Ethan Flint

How to do a Bacterial Streak Plate

Bacterial plating is one of the most widely used laboratory techniques in
Microbiology. Some of the purposes for plating include determination of bacterial
species, determination of bacterial count, and continuation of a bacterial culture.
Although various methods of bacterial plating exist, the following instruction set
technique will be describe the technique of streak plating.
Streak plating is used to isolate single colonies of bacteria. This isolation is
beneficial when trying to distinguish one bacterium from another if using a culture with
multiple specimens. The orientation of the streaking done on the agar plate is what causes
the isolation of single colonies. The purpose of this streaking exercise however, will be
the continuation of the bacterial culture because the original bacterial culture used is a
single species.

Important Items:
Agar Plate
Bacteria Culture
Inoculation Loop
Bunsen Burner
Incubator
Gloves
Fire Sparker

Troubleshooting tips:
When streaking the agar plate, take your time. Rushing can cause mistakes in aseptic
techniques and overlooking important steps.
With maintaining sterility in plating, it is beneficial to think yourself; What in my
technique can cause foreign bacteria from colonizing?
Plating involves a feel for the motion of loop on the plate. Tilting the inoculation loop
too vertical will stab the agar; too many zig-zags on the plate will not dilute the specimen
enough. Dont feel frustrated when plating for the first time, you experience problems.
NOTE:
It is critical that caution and attention for your surroundings must be taken when doing
streak plating. This means cleaning your desk before and after use with the proper solvents and
reducing cross contamination of foreign material (phones, bodily fluids, etc.). Most importantly the
use of gloves for protection is a necessity.


3. Flaming the inoculation loop
Place the loop in the bluest part of the flame.
This is the hottest part of the flame.
Hold loop until it turns a fiery orange color, to
ensure that the loop has been sterilized and no
foreign bacteria will mix with original colony
Cool down the loop for 5-10 seconds so the
heat does not kill the bacteria to be scraped.

1. Labeling the Plate
Near the edge, label the bottom of the clean
agar plate to be streaked with the following:
Your name, Date, Name of Culture
-By labeling the bottom of the plate, the
identification of the plate will still be
possible if an accidental loss of the top of
the petri dish occurs.

2. Turning on the Bunsen burner:
Turn gas lever until hissing comes from burner
Use sparker over Bunsen burner and ignite the
flame
Adjust the level of flame by turning the gas
lever to reduce risk of burn.

NOTE:
It is critical that caution and attention for your surroundings must be taken when doing
streak plating. This means cleaning your desk before and after use with the proper solvents and
reducing cross contamination of foreign material (phones, bodily fluids, etc.). Most importantly the
use of gloves for protection is a necessity.


4. Scraping the Original Culture

Take loop and scrape a single colony from
your original culture until most, if not the
entire colony is gone. Although it may not look
like much is on the loop, there are many cells
that will transfer

NOTE: Try not to scrape into agar, as this
increases the chance of flinging the specimen
into face

5. Streaking the New Agar Plate
Take loop with sample and place on edge of agar near any side.
As horizontally as possible, zig-zag the loop trying to not cross over streaks
and not going too far into the middle of the plate. This is the first streak; do
not continue streaking in a different direction just yet!
Look at Figure 1 for visual assistance.

Figure 1
NOTE:
It is critical that caution and attention for your surroundings must be taken when doing
streak plating. This means cleaning your desk before and after use with the proper solvents and
reducing cross contamination of foreign material (phones, bodily fluids, etc.). Most importantly the
use of gloves for protection is a necessity.


5. Streaking the Plate continued..
Flame the loop until it turns fiery orange. Wait 5-10 seconds for the loop to
cool down
Start the second streak from the edge of the plate near the end of the first
streak. (Shown in Figure 1) Maintain the same zig-zag technique as
previously used, trying not to cross over streaks and staying toward the edge
of the plate.
Repeat this same step for the third and final streak: DO NOT CROSS
OVER INTO THE FIRST STREAK. Doing so will ruin the dilution to
create single colonies
6. Reflaming of the loop
Reflame the loop in order to kill off any
residual bacteria left.

7. Incubating the plate
Place cultured agar plate, upside down
in incubator at 35
o
C. This is to prevent
condensation accumulating on the agar,
which can ruin the colony


NOTE:
It is critical that caution and attention for your surroundings must be taken when doing
streak plating. This means cleaning your desk before and after use with the proper solvents and
reducing cross contamination of foreign material (phones, bodily fluids, etc.). Most importantly the
use of gloves for protection is a necessity.




8. Observing the plate
Take the cultured agar plate out
of the incubator after 24-48 hrs.
The plate should contain single
colonies as circled in Figure 2.

Figure 2