MOL BIO LAB L5 (Midterms- Lab Set-up) Flashcards
The laboratory should be compatible with ________ to prevent contamination
mechanical barrier
This is the most common issue in the molecular laboratory
contamination
What area does the sample preparation room belong to?
Area 1
What area does the Amplification room belong to?
Area 3
What area does the Reagent Preparation room belong to?
Area 2
An amplification room/area where the gel electrophoresis room is located
Analysis of PCR Products
serves as a clean bench area
Dead airbox with UV light
An amplification room/area where the PCR room is located
PCR amplification
Room/ area in the molecular biology lab where biosafety cabinet (optional) is located
Nucleic acid Extraction Room/ Area
The only room/area in the molecular biology lab where there is positive air pressure
Reagent preparation room/area
Air pressure in Sample prep and Post amplification rooms/areas
Negative air pressure
Single entrance, reagents used for amplification should not be exposed to other areas
Reagent prep
Specimens should not be exposed to post-amplification work areas
Specimen prep
The size of each area should consider ________ for equipment and ________ needed for preparation
space; bench space
Alternative to Spatial Separation
- Class II biological safety cabinet
- Dedicated areas for each work phase
- Unidirectional
- Automated specimen processing station/closed tube amplification and detection system
State the other lab design considerations (at least three)
- Temperature and humidity requirements
- Exhaust ventilation
- Water quality
- Electric outlet
- Back-up power system
- Eyewash
- Ergonomic assessment
This is always included when checking for contamiantion
blank (no template) control
What is to be avoided when pipetting?
Production of aerosols
The PCR workstation is cleaned at every?
Start and end of the workday/run
The PCR workstation is cleaned using?
Uv light, 70% ethanol, fresh 10% sodium hypochlorite, DNA Away
This induces thymidine dimers and other modifications that render nucleic acid inactive as a template for amplification
Ultra-violet light irradiation
Work stations should be cleaned with __________, followed by removal with _________
10% sodium hypochlorite solution (bleach; ethanol and water
Substitution of uracil (dUTP) for thymine (dTTP) during PCR amplification
Enzymatic inactivation with uracil-N-glycosylase
Contaminating PCR amplicons are degraded leaving only genomic DNA avaialble for PCR
New PCR sample reactions pre-treated with Uracil-N-glycosylase (UNG)
Wipe test (swab test) is performed?
Monthly
This is used to detect, localize, and remove contamination and identify the source of the contamination
Wipe test (swab test)
Proficiency testing is performed?
Twice a year
Used for assessment of the Competence in Testing
Proficiency Testing
If specimens are not commercially available alternative proficiency testing program has to be?
established (specimen exchange etc.)
Addresses emergencies from an all-hazards
approach. This establishes policy-and guidance ensuring that critical functions continue and that personnel and resources are
relocated to an alternate facility in case of emergencies.
Continuity of Operations Plan (COOP Plan)
Uses of molecular laboratory in the field of clinical diagnostics
- Health
- Medicine
- Forensics
- Pharmaceutical Industry
- Biological warfare
- Drug Discovery
- PCR based Technology
- Fluorescence in situ Hybridization (FISH)
- Biochip
- Peptide Nuicleic acid (PNA)
11.Proteomic Technology : - Electrochemical Detection of DNA
Potential sources of contamination are?
- Cross contamination between specimens
- amplification product contamination
- laboratory surfaces
- ventilation ducts
- reagents/supplies and
- hair, skin, saliva, and clothes of laboratory personnel.
Contamination may cause?
- Incorrect results
- Require extensive-cleanup
- Loss of creditability
- Impact on financial and performance
Contamination can be controlled using proper?
- Laboratory-design
- Laboratory practices
- Chemical and enzymatic controls
Nucleic acids used be stored/ used in what temp?
4-8°C or -15 to -25°C
A fundamental skill in molecular biology, as it ensures that reagents
and samples are measured accurately and reliably
Precise pipetting
These pipettes are used for handling small volumes, typically in
the microliter range
Micropipette
They are essential for PCR, DNA quantification, and sample transfers.
Micropipette
These are used for transferring larger-volumes, typically in
the milliliter range. Often used for media preparation, cell
culture, and larger-scale molecular biology experiments.
Serological pipettes
Multichannel pipettes have multiple channels, usually _______, that allow for simultaneous pipetting of multiple samples. They are commonly used in high-throughput applications.
8 or 12
Two commonly used techniques in pipetting
Forward and reverse
The more traditional-and widely used technique
Forward pipetting
Effective when working with viscous or volatile samples
Reverse pipetting
This technique reduces the risk of over-pipetting and ensures the intended volume is accurately delivered
Reverse pipetting
Suitable for general sample transfers and dilutions
Forward pipetting
This technique is commonly used when precise volume measurements are required, such as in quantitative assays where even minor deviations can affect results
Forward pipetting
This technique is ideal for transferring sample with unique characteristics, like highly viscous substances or volatile solutions
Reverse pipetting
Often used in the preparation of high-concentration reagents and for DNA/RNA extractions
Reverse pipetting
Essential to maintain the accuracy and reproducibility of experiments.
pH measurement
Measures the hydrogen ion concentration in a solution
pH
Formula to calculate dilutions
(Concentration stock) X (volume of stock used) = (Concentration final) X (final volume)
Monitor all steps of analytical procedure
Quality Control Plan
Measurement to monitor and record specific activities as part of the quality
Quality Indicator
- Turn around time
- % of failed runs
- Population medium
- Calibrator parameters
- Graph to identify trend or shift
- Monitor frequency and acceptable range
