def run_custom_protocol(
well_volume: float = 1.0,
pipette_type: StringSelection('p300-Single', 'p50-Single', 'p10-Single',
'p300-Multi', 'p50-Multi',
'p10-Multi') = 'p300-Single'):
pip_name = pipette_type.split('-') # Check which pipette type
if pipette_type == 'p300-Single':
pipette = instruments.P300_Single(mount='left', tip_racks=[tiprack])
elif pipette_type == 'p50-Single':
pipette = instruments.P50_Single(mount='left', tip_racks=[tiprack])
elif pipette_type == 'p10-Single':
pipette = instruments.P10_Single(mount='left', tip_racks=[tiprack])
elif pipette_type == 'p300-Multi':
pipette = instruments.P300_Multi(mount='left', tip_racks=[tiprack])
elif pipette_type == 'p50-Multi':
pipette = instruments.P50_Multi(mount='left', tip_racks=[tiprack])
elif pipette_type == 'p10-Multi':
pipette = instruments.P10_Multi(mount='left', tip_racks=[tiprack])
alternating_wells = []
for column in plate.cols():
if pip_name[1] == 'Multi':
alternating_wells.append(column.wells('A'))
alternating_wells.append(column.wells('B'))
else:
alternating_wells.append(column.wells('A', length=8, step=2))
alternating_wells.append(column.wells('B', length=8, step=2))
pipette.distribute(well_volume, trough.wells('A1'), alternating_wells)
def mount_pipette(pipette_type, mount, tiprack_slot):
if pipette_type == 'p10-multi':
tip_rack = [
labware.load('tiprack-10ul', slot) for slot in tiprack_slot
]
pipette = instruments.P10_Multi(mount=mount, tip_racks=tip_rack)
elif pipette_type == 'p50-multi':
tip_rack = [
labware.load('opentrons-tiprack-300ul', slot)
for slot in tiprack_slot
]
pipette = instruments.P50_Multi(mount=mount, tip_racks=tip_rack)
else:
tip_rack = [
labware.load('opentrons-tiprack-300ul', slot)
for slot in tiprack_slot
]
pipette = instruments.P300_Multi(mount=mount, tip_racks=tip_rack)
return pipette
def run_custom_protocol(pipette_type: StringSelection(
'p300_Multi', 'p50_Single', 'p300_Single', 'p1000_Single', 'p10_Multi',
'p50_Multi', 'p10_Single') = 'p300_Multi',
pipette_mount: StringSelection('left',
'right') = 'left',
sample_number: int = 24,
sample_volume: float = 20,
bead_ratio: float = 1.8,
elution_buffer_volume: float = 200,
incubation_time: float = 1,
settling_time: float = 1,
drying_time: float = 5):
total_tips = sample_number * 8
tiprack_num = total_tips // 96 + (1 if total_tips % 96 > 0 else 0)
slots = ['3', '5', '6', '7', '8', '9', '10', '11'][:tiprack_num]
if pipette_type == 'p1000_Single':
tipracks = [labware.load('tiprack-1000ul', slot) for slot in slots]
pipette = instruments.P1000_Single(mount=pipette_mount,
tip_racks=tipracks)
elif pipette_type == 'p300_Single':
tipracks = [labware.load('tiprack-200ul', slot) for slot in slots]
pipette = instruments.P300_Single(mount=pipette_mount,
tip_racks=tipracks)
elif pipette_type == 'p50_Single':
tipracks = [labware.load('tiprack-200ul', slot) for slot in slots]
pipette = instruments.P50_Single(mount=pipette_mount,
tip_racks=tipracks)
elif pipette_type == 'p10_Single':
tipracks = [labware.load('tiprack-10ul', slot) for slot in slots]
pipette = instruments.P10_Single(mount=pipette_mount,
tip_racks=tipracks)
elif pipette_type == 'p10_Multi':
tipracks = [labware.load('tiprack-10ul', slot) for slot in slots]
pipette = instruments.P10_Multi(mount=pipette_mount,
tip_racks=tipracks)
elif pipette_type == 'p50_Multi':
tipracks = [labware.load('tiprack-200ul', slot) for slot in slots]
pipette = instruments.P50_Multi(mount=pipette_mount,
tip_racks=tipracks)
elif pipette_type == 'p300_Multi':
tipracks = [labware.load('tiprack-200ul', slot) for slot in slots]
pipette = instruments.P300_Multi(mount=pipette_mount,
tip_racks=tipracks)
mode = pipette_type.split('_')[1]
if mode == 'Single':
if sample_number <= 5:
reagent_container = labware.load('tube-rack-2ml', '4')
liquid_waste = labware.load('trough-12row', '5').wells('A12')
else:
reagent_container = labware.load('trough-12row', '4')
liquid_waste = reagent_container.wells('A12')
samples = [well for well in mag_plate.wells()[:sample_number]]
output = [well for well in output_plate.wells()[:sample_number]]
else:
reagent_container = labware.load('trough-12row', '4')
liquid_waste = reagent_container.wells('A12')
col_num = sample_number // 8 + (1 if sample_number % 8 > 0 else 0)
samples = [col for col in mag_plate.cols()[:col_num]]
output = [col for col in output_plate.cols()[:col_num]]
# Define reagents and liquid waste
beads = reagent_container.wells(0)
ethanol = reagent_container.wells(1)
elution_buffer = reagent_container.wells(2)
# Define bead and mix volume
bead_volume = sample_volume * bead_ratio
if bead_volume / 2 > pipette.max_volume:
mix_vol = pipette.max_volume
else:
mix_vol = bead_volume / 2
total_vol = bead_volume + sample_volume + 5
# Mix beads and PCR samples
for target in samples:
pipette.pick_up_tip()
pipette.mix(5, mix_vol, beads)
pipette.transfer(bead_volume, beads, target, new_tip='never')
pipette.mix(10, mix_vol, target)
pipette.blow_out()
pipette.drop_tip()
# Incubate beads and PCR product at RT for 5 minutes
pipette.delay(minutes=incubation_time)
# Engagae MagDeck and incubate
mag_deck.engage()
pipette.delay(minutes=settling_time)
# Remove supernatant from magnetic beads
pipette.set_flow_rate(aspirate=25, dispense=150)
for target in samples:
pipette.transfer(total_vol, target, liquid_waste, blow_out=True)
# Wash beads twice with 70% ethanol
air_vol = pipette.max_volume * 0.1
for cycle in range(2):
for target in samples:
pipette.transfer(200,
ethanol,
target,
air_gap=air_vol,
new_tip='once')
pipette.delay(minutes=1)
for target in samples:
pipette.transfer(200, target, liquid_waste, air_gap=air_vol)
# Dry at RT
pipette.delay(minutes=drying_time)
# Disengage MagDeck
mag_deck.disengage()
# Mix beads with elution buffer
if elution_buffer_volume / 2 > pipette.max_volume:
mix_vol = pipette.max_volume
else:
mix_vol = elution_buffer_volume / 2
for target in samples:
pipette.pick_up_tip()
pipette.transfer(elution_buffer_volume,
elution_buffer,
target,
new_tip='never')
pipette.mix(20, mix_vol, target)
pipette.drop_tip()
# Incubate at RT for 3 minutes
pipette.delay(minutes=5)
# Engagae MagDeck for 1 minute and remain engaged for DNA elution
mag_deck.engage()
pipette.delay(minutes=settling_time)
# Transfer clean PCR product to a new well
for target, dest in zip(samples, output):
pipette.transfer(elution_buffer_volume, target, dest, blow_out=True)
# Trash = labware.load('One-Column-reservoir','3')
temp_deck2 = modules.load('tempdeck', '10')
temp_plate = labware.load('1ml_Covaris', '10', share=True)
#mag_deck = modules.load('magdeck', '7')
#mag_plate = labware.load('biorad-hardshell-96-PCR', '7', share=True)
tipracks_10 = [
labware.load('tiprack-10ul', slot, share=True) for slot in ['8', '5']
]
tipracks_200 = [
labware.load('tiprack-200ul', slot, share=True) for slot in ['9', '11']
]
#### PIPETTE SETUP ####
m10 = instruments.P10_Multi(mount='left', tip_racks=tipracks_10)
m300 = instruments.P300_Multi(mount='right',
min_volume=30,
max_volume=200,
aspirate_flow_rate=100,
dispense_flow_rate=200,
tip_racks=tipracks_200)
## Enzyme SETUP
Enzyme_ER = Cold_plate.wells('A1')
# Enzyme_Lig = Cold_plate.wells('A2')
# Enzyme_Fill = Cold_plate.wells('A3')
## Reagent SETUP
ER_mastermix = Cold_plate.wells('A4')
spacing=(17.5, 0), # distances (mm) between each (column, row)
diameter=5, # diameter (mm) of each well on the plate
depth=30, # depth (mm) of each well on the plate
volume=10000)
PG = labware.load('epMotion_10mL', '11', 'PG')
plate_384 = labware.load('corning_384_wellplate_112ul_flat', '2', 'plate')
sample_1 = labware.load('biorad_96_wellplate_200ul_pcr', '3')
sample_2 = labware.load('biorad_96_wellplate_200ul_pcr', '6')
tiprack_200 = labware.load('tiprack-200ul', '1', 'p200rack')
tiprack_10_1 = labware.load('opentrons_96_tiprack_10ul', '4')
tiprack_10_2 = labware.load('opentrons_96_tiprack_10ul', '7')
P10_8 = instruments.P10_Multi(mount='left',
tip_racks=[tiprack_10_1, tiprack_10_2])
P50_8 = instruments.P50_Multi(mount='right', tip_racks=[tiprack_200])
alternating_wells = []
for column in plate_384.cols():
alternating_wells.append(column.wells('A'))
alternating_wells.append(column.wells('B'))
##Transfer the Picogreen
while assay_start < (assay_end - 6):
P50_8.pick_up_tip()
P50_8.transfer(19,
PG.wells('A1'),
alternating_wells[assay_start:assay_start + 6],
new_tip='never')
def run_custom_protocol(
well_volume: float = 50.0,
reagent_well: StringSelection('A1', 'A2', 'A3', 'A4', 'A5', 'A6', 'A7',
'A8', 'A9', 'A10', 'A11', 'A12') = 'A1',
plate_starting_column: str = '1',
number_of_columns_to_fill: int = 24,
pipette_type: StringSelection('p300-Single', 'p50-Single',
'p10-Single', 'p300-Multi', 'p50-Multi',
'p10-Multi') = 'p50-Multi',
mix_reagent_before_transfer: StringSelection('True',
'False') = 'True'):
if int(plate_starting_column) <= 0 or int(plate_starting_column) > 24:
raise Exception("Plate starting column number must be 1-24.")
if (int(plate_starting_column) + number_of_columns_to_fill) > 25:
raise Exception("Number of columns to fill exceeds plate's limit.")
if mix_reagent_before_transfer == 'False':
mix_times = 0
else:
mix_times = 5
pip_name = pipette_type.split('-') # Check which pipette type
if pip_name[0] == 'p10':
tiprack = labware.load('tiprack-10ul', '1', 'p10rack')
else:
tiprack = labware.load('opentrons-tiprack-300ul', '1', 'p300rack')
if pip_name[0] != 'p10' and well_volume < 5:
raise Exception("Cannot transfer volume this small without p10.")
if pip_name[0] == 'p30' and well_volume < 30:
raise Exception("Cannot transfer volume this small with p300.")
if pipette_type == 'p300-Single':
pipette = instruments.P300_Single(mount='left', tip_racks=[tiprack])
elif pipette_type == 'p50-Single':
pipette = instruments.P50_Single(mount='left', tip_racks=[tiprack])
elif pipette_type == 'p10-Single':
pipette = instruments.P10_Single(mount='left', tip_racks=[tiprack])
elif pipette_type == 'p300-Multi':
pipette = instruments.P300_Multi(mount='left', tip_racks=[tiprack])
elif pipette_type == 'p50-Multi':
pipette = instruments.P50_Multi(mount='left', tip_racks=[tiprack])
elif pipette_type == 'p10-Multi':
pipette = instruments.P10_Multi(mount='left', tip_racks=[tiprack])
if pip_name[1] == 'Multi':
alternating_wells = []
for column in plate.cols(plate_starting_column, to='24'):
alternating_wells.append(column.wells('A'))
alternating_wells.append(column.wells('B'))
dests = alternating_wells[:number_of_columns_to_fill * 2]
else:
dests = plate.cols(plate_starting_column,
length=number_of_columns_to_fill)
pipette.distribute(well_volume,
trough.wells(reagent_well),
dests,
mix_before=(mix_times, pipette.max_volume))
# PCR strips with cDNA
pcr_strips = labware.load('PCR-strip-tall', '3')
# tip rack for p10 pipettes
tip10_rack = labware.load('tiprack-10ul', '10')
tip10_rack2 = labware.load('tiprack-10ul', '11')
# p10 (1 - 10 uL) (single)
p10single = instruments.P10_Single(
mount='right',
tip_racks=[tip10_rack]
)
# p10 (1 - 10 uL) (multi)
p10multi = instruments.P10_Multi(
mount='left',
tip_racks=[tip10_rack2]
)
def run_custom_protocol(vol1: float=8, vol2: float=8, cdna_vol: float=8,
num_strips: int=6):
# set cols of 96 well plate that get each mix
# 1 to 6 as written, will change based on num_strips
mix1_cols = [col for col in destination_plate.cols('1', length=num_strips)]
# 7 to 12 as written, will change based on num_strips
mix2_cols = [
col
for col
in destination_plate.cols(12 - num_strips, length=num_strips)]
# pcr strips location
p10 = instruments.P10_Single(mount='right', tip_racks=tiprack_10)
p10.pick_up_tip()
p10.transfer(Vol,
water,
dest_plate.wells(dest_well),
blow_out=True,
new_tip='never')
p10.drop_tip()
i = 1
tiprack_10_2 = labware.load('tiprack-10ul', '8')
p10_8 = instruments.P10_Multi(mount='left',
max_volume=10,
tip_racks=tiprack_10_2)
##Change the number 2 to the volume of DNA we would like to add.
for i in range(12):
p10_8.pick_up_tip(tiprack_10_2.cols(i))
p10_8.aspirate(2, gDNA_plate.cols(i))
p10_8.dispense(2, dest_plate.cols(i))
p10_8.drop_tip()
#p10_8.transfer(8,gDNA_plate.wells('A1'),dest_plate.wells('A1'))
robot.home()
for c in robot.commands():
print(c)
from opentrons import labware, instruments
source = labware.load('96-deep-well', '11', 'source')
p10rack = labware.load('tiprack-10ul', '10', 'p10rack')
p10 = instruments.P10_Multi(
tip_racks=[p10rack],
mount="left",
)
# target_slots = ['D1', 'E1', 'A2', 'A3', 'B2', 'C2', 'D2', 'D3', 'E2', 'E3']
# plate_type = 'PCR-strip-tall'
# HACK: need to explicitly load each container like this
# instead of using a for loop, so that deck map can be parsed out
# for protocol library
# targets = [
# c o n t a i n e r s.load(plate_type, slot) for slot in target_slots]
targets = [
labware.load('PCR-strip-tall', '1'),
labware.load('PCR-strip-tall', '2'),
labware.load('PCR-strip-tall', '3'),
labware.load('PCR-strip-tall', '4'),
labware.load('PCR-strip-tall', '5'),
labware.load('PCR-strip-tall', '6'),
labware.load('PCR-strip-tall', '7'),
labware.load('PCR-strip-tall', '8'),
labware.load('PCR-strip-tall', '9')
]
# print(targets)
def run_custom_protocol(
number_of_destination_plates: int = 5,
volume_of_mineral_oil_in_ul: float = 15,
distance_from_oil_surface_to_opening_of_trough_in_mm: float = 10):
# check for labware space
if number_of_destination_plates > 5:
raise Exception('Please specify 5 or fewer destination plates.')
# remaining labware
dest_plates = [
labware.load(plate_name, str(slot))
for slot in range(4, 4 + number_of_destination_plates)
]
tips50 = labware.load(tips50_name, str(4 + number_of_destination_plates))
tips10 = [
labware.load(tips10_name, str(slot))
for slot in range(5 + number_of_destination_plates, 12)
]
# pipettes
m10 = instruments.P10_Multi(mount='right', tip_racks=tips10)
m50 = instruments.P50_Multi(mount='left', tip_racks=[tips50])
all_dests = [well for plate in dest_plates for well in plate.rows('A')]
# variables for mineral oil height track
h_oil = -(distance_from_oil_surface_to_opening_of_trough_in_mm + 5)
### length of the trough to work out total volume
length = 71.4
width = dest_plates[0].wells(0).properties['diameter']
def oil_height_track():
nonlocal h_oil
dh = volume_of_mineral_oil_in_ul / (
length * width
) # should this be 8*volume_of_mineral_oil_in_ul for a trough?
h_oil -= dh
# transfer mineral oil
m50.set_flow_rate(aspirate=5, dispense=10)
t_count = 0
m50.pick_up_tip()
for d in all_dests:
# prevent oil buildup in the same tip (replace after each plate fill)
if t_count == 12:
m50.drop_tip()
m50.pick_up_tip()
t_count = 1
oil_height_track()
m50.aspirate(volume_of_mineral_oil_in_ul, mineral_oil.top(h_oil))
m50.delay(seconds=5)
m50.dispense(d.bottom(5))
t_count += 1
m50.blow_out()
m50.drop_tip()
# distribute PCR master mix
m50.set_flow_rate(aspirate=25, dispense=50)
m50.pick_up_tip()
for d in all_dests:
m50.transfer(18, master_mix, d.top(), blow_out=True, new_tip='never')
m50.drop_tip()
# forward primer distribution
for ind, primer in enumerate(forward_primer.rows('A')):
dests = [plate.rows('A')[ind] for plate in dest_plates]
m10.distribute(1, primer, dests)
# reverse primer distribution
for ind, primer in enumerate(reverse_primer.rows('A')):
dests = [plate.rows('A')[ind] for plate in dest_plates]
m10.distribute(1, primer, dests)
temp_deck2 = modules.load('tempdeck', '10')
temp_plate = labware.load('1ml_Covaris', '10', share=True)
#mag_deck = modules.load('magdeck', '7')
#mag_plate = labware.load('biorad-hardshell-96-PCR', '7', share=True)
tipracks_10 = [
labware.load('tiprack-10ul', slot, share=True) for slot in ['8']
]
tipracks_200 = [
labware.load('tiprack-200ul', slot, share=True) for slot in ['9']
]
#### PIPETTE SETUP ####
m10 = instruments.P10_Multi(mount='left',
min_volume=1,
max_volume=10,
tip_racks=tipracks_10)
m300 = instruments.P300_Multi(mount='right',
min_volume=30,
max_volume=200,
aspirate_flow_rate=100,
dispense_flow_rate=200,
tip_racks=tipracks_200)
## Enzyme SETUP
# Enzyme_ER = MM_plate.wells('A1')
# Enzyme_Lig = MM_plate.wells('A2')
Enzyme_Fill = MM_plate.wells('A3')
## Reagent SETUP
##Labware
#Tips
#p300rack = labware.load('opentrons_96_tiprack_300ul', '9')
p10rack1 = labware.load('tiprack-10ul', '4')
p10rack2 = labware.load('tiprack-10ul', '1')
#Plates
p384plate = labware.load('384-plate', '3')
#Samples
sample96plate = labware.load('96-PCR-tall', '2')
##Pippetes and tips definition
#p50 = instruments.P50_Single(mount='right', tip_racks=[p300rack])
p10m = instruments.P10_Multi(mount='right', tip_racks=[p10rack1, p10rack2])
##Commands
def run_custom_protocol():
for r in range(12):
# p10m.transfer(10, sample96plate.cols('1'), p384plate.cols(r).wells('A','C','E','G','I','K','M','O').bottom(5), new_tip='always', dispense_flow_rate=0.5, blow_out=True)
p10m.transfer(10,
sample96plate.cols('1'),
p384plate.cols(r).wells('A', 'C', 'E', 'G', 'I', 'K',
'M', 'O').bottom(5),
new_tip='always',
dispense_flow_rate=0.5)
for r in range(12):
p10m.transfer(10,
primer_plate = labware.load('PCR-strip-tall', '3', share=True)
#Col1 Primer stip 1 (e.g. 1-8)
#Col2 empty - for opening lids
#Col3 Primer stip 2 (e.g. 9-16)
#Col4 empty
#Col5 Primer stip 3 (e.g. 17-24)
#Col6 empty
#### TIP RACKS ####
tiprack_200 = labware.load('labsolute-tiprack-200µl', '5')
tiprack_10 = labware.load('labsolute-tiprack-10µl', '2')
#### PIPETTES ####
s50 = instruments.P50_Single(mount='left', tip_racks=[tiprack_200])
m10 = instruments.P10_Multi(mount='right', tip_racks=[tiprack_10])
#### INPUT FILES ####
#Load reagents data file
water = []
buffer = []
mgcl2 = []
bsa = []
dntp = []
taq = []
primers = []
samples = []
with open("/root/csv/pcr_mix_96plate_v1_reagents.csv", "r") as csvfile:
settings = csv.reader(csvfile, delimiter=',')
for i in settings:
for well in custom_plate.wells():
print(well)
############################ define labware
# pipette and tiprack
if pipette_type == 'p10-Single':
tipracks = [labware.load(tiprack_type, tiprack_slot) for tiprack_slot in tiprack_slots]
pipette = instruments.P10_Single(
mount=pipette_mount,
tip_racks=tipracks)
pipette.start_at_tip(tipracks[0].well(tip_start_from))
elif pipette_type == 'p10-Multi':
tipracks = [labware.load(tiprack_type, tiprack_slot) for tiprack_slot in tiprack_slots]
pipette = instruments.P10_Multi(
mount=pipette_mount,
tip_racks=tipracks)
pipette.start_at_tip(tipracks[0].cols(tip_start_from))
pipette.plunger_positions['drop_tip'] = -6
# container for the drugs
src_container = labware.load(source_type, source_slot)
# where water, op50, and dmso are in the source container
src_water = src_container.wells(source_H2O_well)
src_water = src_water.bottom(0.5)
# container for the agar-filled 96MWP
dst_container = labware.load(destination_type, destination_slot)
################### actions
plate_1 = labware.load('96-flat', '1')
plate_2 = labware.load('96-flat', '2')
plate_3 = labware.load('96-flat', '4')
plate_4 = labware.load('96-flat', '5')
plate_5 = labware.load('96-flat', '7')
plate_6 = labware.load('96-flat', '8')
"""
All four pipettes are loaded into the program, however the app will only
see pipettes that are currently being used.
"""
if pipette_type == 'p300_multi':
pip = instruments.P300_Multi(mount=mount, tip_racks=[tiprack1, tiprack2])
elif pipette_type == 'p10_multi':
pip = instruments.P10_Multi(mount=mount, tip_racks=[tiprack1, tiprack2])
elif pipette_type == 'p50_multi':
pip = instruments.P50_Multi(mount=mount, tip_racks=[tiprack1, tiprack2])
else:
raise ValueError("Incorrect pipette type.")
plates = [plate_1, plate_2, plate_3, plate_4, plate_5, plate_6]
volumes = [
VOLUME_ONE, VOLUME_TWO, VOLUME_THREE, VOLUME_FOUR, VOLUME_FIVE, VOLUME_SIX
]
"""
Helper functions
"""
'description':
'A simple test protocol that checks I can pipette into the same target well on multiple plates'
}
# Set labware to use
trough = labware.load('starlab-E2310-1200', '1')
#tubes = labware.load('opentrons-tuberack-2ml-eppendorf')
tips300 = labware.load('opentrons-tiprack-300ul', '2')
tips10 = labware.load('tiprack-starlab-S1181-3810', '3')
PCR1 = labware.load('starlab-E1403-5200', '4')
PCR2 = labware.load('starlab-E1403-5200', '5')
forward_primer = labware.load('starlab-E1403-0100', '10')
# set pipettes
pipette300 = instruments.P300_Multi(mount='left', tip_racks=[tips300])
pipette10 = instruments.P10_Multi(mount='right', tip_racks=[tips10])
# Define where the mastermix to be distributed is - this version is for bulk setup
master_mix = trough.wells('A3')
# Define where the mastermix to be distributed is - this version is for a single plate with a single channel
#master_mix = tubes.wells('A1')
# List the target plates
dest_plates = [PCR1, PCR2]
# Get a list of all the pipetting locations on every plate in dest_plates
all_dests = [well for plate in dest_plates for well in plate.rows('A')]
# dispence the PCR mastermix across both plates
pipette300.set_flow_rate(aspirate=25, dispense=50)
"""
from opentrons import labware, instruments, robot
####################### user intuitive parameters
tiprack_slot = '3'
tip_start_from = '1' # first nonempty tip in the tip rack
cols_left = 12 # full tiprack
############################ define labware
# pipette and tiprack
tiprack = labware.load('opentrons-tiprack-10ul', tiprack_slot)
# tiprack = labware.load('opentrons_96_tiprack_10ul', tiprack_slot)
pipette = instruments.P10_Multi(mount='left', tip_racks=[tiprack])
pipette.start_at_tip(tiprack.cols(tip_start_from))
pipette.plunger_positions['drop_tip'] = -6
#################### actions
# safety command
# pipette.drop_tip() # actually maybe it hurts the robot to try to drop a tip when there's none
for _ in range(cols_left):
pipette.pick_up_tip()
pipette.delay(seconds=5)
pipette.return_tip()
# print
for c in robot.commands():
volume=200) # as per manufacturer's website
print('Wells in 96WP PCR Thermo Fisher:')
for well in custom_plate.wells():
print(well)
############################ define labware
# i.e. translate user-friendly parameters into opentrons language
# pipette and tiprack
# multi channel
if multi_pipette_type == 'p10-Multi': # this is mostly a check as we don't own other multichannel pipettes (and to not have swapped single/multi)
tiprackdrugs = [labware.load(tiprackdrugs_type, slot) \
for slot in tiprackdrugs_slots]
pipette_multi = instruments.P10_Multi(mount=multi_pipette_mount,
tip_racks=tiprackdrugs)
pipette_multi.start_at_tip(tiprackdrugs[0].well(tiprackdrugs_startfrom))
pipette_multi.plunger_positions['drop_tip'] = -6
# faster dispense
pipette_multi.set_speed(dispense=pipette_multi.speeds['dispense'] * 4)
# I only associated the "drugs" tiprack to the pipette as this is the one I want to handle authomatically
# I'll manually handle pipetting water
# container for water
water_src_container = labware.load(H2O_source_type, H2O_source_slot)
water_src_well = water_src_container.wells(H2O_source_well)
# tiprack for water
tiprackwater = labware.load(tiprackH2O_type, tiprackH2O_slot)
# translate the drugs mapping dict in robot language
wells_mapping = {}
def run_custom_protocol(pipette_type: StringSelection(
'p300_Multi', 'p50_Multi', 'p10_Multi', 'p1000_Single', 'p300_Single',
'p50_Single', 'p10_Single') = 'p300_Multi',
pipette_mount: StringSelection('left',
'right') = 'left',
sample_number: int = 16,
PCR_volume: float = 20,
bead_ratio: float = 1.8,
elution_buffer_volume: float = 20):
incubation_time = 300
settling_time = 50
drying_time = 5
total_tips = sample_number * 8
tiprack_num = total_tips // 96 + (1 if total_tips % 96 > 0 else 0)
slots = ['3', '5', '6', '8', '9', '10', '11'][:tiprack_num]
if pipette_type == 'p1000_Single':
tipracks = [labware.load('tiprack-1000ul', slot) for slot in slots]
pipette = instruments.P1000_Single(mount=pipette_mount,
tip_racks=tipracks)
elif pipette_type == 'p300_Single':
tipracks = [labware.load('tiprack-200ul', slot) for slot in slots]
pipette = instruments.P300_Single(mount=pipette_mount,
tip_racks=tipracks)
elif pipette_type == 'p50_Single':
tipracks = [labware.load('tiprack-200ul', slot) for slot in slots]
pipette = instruments.P50_Single(mount=pipette_mount,
tip_racks=tipracks)
elif pipette_type == 'p10_Single':
tipracks = [labware.load('tiprack-10ul', slot) for slot in slots]
pipette = instruments.P10_Single(mount=pipette_mount,
tip_racks=tipracks)
elif pipette_type == 'p10_Multi':
tipracks = [labware.load('tiprack-10ul', slot) for slot in slots]
pipette = instruments.P10_Multi(mount=pipette_mount,
tip_racks=tipracks)
elif pipette_type == 'p50_Multi':
tipracks = [labware.load('tiprack-200ul', slot) for slot in slots]
pipette = instruments.P50_Multi(mount=pipette_mount,
tip_racks=tipracks)
elif pipette_type == 'p300_Multi':
tipracks = [labware.load('tiprack-200ul', slot) for slot in slots]
pipette = instruments.P300_Multi(mount=pipette_mount,
tip_racks=tipracks)
mode = pipette_type.split('_')[1]
if mode == 'Single':
if sample_number <= 5:
reagent_container = labware.load('opentrons-tuberack-2ml-screwcap',
'7')
liquid_waste = labware.load('trough-12row', '5').wells('A12')
else:
reagent_container = labware.load('trough-12row', '7')
liquid_waste = reagent_container.wells('A12')
samples = [well for well in mag_plate.wells()[:sample_number]]
samples_top = [well.top() for well in samples]
output = [well for well in output_plate.wells()[:sample_number]]
else:
reagent_container = labware.load('trough-12row', '7')
liquid_waste = reagent_container.wells('A12')
col_num = sample_number // 8 + (1 if sample_number % 8 > 0 else 0)
samples = [col for col in mag_plate.cols()[:col_num]]
samples_top = [well.top() for well in mag_plate.rows(0)[:col_num]]
output = [col for col in output_plate.cols()[:col_num]]
# Define reagents and liquid waste
beads = reagent_container.wells(0)
ethanol = reagent_container.wells(1)
elution_buffer = reagent_container.wells(2)
# Define bead and mix volume to resuspend beads
bead_volume = PCR_volume * bead_ratio
if mode == 'Single':
if bead_volume * sample_number > pipette.max_volume:
mix_vol = pipette.max_volume
else:
mix_vol = bead_volume * sample_number
else:
if bead_volume * col_num > pipette.max_volume:
mix_vol = pipette.max_volume
else:
mix_vol = bead_volume * col_num
total_vol = bead_volume + PCR_volume + 15
mix_voltarget = PCR_volume + 10
# Disengage MagDeck
mag_deck.disengage()
# Mix Speed
pipette.set_flow_rate(aspirate=180, dispense=180)
# Mix beads and PCR samples
for target in samples:
pipette.set_flow_rate(aspirate=180, dispense=180)
pipette.pick_up_tip()
# Slow down head speed 0.5X for bead handling
pipette.mix(25, mix_vol, beads)
max_speed_per_axis = {
'x': (50),
'y': (50),
'z': (50),
'a': (10),
'b': (10),
'c': (10)
}
robot.head_speed(combined_speed=max(max_speed_per_axis.values()),
**max_speed_per_axis)
pipette.set_flow_rate(aspirate=10, dispense=10)
pipette.transfer(bead_volume,
beads,
target,
air_gap=0,
new_tip='never')
pipette.set_flow_rate(aspirate=50, dispense=50)
pipette.mix(40, mix_voltarget, target)
pipette.blow_out()
max_speed_per_axis = {
'x': (600),
'y': (400),
'z': (100),
'a': (100),
'b': (40),
'c': (40)
}
robot.head_speed(combined_speed=max(max_speed_per_axis.values()),
**max_speed_per_axis)
pipette.drop_tip()
# Return robot head speed to the defaults for all axes
max_speed_per_axis = {
'x': (600),
'y': (400),
'z': (100),
'a': (100),
'b': (40),
'c': (40)
}
robot.head_speed(combined_speed=max(max_speed_per_axis.values()),
**max_speed_per_axis)
# Incubate beads and PCR product at RT for 5 minutes
robot.comment("Incubating the beads and PCR products at room temperature \
for 5 minutes. Protocol will resume automatically.")
pipette.delay(seconds=incubation_time)
# Engage MagDeck and Magnetize
robot._driver.run_flag.wait()
mag_deck.engage()
robot.comment("Delaying for " + str(settling_time) +
" seconds for beads to \
settle.")
pipette.delay(seconds=settling_time)
# Remove supernatant from magnetic beads
pipette.set_flow_rate(aspirate=25, dispense=120)
for target in samples:
pipette.transfer(total_vol,
target.bottom(0.7),
liquid_waste.top(),
blow_out=True)
# Wash beads twice with 70% ethanol
air_vol = pipette.max_volume * 0.1
for cycle in range(2):
pipette.pick_up_tip()
for target in samples_top:
pipette.transfer(185,
ethanol,
target,
air_gap=air_vol,
new_tip='never')
robot.comment("Delaying for 17 seconds.")
pipette.delay(seconds=17)
for target in samples:
if not pipette.tip_attached:
pipette.pick_up_tip()
pipette.transfer(195,
target.bottom(0.7),
liquid_waste.top(),
air_gap=air_vol,
new_tip='never')
pipette.drop_tip()
# Dry at RT
robot.comment("Drying the beads for " + str(drying_time) +
" minutes. Protocol \
will resume automatically.")
pipette.delay(minutes=drying_time)
# Disengage MagDeck
robot._driver.run_flag.wait()
mag_deck.disengage()
# Mix beads with elution buffer
if elution_buffer_volume / 2 > pipette.max_volume:
mix_vol = pipette.max_volume
else:
mix_vol = elution_buffer_volume / 2
for target in samples:
pipette.pick_up_tip()
pipette.transfer(elution_buffer_volume,
elution_buffer,
target,
new_tip='never')
pipette.mix(45, mix_vol, target)
pipette.drop_tip()
# Incubate at RT for 3 minutes
robot.comment(
"Incubating at room temperature for 3 minutes. Protocol will \
resume automatically.")
pipette.delay(minutes=3)
# Engage MagDeck for 1 minute and remain engaged for DNA elution
robot._driver.run_flag.wait()
mag_deck.engage()
robot.comment("Delaying for " + str(settling_time) +
" seconds for beads to \
settle.")
pipette.delay(seconds=settling_time)
# Transfer clean PCR product to a new well
for target, dest in zip(samples, output):
pipette.transfer(elution_buffer_volume,
target.bottom(1),
dest.top(),
blow_out=True)
# Disengage MagDeck
mag_deck.disengage()