bayer
proposed
to
net
or
cage
Bt
cotton
plants
during
flowering
as
an
alternative
to
the
mandatory
3
mile
isolation
distance.
This
proposal
is
acceptable.

When
conventional
row
spacing
is
used
(
27"­
40")
in
the
experimental
acreage,
planting
density
and
row
spacing
in
border
areas
must
be
within
20%
(
greater
than
or
less
than)
of
that
found
in
the
experiment.
If
unconventional
row
spacing
is
used
in
the
experiment,
planting
density
and
row
spacing
must
be
the
same
in
the
experimental
acreage
and
border
areas.
These
row
spacing
requirements
are
necessary
to
ensure
that
the
prescribed
border
area
serves
as
an
effective
pollen
deposition
site
for
cotton
pollinators.
In
their
September
7,
2005
email
message
and
October
3,
2005
letter
to
EPA,
Bayer
agreed
to
comply
with
these
row
spacing
guidelines.

Acreage
Calculation
3
Total
EUP
acreage
is
equivalent
to
the
sum
of
the
acreage
comprised
in
each
EUP
test
block.
This
calculation
would
include
all
Cry1Ab
cotton
plants,
non­
Cry1Ab
cotton
plants
(
e.
g.
breeding
plants
or
isoline
control
plots),
and
associated
border
rows
contained
within
the
perimeter
of
the
test
block.
Acreage
calculation
methods
were
described
in
Bayer's
September
7
and
December
5,
2005
email
messages
and
October
3,
2005
letter
to
EPA.

BPPD
Review
This
approach
to
EUP
acreage
calculation
is
acceptable.

Trial
Protocols
Trial
protocols
for
the
proposed
EUP
are
described
below.
The
submission
included
a
table
which
lists,
on
a
state
by
state
basis,
the
number
of
acres
and
locations
where
a
protocol
may
be
implemented.
It
should
be
noted
that
the
acreage
listed
for
each
location
is
the
total
acreage
for
all
protocols
implemented
at
that
location.
Consequently,
specific
acreage
per
location,
per
protocol
is
not
provided.

State
Location
County/
Parish
Protocol
Maximum
Acres
Louisiana
Bossier
Pa.
Efficacy/
Agronomic
10
Louisiana
Madison
Pa.
Efficacy/
Agronomic
10
Mississippi
Cohoma
Co.
Efficacy/
Agronomic
10
Mississippi
Oktibbeha
Co.
Efficacy/
Agronomic
10
Mississippi
Washington
Co.
Efficacy/
Agronomic
20
(
2
locations)
North
Carolina
Halifax
Co.
Efficacy/
Agronomic
10
Puerto
Rico
Sabana
Grande
District
Efficacy/
Dissemination
50
South
Carolina
Dillon
Co.
Efficacy/
Agronomic
20
(
2
locations)
Texas
Gaines
Co.
Efficacy/
Agronomic
10
Texas
Lubbock
Co.
Efficacy/
Agronomic
20
(
2
locations)
Texas
Uvalde
Co.
Efficacy/
Agronomic
100
Texas
Wharton
Co.
Efficacy/
Agronomic
100
TOTAL
370
Protocol
descriptions
include
information
on
the
test
sites
(
states
and
number
of
locations
per
state),
objective,
experimental
design,
plot
treatments,
genotypes,
estimated
planting
and
harvest
schedules
for
each
trial,
and
containment
procedures
(
shipping,
pollen
flow,
and
post­
harvest).
Details
of
these
components
may
be
found
in
the
submission.

BPPD
Review
Acreage
requests
are
acceptable.
However,
it
is
recommended
that
future
submissions
should
include
acreage
breakdowns
on
a
per
state,
per
location,
per
protocol,
and
per
experiment
basis
(
e.
g.
50
total
acres
in
Mississippi,
at
two
separate
locations,
for
X,
Y,
and
Z
breeding
trials).
4
DATA
EVALUATION
RECORD
EPA
Secondary
Reviewer:
Tessa
Milofsky,
M.
S.
Agronomist
STUDY
TYPE:
Experimental
Use
Permit
Request
MRID
NO:
46455102,
46455110,
46455112
DP
BARCODE
NO:
DP315394
DECISION
NO:
353022
TEST
MATERIAL:
Bacillus
thuringiensis
subsp.
berliner
Cry1Ab
PROJECT
NO:
EUP
Request
Vol.
I
and
II
SPONSOR:
Bayer
CropScience
TESTING
FACILITY:
Bayer
CropScience
LP­
BioScience
P.
O.
Box
12014
2
T.
W.
Alexander
Dr.
Research
Triangle
Park,
NC
27709
TITLE
OF
REPORT:
Experimental
Use
Permit
Request
for
Bacillus
thuringiensis
subsp.
berliner
Cry1Ab
Insecticidal
Protein
as
Expressed
in
Cotton
Plants
AUTHOR:
Ali
Scott
STUDY
COMPLETED:
January
15,
2005
GOOD
LABORATORY
PRACTICE:
Not
applicable
CONCLUSION:
Bayer
Crop
Science
(
BCS)
is
applying
for
an
Experimental
Use
Permit
(
EUP)
to
allow
further
evaluation
of
transgenic
cotton
[
Gossypium
hirsutum]
plant
lines
which
express
the
insecticidal
protein,
Cry1Ab,
from
Bacillus
thuringiensis
subsp.
berliner
strain
1715
(
events
T303­
3
and
T304­
40).
The
program
will
be
carried
out
in
15
locations
in
6
states
(
including
Puerto
Rico)
on
370
acres.

CLASSIFICATION:
Acceptable
*
CONTAINS
CONFIDENTIAL
BUSINESS
INFORMATION*

There
is
no
MRID
assigned
to
Volume
I
of
the
Experimental
Use
Permit
(
EUP)
Request
submitted
by
Bayer
CropScience
for
the
product
BCS
Cry1Ab
Cotton.
Volume
I
contains
the
majority
of
the
information
for
the
EUP
request
as
well
as
a
confidential
appendix.
Volume
II
references
several
documents
(
MRIDs
46455102,
46455110,
46455112)
which
contain
supporting
information
including
copies
of
literature
citations.
The
ORNL
review
covers
both
Volume
I
and
II
of
the
BCS
EUP.

Background
5
BCS
has
developed
cotton
[
Gossypium
hirsutum]
plants
that
express
an
insecticidal
protein,
Cry1Ab,
from
a
common
soil
bacterium,
Bacillus
thuringiensis
subsp.
berliner
(
B.
t.
berliner).
The
Cry1Ab
protein
is
effective
in
controlling
lepidopteran
larvae
such
as
bollworm
(
CBW,
Helicoverpa
zea)
and
tobacco
budworm
(
TBW,
Heliothis
virenscens)
larvae,
which
are
a
common
pest
of
cotton.
These
pests
cause
severe
economic
damage
to
the
cotton
crop
if
not
controlled.
If
controlled
by
chemical
pesticides,
there
is
the
need
for
large
amounts
of
pesticide
use
annually
to
control
these
pests.
Small
scale
field
trial
experiments
of
cotton
expressing
Cry1Ab
protein,
conducted
under
notifications
granted
by
the
U.
S.
Department
of
Agriculture's
Animal
and
Plant
Health
Inspection
Service
(
APHIS),
have
shown
the
plant's
ability
to
protect
itself
against
these
pests.

Transgenic
cotton
plants
expressing
Cry1Ab
protein
provide
an
addition
to
growers'
options
for
insect
control
that
potentially
reduces
or
eliminates
the
need
for
other
insecticide
inputs
and
fits
well
within
an
integrated
pest
management
program.
Cry1Ab
is
a
protein
familiar
to
the
Agency,
but
is
has
not
been
used
in
commercial
cotton
as
a
Plant
Incorporated
Protectant
(
PIP).
BCS
is
applying
for
an
Experimental
Use
Permit
(
EUP)
to
allow
further
evaluation
of
these
cotton
plant
lines
in
a
larger
range
of
environmental
conditions.
All
cotton
plants
to
be
evaluated
under
the
EUP
have
been
derived
from
either
transformation
event
number
T303­
3
or
T304­
40.
Several
different
experiments
are
planned
and
can
be
grouped
into:
insect
efficacy
trials,
herbicide
efficacy
evaluation,
agronomic
performance
evaluation
and
breeding
(
line
evaluations).
In
addition
to
these
experimental
plans,
seed
blocks
will
be
grown
as
part
of
this
program
to
evaluate
effects
on
seed
production,
dissemination
and
dormancy,
and
seed
may
be
harvested
for
later
plantings
of
experimental
and
regulatory
field
trials.

The
primary
goal
of
this
experimental
research
program
is
to
evaluate
Cry1Ab
proteincontaining
cotton
plants
for
their
efficacy
against
insect
pests
of
cotton,
as
well
as
to
provide
material
for
agronomic
performance
evaluations.
These
plants
also
contain
an
insecticidal­
inert
ingredient
as
a
selectable
marker,
the
phosphinothricin
acetyltransferase
(
PAT)
protein
that
confers
tolerance
to
glufosinate­
ammonium
herbicides.
The
entire
program
will
be
done
on
a
"
Crop
Destruct"
basis;
no
cottonseed
will
enter
commerce.
Some
of
the
plant
material
will
be
retained
for
scientific
research
and/
or
planting
purposes.
All
other
plant
materials
will
be
destroyed.
There
shall
be
no
unintentional
exposure
to
humans
or
domestic
animals
since
the
program
will
be
conducted
using
containment
precautions
and
in
a
crop
destruct
fashion.
Isolation
will
be
maintained
in
order
to
prevent
any
inadvertent
outcrossing
(
pollination)
from
transgenic
plants
to
non­
transgenic
cotton
plants.
No
environmental
impact
issues
related
to
the
testing
of
these
transgenic
cotton
plants
have
been
identified.
The
Cry1Ab
protein
has
a
specific
range
of
toxicity
to
the
target
lepidopteran
pests
without
having
effects
on
non­
target
beneficial
insects.

In
total,
the
program
will
be
carried
out
in
15
locations
in
six
states
on
370
acres
(
Table
3).
The
Cry1Ab
containing
cotton
will
be
planted
at
a
maximum
rate
of
60,000
seeds
per
acre
(
12
lbs/
acre).
The
level
of
Cry1Ab
protein
in
each
seed
is
approximately
180­
1450
ng
(
0.18­
1.45
:
g);
therefore
the
planting
of
these
Cry1Ab­
containing
seeds
represents
an
application
rate
of
approximately
10.9
to
87
mg
of
Cry1Ab
protein
per
acre
(
using
12
lb
or
5442
g
seed/
acre).
Our
proposed
experimental
research
program
would
total
4
to
32
g
of
Cry1Ab
protein
(
or
0.008
to
0.071
pounds
of
Cry1Ab
protein)
for
370
acres.
The
level
of
Cry1Ab
protein
in
the
different
plant
material
is
only
an
estimation
based
on
current
information.
6
TABLE
3.
Proposed
EUP
acreage,
seed
and
a.
i.
quantities
by
state.

State
Location
County/
Parish
Protocol
Maximum
acres
Maximum
lbs
of
seed
Maximum
amount
of
Cry1Ab
in
seed
planted
Louisiana
Bossier
Pa.
Efficacy/
Agronomic
10
120
0.87
g
Louisiana
Madison
Pa.
Efficacy/
Agronomic
10
120
0.87g
Mississippi
Cohoma
Co.
Efficacy/
Agronomic
10
120
0.87g
Mississippi
Oktibbeha
Co.
Efficacy/
Agronomic
10
120
0.87g
Mississippi
Washington
Co.
Efficacy/
Agronomic
20
(
2
locations)
240
1.74g
North
Carolina
Halifax
Co.
Efficacy/
Agronomic
10
120
0.87g
Puerto
Rico
Sabana
Grande
District
Efficacy/
Dissemination
50
600
4.35g
South
Carolina
Dillon
Co.
Efficacy/
Agronomic
20
(
2
locations)
240
1.74g
Texas
Gaines
Co.
Efficacy/
Agronomic
10
120
0.87g
Texas
Lubbock
Co.
Efficacy/
Agronomic
20
(
2
locations)
240
1.74g
Texas
Uvalde
Co.
Efficacy/
Agronomic
100
1200
8.7g
Texas
Wharton
Co.
Efficacy/
Agronomic
100
1200
8.7g
TOTAL
370
4400
32g
or
0.07
lbs
Experimental
protocol
1.
Efficacy
testing
of
insect
resistant
transgenic
cotton:
The
purpose
of
these
trials
is
to
compare
plant
growth,
morphology,
and
agronomic
performance
among
insect
resistant
transgenic
cotton
lines
in
different
genetic
backgrounds
with
their
respective
nontransgenic
counterparts
when
infested
with
target
pests
(
either
naturally
or
artificially)
as
well
as
in
the
absence
of
target
insects.
Another
objective
of
these
trials
is
to
evaluate
insect
resistance
to
transgene
efficacy
and
subsequently
determine
if
transgene
expression
and
resulting
insect
7
resistance
affect
plant
growth,
morphology,
or
any
facet
of
agronomic
performance
including
fiber
characteristics.
8
Specific
areas
to
be
evaluated
are;
a)
Breeding
lines:
This
is
primarily
for
the
purpose
of
transferring
the
Cry1Ab
event
into
elite
cotton
lines,
in
order
to
evaluate
performance
of
the
gene
in
elite
varieties.
This
will
include
backcrossing
programs
for
initial
evaluation
of
lines,
and
for
production
of
hand­
pollinated
seed
to
conduct
preliminary
yield
evaluations.

b)
Insect
efficacy,
Natural
infestations/
Herbicide
efficacy:
These
trials
will
be
conducted
to
evaluate
the
effectiveness
of
events
T303­
3
and
T304­
40
in
controlling
the
primary
target
pests
CBW
and
TBW.
The
trials
will
include
only
sites
where
the
infestations
occur
naturally.
Large
differences
in
the
level
of
severity
of
the
infestation
and
damage
are
expected
across
locations
and
environments;
and
for
this
reason,
the
evaluation
of
efficacy
across
a
broad
spectrum
of
environmental
and
geographical
conditions
is
needed
to
accurately
evaluate
efficacy
of
the
Cry1Ab
events
against
the
targeted
pests
under
field
conditions.
Additionally,
evaluation
by
independent
experts,
such
as
in
trials
conducted
by
university
researchers
and
other
cooperators,
provides
independently
generated
performance
data.
In
addition,
the
tolerance
to
the
herbicide
glufosinate­
ammonium
provided
by
the
marker
gene
will
be
evaluated.

c)
Insect
efficacy
CBW,
artificial
infestations:
These
trials
will
be
conducted
to
evaluate
effectiveness
of
events
T303­
3
and
T304­
40
in
controlling
the
primary
target
pest
CBW.
The
trials
will
include
sites
where
the
infestations
occur
as
natural
infestations,
as
well
as
those
artificially
infested
to
enhance
the
opportunity
to
observe
extreme
CBW
pressure.
Large
differences
in
the
level
of
severity
of
the
infestation
and
damage
are
expected
across
locations
and
environments;
and
for
this
reason,
the
evaluation
of
efficacy
across
a
broad
spectrum
of
environmental
and
geographical
conditions
is
needed
to
accurately
evaluate
efficacy
of
the
Cry1Ab
events
against
the
targeted
pests
under
field
conditions.
Additionally,
evaluation
by
independent
experts,
such
as
in
trials
conducted
by
university
researchers
and
other
cooperators,
provides
independently
generated
performance
data.

d)
Insect
efficacy
and
other
lepidopterans:
Though
the
primary
target
pest
for
the
Cry1Ab
events
are
TBW
and
CBW,
the
Cry1Ab
protein
has
been
shown
to
have
activity
against
other
lepidopteran
pests
of
cotton,
including
Fall
Armyworm
and
Pink
Bollworm.
These
pests
can
be
significant
in
certain
seasons
and
in
certain
geographical
regions,
though
the
overall
incidence
and
damage
attributable
to
these
pests
is
probably
less
significant
than
for
the
two
main
pests.
Since
these
and
other
lepidopteran
pests
can
cause
significant
economic
damage
in
some
environments,
the
opportunity
for
Cry1Ab
to
provide
control
of
additional
lepidopteran
pests
will
be
evaluated.
The
expanded
acreage
available
under
an
EUP
will
allow
the
flexibility
to
test
for
these
additional
insect
pests
in
planned
experiments.

e)
Agronomic
evaluation:
These
trials
will
be
used
to
document
yield
and
agronomic
performance
of
the
events
containing
Cry1Ab.
Some
trials
will
be
small
plot
evaluations
to
look
at
effects
on
flowering,
maturity,
seed
size,
herbicide
sensitivity,
and
yield,
for
example.
Other
trials
will
use
larger
fields
to
evaluate
whether
Cry1Ab
provides
agronomic
and
yield
benefits
for
production
fields
that
affect
the
economics
of
producing
cotton
seed.
One
could
reasonably
expect
that
use
of
Cry1Ab
in
cotton
lines
will
significantly
reduce
the
amount
of
insecticide
that
will
need
to
be
applied
to
production
fields,
and
will
also
substantially
9
increase
seed
yields
in
production
fields
affected
by
the
targeted
pests.
Evaluation
of
these
questions
is
not
possible
under
small
plot
situations,
and
requires
the
acreage
provided
by
an
EUP.

Efficacy
data
will
be
collected
at
the
following
field
sites
specific
to
the
EUP.

County,
State
Total
acres
Pounds
of
seed
per
acre
Bossier
Parish,
LA
10
acres
12
pounds
per
acre
Madison
Parish,
LA
10
acres
12
pounds
per
acre
Cohoma
County,
MS
10
acres
12
pounds
per
acre
Oktibbeha
County,
MS
10
acres
12
pounds
per
acre
Washington
County,
MS
20
acres
12
pounds
per
acre
(
2
locations)
Halifax
County,
NC
10
acres
12
pounds
per
acre
Sabana
Grande,
PR
50
acres
12
pounds
per
acre
Dillon
County,
SC
20
acres
12
pounds
per
acre
(
2
locations)
Gaines
County,
TX
10
acres
12
pounds
per
acre
Lubbock
County,
TX
20
acres
12
pounds
per
acre
(
2
locations)
Uvalde
County,
TX
100
acres
12
pounds
per
acre
Wharton
County,
TX
100
acres
12
pounds
per
acre
10
Genotypes
to
be
planted:
The
BCS
transgenic
cotton
plants
expressing
the
cry1ab
gene
(
derived
from
vector
pTDL004
or
pTDL008)
will
be
tested
in
11
experimental
lines
with
different
backgrounds.
Likewise,
a
transgenic
commercial
control
will
be
planted.
Two
non­
transgenic
plants
Coker
315
and
a
non­
transgenic
commercial
control
will
also
be
planted.

Trial
design:
The
preferred
statistical
design
is
a
split­
plot
where
the
level
of
insect
infestation
(
sprayed
vs.
nonsprayed)
is
the
main
plot
and
line/
transgene
is
the
subplot.
A
split­
strip
design
may
also
be
used
which
facilitates
insecticide
treatments.
A
randomized
complete
block
design
(
RCBD)
would
suffice,
but
would
not
provide
maximal
precision.

Agronomic
treatments:
Typical
agronomic
inputs
for
conventionally
grown
cotton
for
the
area,
including,
but
not
limited
to:

°
Conventional
herbicide
treatments,
both
pre­
and
post­
planting
°
Granular
insecticide
and/
or
fungicide
application
at
planting
°
Fertilizer
applications
°
Necessary
in­
season
insecticide
applications
for
non­
target
and/
or
target
insects
only
(
see
test
treatments
below)
°
Growth
regulator
application
(
this
should
be
done
sparingly
if
at
all)
°
Additional
hand
weeding
as
necessary
°
Chemical
defoliation
without
boll­
opening
desiccants
Test
treatments:
Test
treatments
involve
natural
infestations
of
target
insects,
chemical
control
of
target
insects,
and
chemical
control
of
non­
target
insects.
Treatments
include
but
are
not
limited
to:
°
Complete
insect
control,
both
target
and
non­
target
in
sprayed
plots
°
Insect
control
of
non­
target
insects
that
allows
or
encourages
natural
infestation
of
target
insect,
in
non
sprayed
plots.

Border
rows:
The
EUP
test
plants
or
the
trial
will
be
surrounded
by
one
or
more
border
rows
of
non­
transgenic
cotton.

Schedule:
Planting
dates
for
continental
USA:
May­
June
Harvest
dates
for
continental
USA:
October­
November
Planting
dates
for
PR:
October­
November
Harvest
dates
for
PR:
April­
May
Activities
and
Agronomic
Practices:
Plots
will
be
harvested
by
hand
or
mechanically.
If
by
hand,
the
bolls
will
be
placed
in
cloth
or
paper
bags
of
such
construction
to
avoid
loss
of
seed
outside
of
the
bags.
If
by
machine,
seed
cotton
will
be
harvested,
transported
and
processed
under
conditions
appropriate
for
the
handling
of
regulated
material.
This
11
includes
separate,
redundant
labeled
packaging
of
all
regulated
material
leaving
the
location.

GPS
coordinates,
stakes,
markers
or
other
methods
will
be
used
to
identify
the
area
where
the
transgenic
plants
are
grown,
and
such
an
area
will
be
subsequently
monitored
for
volunteer
plants
for
an
appropriate
period
of
time.
Volunteer
plants
will
be
terminated
by
hand
weeding,
disking,
herbicide
spraying
or
other
method.

Containment:
EUP
test
plants
will
be
isolated
in
accordance
with
USDA­
APHIS
Performance
Standards
for
regulated
cotton
trials.
Isolation
methods
may
include
one
or
more
of
the
following:
EUP
test
plants
will
be
located
at
least
660
feet
from
other
parties'
sexually
receptive
cotton;
(
2)
a
40
foot
wide
perimeter
of
non­
transgenic
cotton
will
surround
the
transgenic
plants
to
act
as
a
pollen
sink
for
insect
pollinators
(
the
perimeter
cotton
would
be
disposed
of
by
harvesting,
disking
and
monitoring);
(
3)
The
EUP
plants
will
be
netted
during
pollen
shed;
(
4)
temporal
isolation,
where
the
flowering
period
for
the
EUP
plants
will
not
coincide
with
the
presence
of
other
parties'
sexually
receptive
cotton
within
660
feet
of
the
EUP
test
plants.
Open
flowering
EUP
test
plants
may
be
located
within
660
feet
of
sexually
receptive
cotton
provided
such
other
cotton
is
used
only
for
experimental
purposes
and/
or
destroyed.

Following
the
trial
completion,
all
remaining
plant
debris
will
be
destroyed
by
incorporation
in
the
soil.
All
equipment
used
during
crop
destruction
practices
will
be
inspected
and
cleaned
before
leaving
the
field.
Seed
cotton
not
destined
for
further
experimentation
will
be
destroyed
by
incineration
or
deep
burial.

If
harvested
material
is
to
be
ginned,
seed
cotton
will
be
securely
transported
to
a
gin.
Processing
of
harvested
seed
cotton
will
consist
of
either
hand
ginning
or
research/
commercial
scale
ginning.
Hand
ginning
will
be
on
small,
table­
top
gins.
Machine
ginning
will
occur
at
the
ginning
facility
on­
site
using
a
limited
number
of
research/
commercial
scale
gins.
All
packaging
and
waste
will
be
destroyed
by
devitalization.
No
seed
cotton
or
ginning
byproducts
will
be
used
for
food
or
feed.
Ginned
seed
will
be
stored
under
containment
practices
for
regulated
materials.
12
Agronomic
data
collection:
If
available,
the
following
data
may
be
collected
from
the
plots,
using
a
1­
9
scale,
where
applicable:
°
Strain
uniformity:
1=
uniform,
9=
highly
variable
°
Leaf
pubescence:
1=
highly
pubescent,
5=
semi­
smooth,
9=
glabrous
°
Disease
reaction
(
verticillium
wilt,
bacterial
blight,
bronze
wilt,
etc.
IF
applicable):
1=
no
symptoms,
5=
some
symptoms
apparent,
9=
severe
°
Stalk
lodging:
1=
upright,
9=
severely
lodged
°
%
open
bolls
as
a
visual
average
when
uninfested
recurrent
parent
is
40­
60%
open
°
Yield
in
lbs.
lint
per
acre
°
%
lint
°
#
seed
per
boll
°
Boll
size
°
Seed
index
°
Fiber
properties:
length,
length
uniformity,
strength,
micronaire,
elongation
°
Plant
mapping:
plant
map
10
plants
per
plot,
each
of
3
replicates
at
maturity
shortly
before
defoliation.
Data
will
include
plant
height,
number
of
nodes,
and
boll
position.
Boll
damage
ratings
may
be
a
part
of
the
mapping
data.
Information
collected
will
reflect
overall
plant
architecture
and
maturity.

Target
insect
evaluation
data
collection:
Data
will
be
taken
on
insect
damage
for
6­
8
weeks
to
measure
resistance
to
the
infesting
insect.
Insect
infestation
and
damage
data
are
collected
by
examining
the
terminal
and
one
square
or
boll
on
10
plants
per
row
on
each
of
the
two
center
rows
in
each
plot.
Data
collected
may
include:

°
Number
of
live
larvae
in
squares
°
Number
of
live
larvae
in
bolls
°
Number
of
squares
damaged
by
larvae
°
Number
of
bolls
damaged
by
larvae
°
Number
of
damaged
white
flowers
°
Number
of
live
larvae
in
white
flowers
Quality
control
sample
collection:
Leaf
samples
are
to
be
taken
from
two
individual
plants
per
plot
for
QC
purposes.

Experimental
protocol
2.
Agronomic
evaluation
of
insect
resistant
transgenic
cotton:
The
purpose
of
these
trials
is
to
compare
total
agronomic
performance
and
fiber
characteristics
among
the
converted
transgenic
lines
(
per
recurrent
parent)
and
with
their
respective
recurrent
parent
variety
counterpart.
The
goal
is
to
select
lines
that
are
equal
to
or
better
than
the
recurrent
parent,
to
be
advanced
to
additional
testing.
13
Agronomic
performance
and
fiber
characteristics
data
will
be
collected
at
the
following
field
sites
specific
to
the
EUP:

County,
State
Total
acres
Pounds
of
seed
per
acre
Bossier
Parish,
LA
10
acres
12
pounds
per
acre
Madison
Parish,
LA
10
acres
12
pounds
per
acre
Cohoma
County,
MS
10
acres
12
pounds
per
acre
Oktibbeha
County,
MS
10
acres
12
pounds
per
acre
Washington
County,
MS
20
acres
12
pounds
per
acre
(
2
locations)
Halifax
County,
NC
10
acres
12
pounds
per
acre
Dillon
County,
SC
20
acres
12
pounds
per
acre
(
2
locations)
Gaines
County,
TX
10
acres
12
pounds
per
acre
Lubbock
County,
TX
20
acres
12
pounds
per
acre
(
2
locations)
Uvalde
County,
TX
100
acres
12
pounds
per
acre
Wharton
County,
TX
100
acres
12
pounds
per
acre
Genotypes
to
be
planted:
The
BCS
transgenic
cotton
plants
expressing
the
cry1ab
gene
(
derived
from
vector
pTDL004
or
pTDL008)
will
be
tested
in
11
experimental
cotton
lines
with
different
backgrounds.
Likewise,
a
transgenic
commercial
control
will
be
planted.
Two
non­
transgenic
plants,
Coker
315
and
a
non­
transgenic
commercial
control,
will
also
be
planted.

Trial
design:
The
statistical
design
may
be
a
RCBD.
Plots
could
be
2­
or
4­
row
with
3­
4
replications.
Multiple
locations
are
necessary.
Seed
availability
will
dictate
replications
and
number
of
locations.
A
lattice
design
may
also
be
used.
The
precision
of
a
split­
plot
is
typically
not
practical
at
early
stages
of
sister
line
evaluation
due
to
the
large
number
of
transgenic
sister
lines,
but
is
encouraged
if
possible.
In
this
case,
variety
or
genetic
background
would
be
the
main­
plot,
and
transgene
+/­
would
be
the
split­
plot.

Agronomic
treatments:
Typical
agronomic
inputs
for
conventionally
grown
cotton
for
the
area,
including,
but
not
limited
to:

°
Conventional
herbicide
treatments,
both
pre­
and
post­
planting
°
Granular
insecticide
and/
or
fungicide
application
at
planting
°
Fertilizer
applications
°
Necessary
in­
season
chemical
insecticide
applications
°
Growth
regulator
application
(
this
should
be
done
sparingly
if
at
all)
°
Additional
hand
weeding
as
necessary
°
Chemical
defoliation
without
boll­
opening
desiccants
Test
treatments:
No
special
test
treatments
are
required.
If
testing
the
herbicide
resistance
transgene,
the
target
herbicide
is
not
to
be
applied
for
these
trials.
If
testing
the
insect
resistance
transgene,
it
is
imperative
that
the
target
insect
is
completely
controlled
so
that
transgenic
lines
do
not
have
an
advantage
over
the
unconverted
recurrent
parent
lines.
14
Border
rows:
The
EUP
test
plants
or
the
trial
will
be
surrounded
by
one
or
more
border
rows
of
cotton.

Schedule:
Planting
dates:
May­
June
Harvest
dates:
October­
November
Activities
and
agronomic
practices:
Plots
will
be
harvested
by
hand
or
mechanically.
If
by
hand,
the
bolls
will
be
placed
in
cloth
or
paper
bags
of
such
construction
to
avoid
loss
of
seed
outside
of
the
bags.
If
by
machine,
seed
cotton
will
be
harvested,
transported
and
processed
under
conditions
appropriate
for
the
handling
of
regulated
material.
This
includes
separate,
redundant
labeled
packaging
of
all
regulated
material
leaving
the
location.

GPS
coordinates,
stakes,
markers
or
other
methods
will
be
used
to
identify
the
area
where
the
transgenic
plants
are
grown,
and
such
an
area
will
be
subsequently
monitored
for
volunteer
plants
for
an
appropriate
period
of
time.
Volunteer
plants
will
be
terminated
by
hand
weeding,
disking,
herbicide
spraying
or
other
method.

Containment:
EUP
test
plants
will
be
isolated
in
accordance
with
USDA­
APHIS
Performance
Standards
for
regulated
cotton
trials.
Isolation
methods
may
include
one
or
more
of
the
following:
EUP
test
plants
will
be
located
at
least
660
feet
from
other
parties'
sexually
receptive
cotton;
(
2)
a
40
footwide
perimeter
of
non­
transgenic
cotton
will
surround
the
transgenic
plants
to
act
as
a
pollen
sink
for
insect
pollinators
(
the
perimeter
cotton
would
be
disposed
of
by
harvesting,
disking
and
monitoring);
(
3)
The
EUP
plants
will
be
netted
during
pollen
shed;
(
4)
temporal
isolation,
where
the
flowering
period
for
the
EUP
plants
will
not
coincide
with
the
presence
of
other
parties'
sexually
receptive
cotton
within
660
feet
of
the
EUP
test
plants.
Open
flowering
EUP
test
plants
may
be
located
within
660
feet
of
sexually
receptive
cotton
provided
such
other
cotton
is
used
only
for
experimental
purposes
and/
or
destroyed.

Following
the
trial
completion,
all
remaining
plant
debris
will
be
destroyed
by
incorporation
in
the
soil.
All
equipment
used
during
crop
destruction
practices
will
be
inspected
and
cleaned
before
leaving
the
field.
Seed
cotton
not
destined
for
further
experimentation
will
be
destroyed
by
incineration
or
deep
burial.

If
harvested
material
is
to
be
ginned,
seed
cotton
will
be
securely
transported
to
a
gin.
Processing
of
harvested
seed
cotton
will
consist
of
either
hand
ginning
or
research/
commercial
scale
ginning.
Hand
ginning
will
be
on
small,
table­
top
gins.
Machine
ginning
will
occur
at
the
ginning
facility
on­
site
on
a
limited
number
of
research/
commercial
scale
gins.
All
packaging
and
waste
will
be
destroyed
by
devitalization.
No
seed
cotton
or
ginning
by­
products
will
be
used
for
food
or
feed.
Ginned
seed
will
be
stored
under
containment
practices
for
regulated
materials.

Agronomic
data
collection
If
available,
the
following
data
may
be
collected
from
the
plots,
using
a
1­
9
scale,
where
15
applicable:
°
Strain
uniformity:
1=
uniform,
9=
highly
variable
°
Leaf
pubescence:
1=
highly
pubescent,
5=
semi­
smooth,
9=
glabrous
°
Disease
reaction
(
verticillium
wilt,
bacterial
blight,
bronze
wilt,
etc.
IF
applicable):
1=
no
symptoms,
5=
some
symptoms
apparent,
9=
severe
°
Stalk
lodging:
1=
upright,
9=
severely
lodged
°
Number
days
to
first
flower:
as
an
average
of
the
plot
°
Number
of
days
to
first
open
boll:
as
an
average
of
the
plot
°
Boll
type:
1=
loose,
5=
intermediate,
9=
storm
proof
°
Plant
height:
in
inches
at
maturity
°
Total
nodes:
10
plants
per
plot
at
maturity
°
%
open
bolls
as
a
visual
average
when
uninfested
recurrent
parent
is
40­
60%
open
°
Yield
in
lbs.
lint
per
acre
°
%
lint
°
#
seed
per
boll
°
Boll
size
°
Seed
index
°
Fiber
properties:
length,
length
uniformity,
strength,
micronaire,
elongation
Experimental
Protocol
3.
Dissemination
Study
Of
Insect
Resistant
Transgenic
Cotton:
The
purpose
of
this
trial
is
to
evaluate
possible
dissemination
of
pollen.
In
addition,
seed
will
be
harvested
to
increase
the
amount
of
seed
from
tested
varieties
for
future
trials.

Pollen
dissemination
data
will
be
collected
at
the
Puerto
Rico
field
site
specific
to
the
EUP.

County,
State
Total
acres
Pounds
of
seed
per
acre
Sabana
Grande,
PR
50
acres
12
pounds
per
acre
Genotypes
to
be
planted:
The
BCS
transgenic
cotton
plants
expressing
the
cry1ab
gene
(
derived
from
vector
pTDL004
or
pTDL008)
will
be
tested
along
with
non­
transgenic
plants,
Coker
315.

Trial
design:
A
planted
block
consisting
of
transgenic
seed
will
be
planted
with
non
transgenic
border
rows
of
cotton
planted
on
all
four
sides
of
the
perimeter
of
the
seed
block.
The
spacing
between
each
row
is
approximately
38
inches.

Agronomic
treatments:
Typical
agronomic
inputs
for
conventionally
grown
cotton
for
the
area,
including,
but
not
limited
to:
°
Conventional
herbicide
treatments,
both
pre­
and
post­
planting
°
Granular
insecticide
and/
or
fungicide
application
at
planting
°
Fertilizer
applications
°
Necessary
in­
season
insecticide
applications
for
non­
target
and/
or
target
insects
only
°
Growth
regulator
application
(
this
should
be
done
sparingly
if
at
all)
°
Additional
hand
weeding
as
necessary
°
Chemical
defoliation
without
boll­
opening
desiccants,
as
this
would
mask
maturity
16
Test
treatments:
Two
applications
of
28
oz
Liberty/
acre.
One
application
at
7
leaf
stage
and
another
at
20%
bloom
of
the
block.

Border
rows:
The
EUP
test
plants
or
the
trial
will
be
surrounded
by
one
or
more
border
rows
of
cotton.

Schedule:
Planting
dates:
October­
November
Harvest
dates:
April­
May
Activities
and
agronomic
practices:
Plots
will
be
harvested
by
hand
or
mechanically.
If
by
hand,
the
bolls
will
be
placed
in
cloth
or
paper
bags
of
such
construction
to
avoid
loss
of
seed
outside
of
the
bags.
If
by
machine,
seed
cotton
will
be
harvested,
transported
and
processed
under
conditions
appropriate
for
the
handling
of
regulated
material.
This
includes
separate,
redundant
labeled
packaging
of
all
regulated
material
leaving
the
location.

GPS
coordinates,
stakes,
markers
or
other
methods
will
be
used
to
identify
the
area
where
the
transgenic
plants
are
grown,
and
such
an
area
will
be
subsequently
monitored
for
volunteer
plants
for
an
appropriate
period
of
time.
Volunteer
plants
will
be
terminated
by
hand
weeding,
disking,
herbicide
spraying
or
other
method.

Containment:
EUP
test
plants
will
be
isolated
in
accordance
with
USDA­
APHIS
Performance
Standards
for
regulated
cotton
trials.
Isolation
methods
may
include
one
or
more
of
the
following:
EUP
test
plants
will
be
located
at
least
660
feet
from
other
parties'
sexually
receptive
cotton;
(
2)
a
40
footwide
perimeter
of
non­
transgenic
cotton
will
surround
the
transgenic
plants
to
act
as
pollen
sink
for
insect
pollinators
(
the
perimeter
cotton
would
be
disposed
of
by
harvesting,
disking
and
monitoring);
(
3)
the
EUP
plants
will
be
netted
during
pollen
shed;
(
4)
temporal
isolation,
where
the
flowering
period
for
the
EUP
plants
will
not
coincide
with
the
presence
of
other
parties'
sexually
receptive
cotton
within
660
feet
of
the
EUP
test
plants.
Open
flowering
EUP
test
plants
may
be
located
within
660
feet
of
sexually
receptive
cotton
provided
such
other
cotton
is
used
only
for
experimental
purposes
and/
or
destroyed.

Following
the
trial
completion,
all
remaining
plant
debris
will
be
destroyed
by
incorporation
in
the
soil.
All
equipment
used
during
crop
destruction
practices
will
be
inspected
and
cleaned
before
leaving
the
field.
Seed
cotton
not
destined
for
further
experimentation
will
be
destroyed
by
incineration
or
deep
burial.

If
harvested
material
is
to
be
ginned,
seed
cotton
will
be
transported
securely
to
a
gin.
Processing
of
harvested
seed
cotton
will
consist
of
either
hand
ginning
or
research/
commercial
scale
ginning.
Hand
ginning
will
be
on
small,
table­
top
gins.
Machine
ginning
will
occur
at
the
ginning
facility
on­
site
on
a
limited
number
of
research/
commercial
scale
gins.
All
packaging
and
waste
will
be
destroyed
by
17
devitalization.
No
seed
cotton
or
ginning
by­
products
will
be
used
for
food
or
feed.
Ginned
seed
will
be
stored
under
containment
practices
for
regulated
materials.

Data
requirements:
Four
hundred
plants
within
the
transgenic
block
will
be
tested
via
dPCR
for
the
target
gene
and
non­
target
genes.
Thirty
thousand
seeds
from
bordering
non­
transgenic
cotton
will
be
harvested
and
tested
for
the
presence
of
the
target
gene
by
lateral
flow
strips
to
determine
if
any
outcrossing
occurred.
If
outcrossing
is
found
to
occur,
the
study
will
determine
at
what
level.

Future
plans:
Transgenic
seed
harvested
from
trial
may
be
used
for
regulatory
trials
at
a
later
date
and
for
a
laboratory
dormancy
study.

Experimental
protocol
4.
Production
of
sample
material
for
use
in
regulatory
studies:
The
purpose
of
these
trials
is
to
produce
sample
materials
for
two
types
of
regulatory
trials,
as
part
of
the
requirement
of
regulatory
agencies
all
over
the
world
for
the
submission
of
registration
packages.
Material
will
be
needed
for:

Production
of
sample
material
for
use
in
poultry
and
cattle
feeding
studies.
Production
of
sample
material
for
use
in
composition
studies.

Material
for
feeding
studies
will
be
collected
from
one
of
these
field
sites.

County,
State
Total
acres
Pounds
of
seed
per
acre
Uvalde
County,
TX
100
acres
12
pounds
per
acre
Wharton
County,
TX
100
acres
12
pounds
per
acre
Material
for
composition
studies
will
be
collected
from
one
acre
at
nine
of
the
following
sites.

County,
State
Total
acres
Pounds
of
seed
per
acre
Bossier
Parish,
LA
10
acres
12
pounds
per
acre
Madison
Parish,
LA
10
acres
12
pounds
per
acre
Cohoma
County,
MS
10
acres
12
pounds
per
acre
Oktibbeha
County,
MS
10
acres
12
pounds
per
acre
Washington
County,
MS
20
acres
12
pounds
per
acre
(
2
locations)
Halifax
County,
NC
10
acres
12
pounds
per
acre
Dillon
County,
SC
20
acres
12
pounds
per
acre
(
2
locations)
Gaines
County,
TX
10
acres
12
pounds
per
acre
Lubbock
County,
TX
20
acres
12
pounds
per
acre
(
2
locations)
Uvalde
County,
TX
100
acres
12
pounds
per
acre
Wharton
County,
TX
100
acres
12
pounds
per
acre
Genotypes
to
be
planted:
The
BCS
transgenic
cotton
plants
expressing
the
cry1ab
gene
(
derived
from
vector
pTDL004
or
pTDL008)
will
be
tested
in
11
experimental
lines
with
different
backgrounds.
Likewise,
a
transgenic
commercial
control
will
be
planted.
Two
18
non­
transgenic
plants,
Coker
315,
and
a
non­
transgenic
commercial
control
will
also
be
planted.

Trial
design:
For
the
trial
to
produce
material
for
the
feeding
studies,
each
plot
will
be
approximately
40
acres
in
order
to
produce
the
required
amount
of
seed
per
treatment.
Thus,
approximately
80
acres
of
the
Cry1Ab
cotton
will
be
planted.
For
the
trial
to
produce
material
for
the
composition
studies,
a
maximum
of
1
acre
will
be
planted
at
each
site
for
sample
production.
Of
this,
a
maximum
of
0.4
acres
will
be
planted
with
transgenic
cotton
seed.
One
to
five
sites
will
be
planted
each
in
the
states
of
Louisiana,
Mississippi,
North
Carolina,
South
Carolina
or
Texas,
but
only
for
a
combined
maximum
of
9
sites
total.

Agronomic
treatments:
Typical
agronomic
inputs
for
conventionally
grown
cotton
for
the
area,
including,
but
not
limited
to:
°
Conventional
herbicide
treatments,
both
pre­
and
post­
planting
°
Granular
insecticide
and/
or
fungicide
application
at
planting
°
Fertilizer
applications
°
Necessary
in­
season
chemical
insecticide
applications
°
Growth
regulator
application
(
this
should
be
done
sparingly
if
at
all)
°
Additional
hand
weeding
as
necessary
°
Chemical
defoliation
without
boll­
opening
desiccants,
as
this
would
mask
maturity
Test
treatments:
For
the
trial
to
produce
material
for
the
feeding
studies,
a
total
of
4
plots
will
be
planted.
The
transgenic
plots
and
the
non­
transgenic
control
plots
may
be
planted
in
the
same
field.
Plots
will
not
be
replicated.
Two
plots
(
one
plot
sprayed
with
glufosinate
ammonium
and
one
unsprayed
plot)
will
be
planted
using
material
derived
from
either
transformation
event
T303_
3
or
T304­
40.
Two
non­
transgenic,
non
glufosinate
ammonium
tolerant
control
plots
will
also
be
planted.
Plot
preparation
and
planting
(
e.
g.,
row
and
plant
spacing,
etc.)
will
follow
local
commercial
practice
for
cotton.

For
the
trial
to
produce
material
for
the
composition
studies,
three
treatments
will
be
replicated
three
times
at
each
site,
for
a
total
of
nine
plots.
One
treatment
will
be
the
nontransgenic
cotton,
one
will
be
the
transgenic
cotton
treated
with
glufosinate
ammonium,
and
the
other
will
be
the
transgenic
cotton
not
treated
with
glufosinate
ammonium.
Glufosinate
ammonium
treatments
will
be
made
to
three
of
the
transgenic
plots
(
the
transgenic
cotton
treated
with
glufosinate
treatment).
Each
plot
will
contain
at
least
six
rows
and
be
large
enough
to
allow
at
least
the
required
amount
of
samples
to
be
obtained
without
sampling
plants
located
on
the
edges
of
the
plot.
The
plot(
s)
will
be
large
enough
to
allow
treatment
with
commercial
type
or
small
plot
application
equipment.

Border
rows:
The
EUP
test
plants
or
the
trial
will
be
surrounded
by
one
or
more
border
rows
of
cotton.

Schedule
Planting
dates:
May­
June
19
Harvest
dates:
October­
November
Activities
and
agronomic
practices:
After
planting,
transgenic
and
control
seeds
that
were
not
planted
will
be
collected.
The
transgenic
seeds
will
be
returned
to
the
attention
of
the
Study
Director.
Any
non­
transgenic
seeds
that
are
not
planted
will
be
disposed
of
at
the
discretion
of
the
Cooperator.
The
disposition
of
the
seeds
will
be
documented.
Plots
will
be
harvested
by
hand
or
mechanically.
The
harvested
seed
cotton
will
be
ginned
at
or
near
the
field
location
to
obtain
required
fuzzy
seed
samples.
A
minimum
of
four
pounds
of
seed
will
be
collected
per
treatment
regimen.
Lint
obtained
from
the
ginning
will
be
disposed
of.
Seed
samples
will
be
packed,
placed
in
frozen
storage
and
ultimately
shipped,
frozen,
to
the
Study
Director
and/
or
to
a
designated
analytical
laboratory
for
composition
analyses.
After
the
field
phase
of
the
study
is
complete,
the
remaining
transgenic
and
non­
transgenic
cotton
plants
will
be
destroyed
by
plowing
the
crop
into
the
ground.

Containment:
EUP
test
plants
will
be
isolated
in
accordance
with
USDA­
APHIS
Performance
Standards
for
regulated
cotton
trials.
Isolation
methods
may
include
one
or
more
of
the
following:
EUP
test
plants
will
be
located
at
least
660
feet
from
other
parties'
sexually
receptive
cotton;
(
2)
a
40
footwide
perimeter
of
non­
transgenic
cotton
will
surround
the
transgenic
plants
to
act
as
a
pollen
sink
for
insect
pollinators
(
the
perimeter
cotton
would
be
disposed
of
by
harvesting,
disking
and
monitoring);
(
3)
the
EUP
plants
will
be
netted
during
pollen
shed;
(
4)
temporal
isolation,
where
the
flowering
period
for
the
EUP
plants
will
not
coincide
with
the
presence
of
other
parties'
sexually
receptive
cotton
within
660
feet
of
the
EUP
test
plants.
Open
flowering
EUP
test
plants
may
be
located
within
660
feet
of
sexually
receptive
cotton
provided
such
other
cotton
is
used
only
for
experimental
purposes
and/
or
destroyed.
Following
the
trial
completion,
all
remaining
plant
debris
will
be
destroyed
by
incorporation
in
the
soil.
All
equipment
used
during
crop
destruction
practices
will
be
inspected
and
cleaned
before
leaving
the
field.
Seed
cotton
not
destined
for
further
experimentation
will
be
destroyed
by
incineration
or
deep
burial.

If
harvested
material
is
to
be
ginned,
seed
cotton
will
be
securely
transported
to
a
gin.
Processing
of
harvested
seed
cotton
will
consist
of
either
hand
ginning
or
research/
commercial
scale
ginning.
Hand
ginning
will
be
on
small,
table­
top
gins.
Machine
ginning
will
occur
at
the
ginning
facility
on­
site
on
a
limited
number
of
research/
commercial
scale
gins.
All
packaging
and
waste
will
be
destroyed
by
devitalization.
No
seed
cotton
or
ginning
by­
products
will
be
used
for
food
or
feed.
Ginned
seed
will
be
stored
under
containment
practices
for
regulated
materials.

Data
requirements:
Two
treatments
of
Cry1Ab
cotton
seed
will
be
conducted.
One
treatment
will
be
from
plants
that
were
treated
with
glufosinate
ammonium
and
one
treatment
will
be
from
plants
that
were
not
treated
with
glufosinate
ammonium.
For
the
feeding
studies,
a
total
of
22,000
pounds
of
BCS
Cry1Ab
cotton
seed
must
be
produced.
In
addition
to
the
Cry1Ab
cotton
material,
a
nongenetically
modified
line
of
the
same
genetic
background
and
another
commercially
available
line
will
also
be
produced
to
provide
material
for
comparative
treatments.
Cotton
seed
will
be
harvested
from
each
20
plot,
including
edge
rows,
at
normal
maturity.
Material
from
the
control
non­
transgenic
plots
will
be
harvested
before
harvesting
the
transgenic
plots,
if
they
were
established
in
the
same
field.
The
harvested
seed
cotton
will
be
ginned
to
obtain
the
required
seed
samples
to
prepare
meal
for
feeding
studies.
Ginning
will
be
done
at
or
near
the
field
site.
If
the
seed
cotton
is
transported
to
the
gin,
the
samples
will
be
contained
in
an
enclosed
or
covered
vehicle
or
module.
Ginning
may
be
performed
on
a
commercial
gin,
under
the
supervision
of
the
Cooperator.

The
ginned
seed
samples
will
be
stored
for
transport
at
ambient
temperature
in
suitable
containers
such
as
large
bulk
bags.
Sub­
samples
will
be
sent
to
BCS.
The
bulk
samples
will
be
shipped
to
a
processing
facility
for
processing
into
meal.
Transgenic
cotton
lint
may
be
retained
by
BCS
after
ginning
for
immediate
storage
by
Bayer
and
possible
future
commercial
use
after
registration.
Alternatively,
it
will
be
disposed
of.
Any
"
waste"
transgenic
lint
(
e.
g.,
spillage
or
clean­
out
waste)
will
be
disposed
of.
After
samples
are
collected,
any
remaining
transgenic
crop
(
left
in
the
field
after
harvesting)
will
be
destroyed.

Future
plans:
Transgenic
seed
harvested
may
be
used
for
regulatory
trials
at
a
later
date.

Reviewer's
comments:
1)
The
Cry1Ab
concentration
in
T303­
3
and
T304­
40
seeds
needs
to
be
clarified.
The
EUP
states
the
level
of
Cry1Ab
protein
in
each
seed
is
approximately
180­
1450
ng
(
0.18­
1.45
:
g).
In
contrast,
the
preliminary
protein
expression
data
presented
in
MRID
46477404
has
concentrations
of
Cry1Ab
in
seeds
from
event
T303­
3
and
T304­
40
being
15.6
:
g/
g
and
2.2
:
g/
g
fresh
weight,
respectively
(
2
to16
:
g
of
Cry1Ab
per
0.91g
dry
seed).

2)
A
breeding
tree
is
needed
to
clarify
the
parental
lines
and
backcrossing
for
BCS
events
T303­
3
and
T304­
40.