Streak Plate Method : Microbial Technique

Microbiology Lab Essentials Techniques :

2)Streak Plate Technique :

✓Principle :

The techniques commonly used for isolation of discrete colonies initially require that the number of organisms in the inoculum be reduced.The resulting diminution of the population size ensures that, following inoculation, individual cells will be sufficiently far apart on the surface of the agar medium to separate the different species. The   following are techniques that can be used to accomplish this necessary dilution:

✓Materials :

i)Cultures :

24 to 48hour nutrient broth cultures of a mixture of one part  Serratia marcescens  and three parts  Micrococcus luteus  and a mixture of one part  Escherichia coli  and ten parts Micrococcus luteus. 

ii)Sources :

mixed cultures from the environ ment could include cultures from a tabletop, bathroom sink, water fountain, or inside of an incubator. Each student should obtain a mixed culture from one of the environmental sources listed above. 

ii)Media : 

Three Trypticase soy agar plates per designated student group for each inoculation technique to be performed. Equipment Microincinerator or Bunsen burner, inoculating loop, turntable, glassware marking pencil, culture tubes containing 1 ml of sterile water, test tube rack, and sterile cotton swabs.

#The  streak-plate  method is a rapid qualitative isolation method. It is essentially a dilution technique that involves spreading a loopful of culture over the surface of an agar plate. Although many types of procedures are per formed, the #fourway, or #quadrant, streak  is described. Refer to above Figure which schematically illustrates this technique.

i)Place a loopful of culture on the agar surface in Area 1. Flame the loop, cool it by touching an unused part of the agar surface close to the periphery of the plate, and then drag it rapidly several times across the surface of Area 1.
ii)Reflame and cool the loop, and turn the Petri dish 90°.Then touch the loop to a corner of the culture in Area 1 and drag it several times across the agar in Area 2.The loop should never enter Area 1 again.
iii)Reflame and cool the loop and again; turn the dish 90°.Streak Area 3 in the same manner as Area 2.  Without reflaming go the loop, again turn the dish 90° and then drag the culture from a corner of Area 3 across Area 4, using a wider streak. Don’t let the loop touch any of the previously streaked areas.
iv)The flaming of the loop at the points indicated is to dilute the culture so that fewer organisms are streaked in each area, resulting in the final desired separation.A photograph of a streak plate inoculation is shown in the following figure.

An alternative streakplate method is for students new to the laboratory who have
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yet to master the necessary lab skills that would allow them to use the rapid method listed above. This alternative method involves spreading a loop ful of culture over the surface of an agar plate that has the quadrants laid out visibly for quick reference. Refer figure shown above which illustrates this technique.
a)Using a marker, draw two bisecting lines on the bottom of the Petri dish to divide the plate into 4 equal parts. Label each quadrant 1 through 4, starting with the top right quadrant and labeling counter clockwise. Sterilizing the loop at the points indicated is to dilute the culture due to fewer organisms available to be streaked into each area, resulting in the final desired separation.
b)Turn the Petri dish over and place a loopfull of culture on the agar surface in quadrant 1.  Using the edge of the loop and holding the loop at a shallow angle so as not to gouge the agar, quickly spread the bacteria throughout the quadrant.
c)Reflame and cool the loop, and turn the Petri dish 90°.Then touch the loop into an area that has been streaked in quadrant 1 and drag it across the agar into quadrant 2, repeat this twice without flaming the loop.
d)Reflaming and cool the loop and again turn the dish 90°.Streak the bacteria into   quadrant 3 in the same manner used for quadrant 2. e.  Reflame and cool the loop and again turn the dish 90°. Streak the bacteria into quadrant 4 in the same manner used for quadrant 3.
In the Following figure after incubation onverted form the results can observe as Follow 👇
You can Also observe that IV quadrant is having isolated Colonies so thats why this isolation of bacterial pure colonies or single Colony can be picked up .