PRAK. KE-1C. ISOLASI AZOTOBACTER - BIOTEK PERTANIAN (Lab. Biotan Faperta Unpad)

Summary

This practical session demonstrates the isolation of non-symbiotic azotobacter fixative bacteria from corn rhizosphere soil. The process begins with preparing a sample in liquid Asbi media, followed by incubation. After incubation, rims that grow on the media are isolated onto solid Asbi media using a scratch method. The session also covers the aseptic technique, including heating equipment, isolating colonies, and transferring them to slant agar for further incubation. After 3-7 days of incubation, the resulting azotobacter isolates are observed for characteristics such as pigmentation, marking the completion of the isolation process.

Takeaways

• 😀 Proper preparation of corn rhizosphere soil in liquid ASBI media is key for isolating Azotobacter fixative bacteria.
• 😀 After preparing the sample, the soil should be inoculated into liquid ASBI media and incubated at 30°C for 5-7 days.
• 😀 Following incubation, the rims will grow on the surface of the media, which can then be isolated onto solid ASBI media.
• 😀 To prepare solid ASBI media, dissolve the media into sterile Petri dishes and allow it to freeze and cool.
• 😀 The pellicle should be isolated onto the solid media using the scratch method to ensure proper aseptic technique.
• 😀 The nozzle used for the scratch method should be heated until it glows, then cooled before applying it to the media.
• 😀 The scratch method involves dividing the Petri dish into three quadrants and scratching in a zigzag manner across each quadrant.
• 😀 After completing the scratching, heat the nozzle again until it glows and cool before continuing with each quadrant.
• 😀 Label the Petri dish after completing the inoculation and then incubate at 30°C for further bacterial growth.
• 😀 Once incubation is completed, isolated colonies can be transferred to slant agar to observe bacterial growth and pigmentation.
• 😀 After a 3-5 day incubation period on slant agar, pigmentation and growth will be observed, indicating successful isolation of Azotobacter bacteria.

Q & A

What is the purpose of isolating Azotobacter fixative bacteria in this experiment?

The purpose is to isolate non-symbiotic Azotobacter fixative bacteria from corn rhizosphere soil, which are capable of fixing nitrogen.

What is the first step in preparing the sample for bacterial isolation?

The first step is to prepare a sample of corn rhizosphere soil in liquid ASBI media, which is then inoculated and incubated.

At what temperature should the sample be incubated after inoculation?

The sample should be incubated at a temperature of 30°C for 5-7 days.

What happens during the incubation period in terms of bacterial growth?

During the incubation, a pellicle (a thin film of bacteria) will form on the surface of the media, indicating bacterial growth.

How is the pellicle transferred to solid ASBI media?

The pellicle is transferred to solid ASBI media using the scratch method, where the bacteria are inoculated onto the solid media with a sterile loop.

What is the purpose of the scratch method?

The scratch method is used to spread the inoculum across the surface of the solid media in a zigzag pattern, which helps isolate individual bacterial colonies.

What are the steps involved in the scratch method for inoculating the solid media?

In the scratch method, the inoculation loop is heated until it glows, cooled briefly, and then used to make zigzag scratches in three quadrants of the solid media plate.

How long should the inoculated solid media be incubated?

The inoculated solid media should be incubated at 30°C, and bacterial growth will be observed after a few days.

What should be done after observing bacterial growth on the Petri dish?

After observing bacterial growth, individual colonies should be separated and transferred to slant agar for further growth and analysis.

What is the final incubation step after transferring bacteria to slant agar?

After transferring the bacteria to slant agar, it should be incubated at 30°C for 3-5 days to allow for further colony growth and pigmentation.