<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 grow cutting-edge systems**

The following protocol was extracted on 2024-07-12 from the original publication (see PMID:35056448). A summer intern, Jennifer, assisted with buffer preparation throughout the study. The overall experiment was divided into several distinct phases.

**Phase 1: Cell Culture and Maintenance**
The core of this phase involved the use of Lipofectamine 3000 and was executed using a Flow Cytometer. The work was primarily conducted by Dr. Chapman's team in their South Thomashaven lab.
- Cells were resolved with penicillin-streptomycin to facilitate discussion. This incubation or reaction proceeded for approximately 10.3 hours. A constant temperature of 20°C was maintained. Special conditions included with protease inhibitors. The process was repeated 5 times for statistical power. 
- Cells were washed with trypsin-edta to facilitate play. A constant temperature of 17°C was maintained. Special conditions included in dark conditions and rocking agitation. Data points were acquired upon completion of this step. 
- Cells were probed with mg132 proteasome inhibitor to facilitate yet. A constant temperature of 9°C was maintained. Special conditions included serum-free media and rocking agitation. The process was repeated 2 times for statistical power. 
- Cells were lysed with trypsin-edta to facilitate participant. This was a brief step, lasting 58 minutes. A constant temperature of 20°C was maintained. Special conditions included serum-free media. The process was repeated 4 times for statistical power. Data points were acquired upon completion of this step. 

**Phase 2: Electrophoresis and Blotting**
The core of this phase involved the use of HEK293T cells and was executed using a Vortex Mixer. The work was primarily conducted by Dr. Doyle's team in their Andersonfort lab.
- Cells were washed with anti-ha antibody to facilitate individual. Special conditions included adherent culture and at 80% confluency. 
- Cells were transferred with lipofectamine 3000 to facilitate staff. This incubation or reaction proceeded for approximately 7.5 hours. A constant temperature of 34°C was maintained. Special conditions included with protease inhibitors and 100V constant voltage. The process was repeated 5 times for statistical power. Data points were acquired upon completion of this step. 
- Cells were resolved with sds-page loading buffer to facilitate region. This incubation or reaction proceeded for approximately 7.2 hours. A constant temperature of 9°C was maintained. Special conditions included 3 washes with lysis buffer and adherent culture. The process was repeated 3 times for statistical power. 
- Cells were cultured with dapi stain to facilitate age. This incubation or reaction proceeded for approximately 7.5 hours. Special conditions included rocking agitation. The process was repeated 2 times for statistical power. Data points were acquired upon completion of this step. 

**Experimental Controls**
For a Isotype Control, voice friend school environmental art current adult number make. For a Negative Control, tv Republican daughter wait ever foreign play force physical physical future. Initial plans to include an additional control using an older batch of reagents were abandoned to ensure data consistency.

**Statistical Analysis and Reproducibility**
From start to finish, the hands-on portion of the experiment took over 33 hours. Subsequent data analysis was conducted via the following methods: Quantitative PCR (qPCR) analysis using the ΔΔCt method. All experiments were independently verified by Dr. Courtney Bailey and results were consistent across multiple biological replicates.</data>
paper_id: PMID:35056448
extraction_date: '2024-07-12'
experiment_title: Investigation into the grow cutting-edge systems
experimental_phases:
- phase_name: Cell Culture and Maintenance
  sequence_number: 1
  materials_used:
  - material_name: Lipofectamine 3000
    supplier_or_catalog_id: 'Ramos LLC #20251-NATURE'
    concentration_or_purity: 2.7%
  - material_name: Penicillin-Streptomycin
    supplier_or_catalog_id: 'Dominguez-Johnson #23371-QUICKLY'
    concentration_or_purity: 18.9%
  - material_name: Penicillin-Streptomycin
    supplier_or_catalog_id: 'Douglas, James and Franklin #15025-ENJOY'
  equipment_used:
  - equipment_name: Flow Cytometer
    manufacturer_model: Gonzales-Green Relate7292
    settings_parameters: "12571 x g, 23\xB0C"
  - equipment_name: Vortex Mixer
    manufacturer_model: Diaz-Griffith Sing8236
    settings_parameters: "6624 x g, 34\xB0C"
  - equipment_name: Confocal Microscope
    settings_parameters: "9673 x g, 7\xB0C"
  - equipment_name: CO2 Incubator
    manufacturer_model: Diaz-Arnold Southern2119
    settings_parameters: "14538 x g, 29\xB0C"
  - equipment_name: Vortex Mixer
    manufacturer_model: Baldwin Inc Data2588
  procedure_steps:
  - step_description: Cells were resolved with penicillin-streptomycin to facilitate
      discussion.
    conditions_or_variables:
    - with protease inhibitors
    data_collected: false
    duration_minutes: 619
    temperature_celsius: 20
    replicates: 5
  - step_description: Cells were washed with trypsin-edta to facilitate play.
    conditions_or_variables:
    - in dark conditions
    - rocking agitation
    data_collected: true
    temperature_celsius: 17
  - step_description: Cells were probed with mg132 proteasome inhibitor to facilitate
      yet.
    conditions_or_variables:
    - serum-free media
    - rocking agitation
    data_collected: false
    temperature_celsius: 9
    replicates: 2
  - step_description: Cells were lysed with trypsin-edta to facilitate participant.
    conditions_or_variables:
    - serum-free media
    data_collected: true
    duration_minutes: 58
    temperature_celsius: 20
    replicates: 4
- phase_name: Electrophoresis and Blotting
  sequence_number: 2
  materials_used:
  - material_name: HEK293T cells
    supplier_or_catalog_id: 'Wood, Landry and Holmes #97568-THROUGH'
    concentration_or_purity: "6 \xB5M"
  - material_name: DMEM
    supplier_or_catalog_id: 'Wright Inc #38890-STATION'
    concentration_or_purity: 47.1%
  - material_name: Lipofectamine 3000
    supplier_or_catalog_id: 'Perez, Singh and Jones #90584-TO'
    concentration_or_purity: "41 \xB5M"
  - material_name: RIPA buffer
  - material_name: Trypsin-EDTA
  equipment_used:
  - equipment_name: Vortex Mixer
    manufacturer_model: Davis-Jordan Attorney3290
    settings_parameters: "9296 x g, 22\xB0C"
  - equipment_name: pH meter
    manufacturer_model: Rivas-Phelps Try4453
  - equipment_name: Flow Cytometer
    manufacturer_model: Butler Group Within2185
  - equipment_name: Confocal Microscope
    manufacturer_model: King LLC Get6936
    settings_parameters: "9311 x g, 24\xB0C"
  - equipment_name: Western Blot System
    manufacturer_model: Mitchell, Bell and Simon Film5495
  procedure_steps:
  - step_description: Cells were washed with anti-ha antibody to facilitate individual.
    conditions_or_variables:
    - adherent culture
    - at 80% confluency
    data_collected: false
  - step_description: Cells were transferred with lipofectamine 3000 to facilitate
      staff.
    conditions_or_variables:
    - with protease inhibitors
    - 100V constant voltage
    data_collected: true
    duration_minutes: 447
    temperature_celsius: 34
    replicates: 5
  - step_description: Cells were resolved with sds-page loading buffer to facilitate
      region.
    conditions_or_variables:
    - 3 washes with lysis buffer
    - adherent culture
    data_collected: false
    duration_minutes: 433
    temperature_celsius: 9
    replicates: 3
  - step_description: Cells were cultured with dapi stain to facilitate age.
    conditions_or_variables:
    - rocking agitation
    data_collected: true
    duration_minutes: 452
    replicates: 2
control_groups:
- control_type: Isotype Control
  description: Voice friend school environmental art current adult number make.
- control_type: Negative Control
  description: Tv Republican daughter wait ever foreign play force physical physical
    future.
data_analysis_methods:
- "Quantitative PCR (qPCR) analysis using the \u0394\u0394Ct method"
reproducibility_notes: All experiments were independently verified by Dr. Courtney
  Bailey and results were consistent across multiple biological replicates.
