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DEPARTMENT OF BIOLOGY FACULTY OF SCIENCE & MATHEMATICS UNIVERSITI PENDIDIKAN SULTAN IDRIS SBL 1023 LABORATORY TECHNIQUE

DEPARTMENT OF BIOLOGY FACULTY OF SCIENCE & MATHEMATICS UNIVERSITI PENDIDIKAN SULTAN IDRIS

SBL 1023 LABORATORY TECHNIQUE IN BIOCHEMISTRY

STUDENT’S NAME

NAZIRA BINTI NASLAMROS

LECTURER’S NAME

ASSOC.PROF SHAKINAZ DESA

MATRIC NO

E20161013757

LABORATORY

MP 13

Introduction

Bacteria, yeast and fungi, protozoa and viruses are known as microorganism. These are most commonly found microorganisms in the surrounding, capable of settling and living in various environments such as soil, water, plant and animal tissues also human body. Some simple methods include cultivation and microscopic techniques could be applied to identify and isolate most of microbiological culture.

When working in a microbiology laboratory, aseptic technique is fundamental to the safety and success of an experiment. Generally, aseptic techniques performed by scientist under sterile conditions to ensure that microbial contaminants not doing harm to the colleagues, supply or experimental cultures. One of the aseptic technique is streak plate. The streak plate method is a rapid qualitative isolation method commonly used for isolation of discrete colonies that require the number of organisms in the inoculums be reduced. Inoculation is performed from a sector in order to isolate a pure culture.

Also in this lab, we are going to determine the effect of handwashing on bacteria on thumb. People used their hands for variety of activities everyday so it is extremely easy to come in contact with microbes. Handwashing is thought to be effective for the prevention of pathogens transmission. However, there is no conclusion that handwashing with soap is more effective at reducing bacteria contamination than using water only.

The gram stain is a differential stain commonly used to differentiates bacteria on the basis of their cell wall structure.

The gram stain is key procedure in the identification of bacteria based on staining characteristics, and enable the bacteria to be examined using light microscope. Most bacteria can be divided into 2 groups based on the cell wall structure.

Gram staining positive cell walls have single membrane enclosed by thick, cross link peptidoglycan. The thick peptidoglycan takes up dye and appears in purple. Alcohol cannot wash out the dye-mordant complex because it firmly secured in the thick peptidoglycan layer. Saturated with crystal violet dye. The cell counter stain has no little to effect. Cell wall ranges in colour from mid to dark purple.

Gram staining negative cell wall have thin layer of peptidoglycan in the periplasmic space within its inner and outer lipid membrane. Cell wall takes up dye. It appears in purple. Dye and mordant complex, but does not adhere to the thin layer of peptidoglycan. It easily removed by alcohol and turn to colourless. It easy to take the counterstain. It colour with safranin and turn to pink.

Objective :

To produce isolated colonies of an organisms ( bacteria ) on an agar plate

using streak plate technique To determine the effect of handwashing using water, hand soap and hand

sanitizer on bacteria. To differentiate between the 2 major categories of bacteria which is gram

positive and gram negative. To understand how gram stain affects the gram (+) and gram (-) based on the structural differences of their cell wall

Result :

  • 1. Streak plate technique.

Result : 1. Streak plate technique. 2. Effect of handwashing using water, hand soap and hand
  • 2. Effect of handwashing using water, hand soap and hand sanitizer on bacteria.

Result : 1. Streak plate technique. 2. Effect of handwashing using water, hand soap and hand
  • 3. Gram staining

Result : 1. Streak plate technique. 2. Effect of handwashing using water, hand soap and hand

Discussion :

Aseptic techniques are performed to prevent contamination of the growth medium. Bacteria often occur in mixed populations and very rare to find a single species of bacteria. So, it is vital to divide the bacteria from the other species that originate in the environment so that we can study the cultural, physiological and morphological characteristic of the individual species. The streak plate method is commonly used as isolation method to grow bacteria on a media surface (agar plate) so that individual bacterial colonies are isolated and can be sampled.

When the selected culture media is inoculate using a single isolated colony, the resulting culture will grow from the selected single colony. This techniques is useful when we need to separate an organism in a mixed culture. It is also useful when we need to study the colony morphology of an organism because biochemical test are only valid when performed on pure culture. In this lab, we must use sterile tool because the crucial aspects is to maintain a sterile work area surrounding us.

Before we start the experiment, it is important to disinfect all of our work surface using 70% ethanol or any other disinfectant reagent. Sterile technique is absolutely necessary in many laboratory practical. It is a method that prevents foreign cells from getting into our plates and cultures. Generally, the key to successful experiment is to work quickly and efficiently otherwise, we will get a contaminant resulting culture.

To differentiate between the two major categories of bacteria which is gram positive and gram negative. S. aures are choose as experimental species. S.aures is a facultative anaerobic that can grow without need of oxygen and coccal (round) bacterium.

S. aures is not always pathogenic, it is a common cause of skin and respiratory infections also food poisoning. After we examined the bacteria under light microscope, we identifying the bacteria to be gram- positive bacteria as it stains purple in colour. Gram-positive differ from gram-negative through their cell walls structure and composition. Gram-positive bacteria have a thick cell walls made up of peptidoglycan layer beyond the plasma membrane which is cross linked by short chains of peptides. It is then resulting in the shape and rigidity of the cell wall. The thick layer of peptidoglycan allows to retain the dye-mordant complex (crystal violet and iodine). The alcohol cannot wash out the dye-mordant complex because it firmly secured in the thick layer of peptidoglycan. As a result the cells are stain as purple

Conclusion:

Staining is a technique used to enhance the clarify of the microscopic image. Gram

stain is a very important preliminary step in the classification and characterization of

bacteria. Gram stain procedure enables bacteria to retain the colour of stains based

on the differences in the composition of their cell walls. From this experiment, we can

differentiate between the two major categories (gram-positive and gram-negative).

We can obtain the individual colonies from a bacterial culture using streak

plate method. This is ideal for experimentation because the colonies are genetically

identical. When doing a microbiological laboratory, there are many aspects that we

need to pay attentions. Firstly, it is very crucial to maintain a sterile work area

surrounding us. This requires common sense and a continuous practices.

Reference :