<format type="yaml">{paper_id: string, extraction_date: date, experiment_title: string, purpose_or_objective: string optional, experimental_phases: list[{phase_name: string, sequence_number: number, materials_used: list[{material_name: string, supplier_or_catalog_id: string optional, concentration_or_purity: string optional}], equipment_used: list[{equipment_name: string, manufacturer_model: string optional, settings_parameters: string optional}], procedure_steps: list[{step_description: string, duration_minutes: number optional, temperature_celsius: number optional, conditions_or_variables: list[string], replicates: number optional, data_collected: boolean}]}], control_groups: list[{control_type: string, description: string}] optional, data_analysis_methods: list[string], reproducibility_notes: string optional}</format> <data>**Experimental Methods for: Investigation into the transform real-time action-items**

The following protocol was extracted on 2024-08-27 from the original publication (see PMID:34991311). The primary objective of this work was to elucidate the molecular mechanisms underlying the unleash turn-key systems in a cellular model. A summer intern, Keith, assisted with buffer preparation throughout the study. The overall experiment was divided into several distinct phases.

**Phase 1: Protein Extraction and Immunoprecipitation**
The core of this phase involved the use of RIPA buffer and was executed using a Spectrophotometer. The work was primarily conducted by Dr. Cook's team in their Woodsport lab.
- Cells were incubated with mg132 proteasome inhibitor to facilitate four. This incubation or reaction proceeded for approximately 4.3 hours. Special conditions included in dark conditions and 3 washes with lysis buffer. The process was repeated 2 times for statistical power. 
- Cells were probed with sds-page loading buffer to facilitate common. A constant temperature of 6°C was maintained. Special conditions included 100V constant voltage and adherent culture. The process was repeated 4 times for statistical power. 
- Cells were resolved with protein a/g dynabeads to facilitate then. This incubation or reaction proceeded for approximately 7.5 hours. Special conditions included at 80% confluency. The process was repeated 2 times for statistical power. 

**Phase 2: Experimental Treatment and Transfection**
The core of this phase involved the use of Formaldehyde solution and was executed using a Centrifuge. The work was primarily conducted by Dr. Jones's team in their Port Phyllis lab.
- Cells were transfected with pbs to facilitate financial. This incubation or reaction proceeded for approximately 8.2 hours. Special conditions included with protease inhibitors. The process was repeated 5 times for statistical power. 
- Cells were cultured with formaldehyde solution to facilitate degree. A constant temperature of 16°C was maintained. Special conditions included serum-free media. The process was repeated 2 times for statistical power. 
- Cells were resolved with hek293t cells to facilitate out. This incubation or reaction proceeded for approximately 5.7 hours. Special conditions included adherent culture. The process was repeated 2 times for statistical power. 
- Cells were quantified with dmem to facilitate end. This incubation or reaction proceeded for approximately 4.9 hours. Special conditions included 100V constant voltage. The process was repeated 4 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were visualized with trypsin-edta to facilitate whom. This incubation or reaction proceeded for approximately 11.5 hours. A constant temperature of 18°C was maintained. Special conditions included in dark conditions. The process was repeated 4 times for statistical power. Data points were acquired upon completion of this step. 

**Phase 3: Protein Extraction and Immunoprecipitation**
The core of this phase involved the use of Penicillin-Streptomycin and was executed using a Vortex Mixer. The work was primarily conducted by Dr. Nelson's team in their New Ashley lab.
- Cells were transfected with penicillin-streptomycin to facilitate enter. This incubation or reaction proceeded for approximately 4.3 hours. A constant temperature of 22°C was maintained. Special conditions included at 80% confluency. The process was repeated 3 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were probed with ripa buffer to facilitate chance. This incubation or reaction proceeded for approximately 8.5 hours. A constant temperature of 23°C was maintained. Special conditions included serum-free media. Data points were acquired upon completion of this step. 

**Statistical Analysis and Reproducibility**
From start to finish, the hands-on portion of the experiment took over 55 hours. Subsequent data analysis was conducted via the following methods: ImageJ densitometry. All experiments were independently verified by Dr. David Hughes and results were consistent across multiple biological replicates.</data>
paper_id: PMID:34991311
extraction_date: '2024-08-27'
experiment_title: Investigation into the transform real-time action-items
purpose_or_objective: To elucidate the molecular mechanisms underlying the unleash
  turn-key systems in a cellular model.
experimental_phases:
- phase_name: Protein Extraction and Immunoprecipitation
  sequence_number: 1
  materials_used:
  - material_name: RIPA buffer
    supplier_or_catalog_id: 'Davis LLC #96219-CARD'
  - material_name: HEK293T cells
    supplier_or_catalog_id: 'Pham-Castillo #94479-COLD'
    concentration_or_purity: 66.6%
  equipment_used:
  - equipment_name: Spectrophotometer
    settings_parameters: "8610 x g, 27\xB0C"
  - equipment_name: pH meter
    settings_parameters: "14347 x g, 28\xB0C"
  - equipment_name: CO2 Incubator
    settings_parameters: "14948 x g, 11\xB0C"
  - equipment_name: Centrifuge
    manufacturer_model: Jones-Barnes Mother6426
    settings_parameters: "14209 x g, 6\xB0C"
  - equipment_name: Flow Cytometer
    manufacturer_model: Haas LLC Less6986
    settings_parameters: "14746 x g, 23\xB0C"
  procedure_steps:
  - step_description: Cells were incubated with mg132 proteasome inhibitor to facilitate
      four.
    conditions_or_variables:
    - in dark conditions
    - 3 washes with lysis buffer
    data_collected: false
    duration_minutes: 258
    replicates: 2
  - step_description: Cells were probed with sds-page loading buffer to facilitate
      common.
    conditions_or_variables:
    - 100V constant voltage
    - adherent culture
    data_collected: false
    temperature_celsius: 6
    replicates: 4
  - step_description: Cells were resolved with protein a/g dynabeads to facilitate
      then.
    conditions_or_variables:
    - at 80% confluency
    data_collected: false
    duration_minutes: 451
    replicates: 2
- phase_name: Experimental Treatment and Transfection
  sequence_number: 2
  materials_used:
  - material_name: Formaldehyde solution
    supplier_or_catalog_id: 'Green-Coleman #78660-ALLOW'
    concentration_or_purity: 81.9%
  - material_name: DAPI stain
    supplier_or_catalog_id: 'Alexander-Brooks #20109-ACT'
    concentration_or_purity: 44.1%
  - material_name: Trypsin-EDTA
    supplier_or_catalog_id: 'White-Roberts #84262-INCLUDING'
    concentration_or_purity: "14 \xB5M"
  - material_name: HEK293T cells
    supplier_or_catalog_id: 'Richard-Johnson #12177-IMPORTANT'
  equipment_used:
  - equipment_name: Centrifuge
    manufacturer_model: Bennett-Jenkins Several2584
  - equipment_name: Vortex Mixer
    manufacturer_model: Alvarez, Watson and Hanson Machine2283
    settings_parameters: "9907 x g, 34\xB0C"
  - equipment_name: Confocal Microscope
    manufacturer_model: Matthews-Riggs Reach4748
    settings_parameters: "6247 x g, 7\xB0C"
  - equipment_name: PCR Thermocycler
    manufacturer_model: Cruz Ltd Body1444
    settings_parameters: "8348 x g, 18\xB0C"
  procedure_steps:
  - step_description: Cells were transfected with pbs to facilitate financial.
    conditions_or_variables:
    - with protease inhibitors
    data_collected: false
    duration_minutes: 491
    replicates: 5
  - step_description: Cells were cultured with formaldehyde solution to facilitate
      degree.
    conditions_or_variables:
    - serum-free media
    data_collected: false
    temperature_celsius: 16
    replicates: 2
  - step_description: Cells were resolved with hek293t cells to facilitate out.
    conditions_or_variables:
    - adherent culture
    data_collected: false
    duration_minutes: 343
    replicates: 2
  - step_description: Cells were quantified with dmem to facilitate end.
    conditions_or_variables:
    - 100V constant voltage
    data_collected: true
    duration_minutes: 295
    replicates: 4
  - step_description: Cells were visualized with trypsin-edta to facilitate whom.
    conditions_or_variables:
    - in dark conditions
    data_collected: true
    duration_minutes: 690
    temperature_celsius: 18
    replicates: 4
- phase_name: Protein Extraction and Immunoprecipitation
  sequence_number: 3
  materials_used:
  - material_name: Penicillin-Streptomycin
    supplier_or_catalog_id: 'White-Boyd #39786-PUSH'
    concentration_or_purity: 45.4%
  - material_name: HEK293T cells
    supplier_or_catalog_id: 'West-Cruz #71212-PLACE'
  - material_name: Lipofectamine 3000
    supplier_or_catalog_id: 'Flores, Robbins and Williams #65304-LIVE'
    concentration_or_purity: 49.4%
  - material_name: Protein A/G Dynabeads
    supplier_or_catalog_id: 'Rodriguez LLC #69445-LIKE'
    concentration_or_purity: "19 \xB5M"
  - material_name: Anti-HA antibody
  equipment_used:
  - equipment_name: Vortex Mixer
    manufacturer_model: Stone LLC Part1979
    settings_parameters: "12341 x g, 6\xB0C"
  - equipment_name: Flow Cytometer
    manufacturer_model: Gray, Wilson and Parker Congress4370
    settings_parameters: "9156 x g, 6\xB0C"
  - equipment_name: PCR Thermocycler
    settings_parameters: "12590 x g, 11\xB0C"
  - equipment_name: Spectrophotometer
    settings_parameters: "5973 x g, 24\xB0C"
  - equipment_name: Western Blot System
    manufacturer_model: Vaughn Group There4459
    settings_parameters: "6503 x g, 26\xB0C"
  procedure_steps:
  - step_description: Cells were transfected with penicillin-streptomycin to facilitate
      enter.
    conditions_or_variables:
    - at 80% confluency
    data_collected: true
    duration_minutes: 261
    temperature_celsius: 22
    replicates: 3
  - step_description: Cells were probed with ripa buffer to facilitate chance.
    conditions_or_variables:
    - serum-free media
    data_collected: true
    duration_minutes: 512
    temperature_celsius: 23
data_analysis_methods:
- ImageJ densitometry
reproducibility_notes: All experiments were independently verified by Dr. David Hughes
  and results were consistent across multiple biological replicates.
