Introduction
There are countless varieties of microorganisms in different habitats. The majority of the time, those who engage with the bacteria don't know anything about them. Thus, the identification procedure is essential to understanding the microorganism being studied as well as to determining whether a certain microbe has the potential to cause disease or provide a research or commercial advantage (Maxwell, 2017). The identification procedure is used for academic purposes to determine the identity of an unidentified sample or to verify the identity of an organism that has already been identified. The establishment and separation of possible pollutants depend on the industrial identification of unknown microorganisms. In clinical settings identification are used in establishing various pathogens that are existing in various environments (Benson, 2002). Identification of unknown microorganisms is thus a significant aspect of study especially regarding the study of microorganisms in the laboratory.
Characteristics for Identifying Microorganisms
Microorganisms like all other living organisms possess special characteristics and features that are distinct from others (Valentine et al., 2005). It is thus significant that specific behaviors and bodily features are used in the process of identifying an organism that is not known. The diversity of the characteristics of the microorganisms is thus utilized as the basis for identification of various samples in the microbial laboratory. Some of the specific characteristics that are used for identification include the ability to grow on certain substrates; production of particular pigments and enzymes and others. Some of the techniques and tools applied in identifying unknown microorganisms include dichotomous keys with Gram stains after isolating the specimen of interests (Johnson & Case, 2015). The present pre-lab report discusses the isolation/separation as a step in the identification of unknown microorganisms.
Goals of the Lab Exercise
To describe and apply the procedure necessary for isolating and identifying a particular unknown organism
To use the learned microbiological techniques to identify an unknown microorganism in a sample and to apply a step-wise manner that leads to the identification of the specimen under study.
Procedure
Isolation/Separation
The goal of the isolation/separation step in week 1 is to isolate the unknown microorganism from the soil using serial dilution.
Serial Dilution Protocol
Sample is received and the unknown number is immediately recorded in a notebook
Dilution is performed in the order of 10-2, 10-4, 10-5, and 10-6 as described in the dilution scheme that is provided in the diagram. Different 1ml pipettes are used in a particular dilution under consideration.
To ensure a homogenous mixture, the unknown sample is vortexed and 0.1ml of the unknown sample is transferred into a culture tube containing 9.9ml sterile water leading to a 10-2 or 1/100 dilution.
The second dilution is performed in the same manner leading to 10-4 as the dilution is made from 10-2. Subsequent dilutions are made by pipetting 1ml from 10-4 and 10-5 dilutions and added to 9ml sterile water to attain 10-5, and 10-6 respectively.
In each of the dilution procedures, the sample is vortexed to ensure homogenous mixture is attained.
The dilution scheme is then followed by spread plating. Spread plating is done to isolate the organisms from each other.
The 10-4, 10-5, and 10-6 are vortexed one at a time to ensure a homogenous mixture. Using 1ml pipette 0.1ml of each solution is transferred into a TSA plate and labeled according to the dilution from which the sample is pipetted such as 10-4.
In total there should be three plates labeled 10-4, 10-5, and 10-6 according to the serial dilution scheme.
The plate spreader is sterilized by dipping it in ethanol and flaming it in each case of the spreading procedure.
The TSA plates are then inverted and incubated at 37°C.
References
Benson, H. J. (2002). Microbiological applications: laboratory manual in general microbiology. McGraw-Hill Science, Engineering & Mathematics.
Johnson, T. R., & Case, C. L. (2015). Laboratory experiments in microbiology. Pearson.
Maxwell, R. (2017). Lab Experiment 7: Microbiological Media. Retrieved 03 March 2017 from https://gsu.edu/lab1.pdf.
Valentine, N., Wunschel, S., Wunschel, D., Petersen, C., & Wahl, K. (2005). Effect of culture conditions on microorganism identification by matrix-assisted laser desorption ionization mass spectrometry. Applied and environmental microbiology, 71(1), 58-64.
