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A question about microbes growing media


I was told in my microbiology course that blood agar can be put in GasPak system (anearobic) to grow strict anaerobes, but why would someone use blood-dependent bacteria in this experiment as these blood-dependent bacteria will be aerobes or at least facultative


You can put any media in a GasPak or other anaerobic system depending the type of anaerobic growth you are looking for. Sometimes blood is simply added to media to meet nutritional requirements of an organism. I use Columbia Blood agar often as a general purpose anaerobic medium. In a collection of 100+ obligate anaerobes I am working with, around half grow well on columbia agar with 5% sheeps blood, and many of those will not grow on Columbia without this blood amendment. As an added bonus, I can easily spot certain common contaminants growing on my plates by their hemolysis paterns.


Blood agar is prepared by heating the blood so that there will be lysis of RBC. Blood agar is mostly used to grow pathogenic organisms such as Haemophilus influenzae and Neisseria gonorrhoeae which cannot grow on other media.Neisseria gonorrhoeae can grow in the absence of oxygen. Haemophilus influenzae which is a facultative anaerobe also can carry out anaerobic respiration when there is no oxygen.Blood agar is very important to grow fastidious organisms, i.e the organism which is capable to lyse the RBC.


Growth of Microorganisms in Meat | Microbiology

In this article we will discuss about the growth of microorganisms in meat.

Due to high moisture, rich in nitrogenous food, plentified supply of minerals and accessory growth factors usually has some fermentable carbohydrate (glycogen) and a favourable pH allows most of the microbes to grow.

The following factors influence the growth of microorganism:

(a) The kind and amount of contamination with microorganisms and the spread of these organisms in the meat,

(b) Physical properties of meat,

(c) Chemical properties of meat,

(d) Availability of oxygen, and

Fish and other Sea Food:

Various bacteria involved in the spoilage are part of the natural flora of the external slime of fishes and their intestinal contents. At higher temperature Micrococcus and Bacillus species involved in fish spoilage while at ordinary temperature, species of Escherichia, Proteus, Serratia, Sarcina and Clostridium are found.

Seafood includes fresh, frozen, dried, pickled and salted fish, as well as various shellfish. Fresh water fishes also get contamination from the surrounding microflora present in the water in which they live. The main genera that covers outer surfaces are: Pseudomonas, Alcaligenes, Micrococcus, Flavobacterium, Corynehacterium, Sarcina, Serratia, Vibrio, Salmonella and Bacillus species.

The presence of bacteria in the water content depends upon the temperature of water. Both psychrophiles and mesophiles are present in the water. Fresh water fish carry fresh water bacteria.

Boats, boxes, bins, fish ponds and fish houses including fisherman soon become heavily contami­nated with these bacteria and transfer them during cleaning. Most of the fishes pass large amount of water through their bodies pick up soil and water microorganism in this way, including pathogens if they are present.

The numbers of microorganisms on the skin of fish can be influenced by the method of catching. The other sources of contamination includes potatoes, spices and flavours used in fish cake. The microbial contents of fresh fish vary in the microbial fish products.

(i) Microbiology of Fish Brines:

The temperature of brine and salt concentration influences the bacterial activity. The kind and amount of bacteria also vary which result to fish spoilage. Contamination comes from the fish, which ordinarily introduces species of Pseudomonas, Alcali­genes and Flavobacterium. The continuous use of fish brine may contribute additional pathogens from successive lots of fish and because of salt tolerant bacteria (micrococci, etc.).

Autolysis, oxidation or bacterial activity spoil fish like meat. The fish flesh is autolysed more quickly due to the presence of fish enzymes and because of the less acid reaction of fish flesh that favours microbial growth.

Many of the fish oils seem to be more susceptible to oxida­tive deterioration than most animal fats. The lower the pH of fish flesh, the slower in general bacterial decomposi­tion. Lowering the pH of fish results from the conversion of muscle glycogen to lactic acid.

(iii) Factors Influencing Kind and Rate of Spoilage

(a) The kind of fish:

The various kinds of fish differ considerably in their perishability. Certain fatty fish deteriorate rapidly because of oxidation of unsaturated fats of their oils.

(b) The condition of fish when caught:

Fish full of food when caught, are more readily perishable than those with an empty intestinal tract.

(c) The kind and extent of contamination of the fish flesh with bacteria:

These may come from mud, water handlers, exterior slime, and the intestinal content of the fish and are supposed to enter the gill of the fish, from which they pass through the vascular system and thus invade the flesh, or to penetrate the intestinal tract and thus enter the body cavity.

In general, the greater the load of bacteria, the more rapid the spoilage. The contamination may take place in the net, in the fishing boat, on the docks, or later, in the plants. This process is accelerated by the digestive enzymes attacking and perforating the gut wall and viscera, which in themselves have a high rate of autolysis.

Bacterial growth is delayed at lower temperature. Cooling or chilling should be as rapid as possible 0 to -1°C. The high temperature reduce the life of fish. The prompt and rapid freezing of the fish is more effective in preservation.

(e) Use of an Antibiotic Ice or Dip:

Some antibiotics are recommended to avoid spoilage.

(iv) Characteristics of Spoilage:

The bright colours of fish fade and dirty, yellow-brown discolouration appear. The slime on the skin of fish increases, especially on the flaps and gills. The eyes gradually sink, and oh shrinkage the pupil becomes cloudy and the cornea opaque.

Flesh becomes soft and juice erodes when squeezed. The discolouration takes place towards tail due to oxidation of haemoglobin and the different types of odours evolved during spoilage and cooking will bring out these odours more strongly.

(v) Control of Spoiling Microorganisms:

Keeping microorganisms away from meats is called Asepsis. This process begins with avoidance from contamination. Before slaughtering, the animal should be carefully washed with water so as to remove dust from hair and hoof.

The knife may also introduce microorganism in the circulating blood. During evisceration contamination may come from the internal body parts like intestine, the air, the water, cloths, and brushes used on the carcass. Some organism may come from the surface soil and from workers.

Once meat is contaminated with microbes their removal is difficult. The use of hot water or sanitizer sprays under pressure is a procedure of decreasing the bacterial number. Moldy or spoiled portion of large piece of meat may be trimmed off.

Meats have been reported to have a shorter storage life in films with less permeable to water. Cured meats are packed in an oxygen- tight film with evacuation. It checks the growth of aerobes especially molds, reduces the rate of growth of staphylococci.

Canning of meat differs from product to product to be preserved. Most meat products are low-acid foods and are good culture media. Rates of heat penetration range from fairly effective in meat soups to very slow in tightly packed meats or in paste. Various additives such as spices, chemical salts and flavour also affect the heat processing. The process becomes more effective.

On the basis of heat processing, canned meat can be divided into two groups:

(a) Meat that are processed in one attempt (shelf-stable canned meat), and

(b) Meat that are heated enough to kill part of the spoilage organisms but must be kept refrigerated to prevent spoilage. This type of meat is known as non-shelf stable. The shelf-stable meat is processed at 98°C and the size of container is usually less than a kilogram. Cured meat temperature for processing should be 65°C and the container used during packing is of up to 22 kg.

Hot water treatment is also a method to remove the microbes from meat surfaces. But this may lessen nutrients and can damage colour. Heat applied during the smoking of meat and meat products helps in reducing microbes.

The cooking of meats for direct consumption greatly reduces the microbial content and hence lengthen the keeping time. Precooked frozen meat should contain few viable microbes. More meat is preserved by the use of low temperature either by chilling or freezing. Chilling is more common.

Meat can be preserved promptly and rapidly to temperature near freezing and chilling at only slightly above the freezing point. Meat may be held in chilling storage for a limited time with little change from their original constitution. Enzymatic and microbial changes in the foods are not prevented but are slowed down considerably. Cooler temperature prevents growth but slow metabolic activity may continue.

The storage time can be prolonged in an atmosphere containing added CO2 and O3. Ships equipped for storage of meat in a controlled atmosphere of CO2 have been employed successfully. Increasing amounts of CO2 inhibit microbial growth but also enhance the formation of met-myoglobin resulting into loss of colour. Storage time can be increased by the pressure of 2.5 to 3 ppm of ozone in the atmosphere.

Ozone is an active oxidizing agent, that may give an oxidized or tallowy flavour to fats. Few bacteria, molds and yeasts can grow in meats at low temperature are known as psychrotrophic bacteria (Sta­phylococcus, Alcaligenes, Micrococcus, Leuconostoc, Flavobacterium and Proteus).

(b) Freezing or Frozen Storage:

Under the usual conditions of storage of frozen foods microbial growth is prevented entirely and action of food enzymes is greatly retarded. The lower the storage temperature the slower will be any chemical or enzymatic reaction. The preservation of frozen meats is increasingly effective as the storage temperature drops from -12.2°C to -28.9°C.

The freezing process kills the bacteria. The rate of freezing of meat and other food items depends upon a number of factors, such as the temperature, circulation of air, or refrigerant, size and shape of package, kind of food etc. Sharp freezing refers to freezing in air with only natural air circulation or at best with electric fans. The freezing temperature is usually -23.3°C or lower but may vary from -15 to -29°C, and may take from 3 to 72 hours.

This is called slow freezing. Quick freezing is accomplished by one of the three methods:

(a) Direct immersion in a refrigerant,

(b) Indirect contact where the meat is in contact with the passage through which the refrigerant at -17.8 to -45.6°C flows, and

(c) Air-blast freezing where frigid air at -17.8 to -34.4°C is blown across the materials being frozen. Certain items now are being frozen into liquid nitrogen.

The advantages of quick freezing are:

(a) Smaller ice crystals formation and less destruction of intact cells of the food,

(b) A shorter period of sodification and therefore, less time for diffusion of soluble materials and for separation of ice,

(c) More prompt preservation of microbial growth, and

(d) More rapid enzyme action.

The ultraviolet rays serve to reduce number of microorganisms in the air and to inhibit or kill them on the surfaces of the meat reached directly by the rays. Irradiation also is used in the rapid aging of meats at higher than the usual chilling temperature to reduce the growth of microorganisms, especially molds, on the surface. Some oxidation, favoured by UV rays, and hydrolysis of fats may take place during aging.

Some types of sausages are preserved primarily by their dryness. In dried beef, made mostly from cured, smoked beef hams, growth of microorganism may take place before processing and may develop in the “pickle” during curing, but numbers of organism are reduced by the smoking and drying process.

Organisms may contaminate the dried ham during storage and the slices during cutting and packing. Salting and smoking are usually employed during meat drying.

Another method of drying pork involves a short addition of lecithin as an antioxidant and stabilizer. Drying may be in vacuum in trays, or by other methods. The meat for drying should be of good bacteriological quality.


Bacterial Division

Bacteria and archaea reproduce asexually only, while eukartyotic microbes can engage in either sexual or asexual reproduction. Bacteria and archaea most commonly engage in a process known as binary fission, where a single cell splits into two equally sized cells. Other, less common processes can include multiple fission, budding, and the production of spores.

The process begins with cell elongation, which requires careful enlargement of the cell membrane and the cell wall, in addition to an increase in cell volume. The cell starts to replicate its DNA, in preparation for having two copies of its chromosome, one for each newly formed cell. The protein FtsZ is essential for the formation of a septum, which initially manifests as a ring in the middle of the elongated cell. After the nucleoids are segregated to each end of the elongated cell, septum formation is completed, dividing the elongated cell into two equally sized daughter cells. The entire process or cell cyclecan take as little as 20 minutes for an active culture of E. coli bacteria.


Generation Time

In eukaryotic organisms, the generation time is the time between the same points of the life cycle in two successive generations. For example, the typical generation time for the human population is 25 years. This definition is not practical for bacteria, which may reproduce rapidly or remain dormant for thousands of years. In prokaryotes (Bacteria and Archaea), the generation time is also called the doubling time and is defined as the time it takes for the population to double through one round of binary fission. Bacterial doubling times vary enormously. Whereas Escherichia coli can double in as little as 20 minutes under optimal growth conditions in the laboratory, bacteria of the same species may need several days to double in especially harsh environments. Most pathogens grow rapidly, like E. coli, but there are exceptions. For example, Mycobacterium tuberculosis, the causative agent of tuberculosis, has a generation time of between 15 and 20 hours. On the other hand, M. leprae, which causes Hansen’s disease (leprosy), grows much more slowly, with a doubling time of 14 days.

Calculating Number of Cells

It is possible to predict the number of cells in a population when they divide by binary fission at a constant rate. As an example, consider what happens if a single cell divides every 30 minutes for 24 hours. The diagram in Figure 3 shows the increase in cell numbers for the first three generations.

The number of cells increases exponentially and can be expressed as 2 n , where n is the number of generations. If cells divide every 30 minutes, after 24 hours, 48 divisions would have taken place. If we apply the formula 2 n , where n is equal to 48, the single cell would give rise to 2 48 or 281,474,976,710,656 cells at 48 generations (24 hours). When dealing with such huge numbers, it is more practical to use scientific notation. Therefore, we express the number of cells as 2.8 × 10 14 cells.

In our example, we used one cell as the initial number of cells. For any number of starting cells, the formula is adapted as follows:

Nn is the number of cells at any generation n, N0 is the initial number of cells, and n is the number of generations.

Figure 3. The parental cell divides and gives rise to two daughter cells. Each of the daughter cells, in turn, divides, giving a total of four cells in the second generation and eight cells in the third generation. Each division doubles the number of cells.

Think about It

  • With a doubling time of 30 minutes and a starting population size of 1 × 10 5 cells, how many cells will be present after 2 hours, assuming no cell death?

Culture Media

Culture media are a combination of pre-defined ingredients that support the growth and proliferation of microorganisms in an artificial/in vitro environment. They are used for the isolation of organisms from various samples such as clinical specimens, food, or the environment.

Based on the consistency, culture media are further categorized as solid, semisolid, and liquid (broth). Functionally, culture media can be classified as differential, selective, enriched, enrichment, assay, transport, etc.

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Calling Startups: There’s A Big Niche to Fill

How can synthetic biology eliminate the screening bottleneck to unlock the potential of novel organisms? One way is to completely skip screening a broad range of organisms and take a more targeted approach instead.

“I think there’s a market for a company that could systematically explore the vast diversity of [microbes] on Earth,” muses Solomon. Such an approach would be critical half-joking, Solomon says that many enzymes used now are essentially “random strains that people found on the bottom of someone’s shoe.” Instead, he envisions a catalog of “all the weird organisms” and their properties, which could be licensed out to researchers.

“There are literally billions of species on this planet. There’s no reason why synthetic biology should be limited to E.coli and yeast and other emerging strains,” he projects. With such an expanded search space available, one of the few remaining questions is where the next transformative microbe may be found.


Complex and Synthetic Media

In defined media all the chemical compounds are known, while undefined media has partially unknown chemical constituents.

Learning Objectives

Differentiate complex and synthetic medias

Key Takeaways

Key Points

  • Defined media is made from constituents that are completely understood.
  • Undefined media has some part of which is not entirely defined.
  • The presence of extracts from animals or other microbes makes a media undefined as the entire chemical composition of extracts are not completely known.

Key Terms

  • recombinant: This term refers to something formed by combining existing elements in a new combination. Thus, the phrase recombinant DNA refers to an organism created in the lab by adding DNA from another species.
  • serum: The clear yellowish fluid obtained upon separating whole blood into its solid and liquid components after it has been allowed to clot. Also called blood serum.

There are many types of culture media, which is food that microbes can live on. Two major sub types of media are complex and synthetic medias, known as undefined and defined media.

Undefined Media: Luria Broth as shown here is made with yeast extract, as yeast extract is not completely chemically defined Luria Broth is therefore an undefined media. By Lilly_M [GFDL (http://www.gnu.org/copyleft/fdl.html), CC-BY-SA, via Wikimedia Commons

An undefined medium has some complex ingredients, such as yeast extract or casein hydrolysate, which consist of a mixture of many, many chemical species in unknown proportions. Undefined media are sometimes chosen based on price and sometimes by necessity – some microorganisms have never been cultured on defined media.A defined medium (also known as chemically defined medium or synthetic medium) is a medium in which all the chemicals used are known, no yeast, animal, or plant tissue is present. A chemically defined medium is a growth medium suitable for the culture of microbes or animal cells (including human) of which all of the chemical components are known. The term chemically defined medium was defined by Jayme and Smith as a ‘Basal formulation which may also be protein-free and is comprised solely of biochemically-defined low molecular weight constituents.

A chemically defined medium is entirely free of animal-derived components (including microbial derived components such as yeast extract) and represents the purest and most consistent cell culture environment. By definition chemically defined media cannot contain either fetal bovine serum, bovine serum albumin, or human serum albumin as these products are derived from bovine or human sources and contain complex mixes of albumins and lipids. The term ‘chemically defined media’ is often misused in the literature to refer to serum albumin-containing media. Animal serum or albumin is routinely added to culture media as a source of nutrients and other ill-defined factors, despite technical disadvantages to its inclusion and its high cost. Technical disadvantages to using serum include the undefined nature of serum, batch-to-batch variability in composition, and the risk of contamination. There are increasing concerns about animal suffering inflicted during serum collection that add an ethical imperative to move away from the use of serum wherever possible. Chemically defined media differ from serum-free media in that bovine serum albumin or human serum albumin with either a chemically defined recombinant version (which lacks the albumin associated lipids) or synthetic chemical such as the polymer polyvinyl alcohol which can reproduce some of the functions of serums.


III. CULTURING BACTERIA

A. Methods of Obtaining Pure Cultures (a culture that contains only 1 species of organism)

1. The Streak Plate Method – Bacteria are picked up on a sterile wire loop, and the wire is moved lightly along the agar surface, depositing streaks of bacteria on the surface. The loop is flamed and a few bacteria are picked up from the region already deposited and streaked onto a new region. Fewer and fewer bacteria are deposited as the streaking continues, and the loop is flamed after each streaking. Individual organisms (individual cells) are deposited in the region streaked last. After the plate is incubated at a suitable growth temperature for the organism, small colonies (each derived from a single bacterial cell) appear. The loop is used to pick up a portion of an isolated colony and transfer it to another medium for study. The use of aseptic technique assures that the new medium will contain organisms of a single species. We’ll do this in lab.


Principle:


The increase in the cell size and cell mass during the development of an organism is termed as growth. It is the unique characteristics of all organisms. The organism must require certain basic parameters for their energy generation and cellular biosynthesis. The growth of the organism is affected by both physical and Nutritional factors. The physical factors include the pH, temperature, Osmotic pressure, Hydrostatic pressure, and Moisture content of the medium in which the organism is growing. The nutritional factors include the amount of Carbon, nitrogen, Sulphur, phosphorous, and other trace elements provided in the growth medium. Bacteria are unicellular (single cell) organisms. When the bacteria reach a certain size, they divide by binary fission, in which the one cell divides into two, two into four and continue the process in a geometric fashion. The bacterium is then known to be in an actively growing phase. To study the bacterial growth population, the viable cells of the bacterium should be inoculated on to the sterile broth and incubated under optimal growth conditions. The bacterium starts utilising the components of the media and it will increase in its size and cellular mass. The dynamics of the bacterial growth can be studied by plotting the cell growth (absorbance) versus the incubation time or log of cell number versus time. The curve thus obtained is a sigmoid curve and is known as a standard growth curve. The increase in the cell mass of the organism is measured by using the Spectrophotometer. The Spectrophotometer measures the turbidity or Optical density which is the measure of the amount of light absorbed by a bacterial suspension. The degree of turbidity in the broth culture is directly related to the number of microorganism present, either viable or dead cells, and is a convenient and rapid method of measuring cell growth rate of an organism. Thus the increasing the turbidity of the broth medium indicates increase of the microbial cell mass (Fig 1) .The amount of transmitted light through turbid broth decreases with subsequent increase in the absorbance value.

Fig 1: Absorbance reading of bacterial suspension

The growth curve has four distinct phases (Fig 2)

1. Lag phase

When a microorganism is introduced into the fresh medium, it takes some time to adjust with the new environment. This phase is termed as Lag phase, in which cellular metabolism is accelerated, cells are increasing in size, but the bacteria are not able to replicate and therefore no increase in cell mass. The length of the lag phase depends directly on the previous growth condition of the organism. When the microorganism growing in a rich medium is inoculated into nutritionally poor medium, the organism will take more time to adapt with the new environment. The organism will start synthesising the necessary proteins, co-enzymes and vitamins needed for their growth and hence there will be a subsequent increase in the lag phase. Similarly when an organism from a nutritionally poor medium is added to a nutritionally rich medium, the organism can easily adapt to the environment, it can start the cell division without any delay, and therefore will have less lag phase it may be absent.

2. Exponential or Logarithmic (log) phase

During this phase, the microorganisms are in a rapidly growing and dividing state. Their metabolic activity increases and the organism begin the DNA replication by binary fission at a constant rate. The growth medium is exploited at the maximal rate, the culture reaches the maximum growth rate and the number of bacteria increases logarithmically (exponentially) and finally the single cell divide into two, which replicate into four, eight, sixteen, thirty two and so on (That is 2 0 , 2 1 , 2 2 , 2 3 . 2 n , n is the number of generations) This will result in a balanced growth. The time taken by the bacteria to double in number during a specified time period is known as the generation time. The generation time tends to vary with different organisms. E.coli divides in every 20 minutes, hence its generation time is 20 minutes, and for Staphylococcus aureus it is 30 minutes.

3. Stationary phase

As the bacterial population continues to grow, all the nutrients in the growth medium are used up by the microorganism for their rapid multiplication. This result in the accumulation of waste materials, toxic metabolites and inhibitory compounds such as antibiotics in the medium. This shifts the conditions of the medium such as pH and temperature, thereby creating an unfavourable environment for the bacterial growth. The reproduction rate will slow down, the cells undergoing division is equal to the number of cell death, and finally bacterium stops its division completely. The cell number is not increased and thus the growth rate is stabilised. If a cell taken from the stationary phase is introduced into a fresh medium, the cell can easily move on the exponential phase and is able to perform its metabolic activities as usual.

4. Decline or Death phase

The depletion of nutrients and the subsequent accumulation of metabolic waste products and other toxic materials in the media will facilitates the bacterium to move on to the Death phase. During this, the bacterium completely loses its ability to reproduce. Individual bacteria begin to die due to the unfavourable conditions and the death is rapid and at uniform rate. The number of dead cells exceeds the number of live cells. Some organisms which can resist this condition can survive in the environment by producing endospores.

Fig 2: Different phases of growth of a bacteria

CALCULATION:


The generation time can be calculated from the growth curve(Fig 3).

Fig 3: Calculation of generation time

The exactly doubled points from the absorbance readings were taken and, the points were extrapolated to meet the respective time axis.


Generation Time = (Time in minutes to obtain the absorbance 0.4) &ndash (Time in minutes to obtain the absorbance 0.2)

Let No = the initial population number


N = the number of generations in time t



The growth rate can be expressed in terms of mean growth rate constant (k), the number of generations per unit time.



Mean generation time or mean doubling time (g), is the time taken to double its size.

Substituting equation 4 in equation 3


(Since the population doubles t= g)

Mean growth rate constant,
Mean generation time,


Coupling Specific Genes to Specific Organisms Using PCR

PCR allows for the amplification and mutation of DNA and allowing researchers to study very small samples.

Learning Objectives

Describe how polymerase chain reaction (PCR) allows for the amplification and mutation of DNA and enables researchers to study very small samples

Key Takeaways

Key Points

  • PCR allows for identification of an infectious agent without the need for culturing.
  • Researchers can use PCR as a method of searching for specific genes and/or mutations.
  • PCR, coupled with other biochemical techniques, allows us to analyze the very core of organisms and the processes by which they function.

Key Terms

  • polymerase chain reaction: A technique in molecular biology for creating multiple copies of DNA from a sample used in genetic fingerprinting etc.

Polymerase chain reaction (PCR) is a useful technique for scientists, because it allows for the amplification and mutation of DNA. Through PCR, small quantities of DNA can be replicated by orders of magnitude, not only essentially preserving the sample if successful, but allowing for study on a much larger scale.. Without PCR, the studies we perform would be limited by the amount of DNA we were able to isolate from samples. Through PCR, the original DNA is essentially limitless, allowing scientists to induce various mutations in different genes for further study.

Polymerase chain reaction: a schematic of the polymerase chain reaction

Through site-directed mutagenesis or customized primers, individual mutations in DNA can be made. By changing the amino acids transcribed from DNA through individual mutations, the importance of those amino acids with respect to gene function can be analyzed. However, this process can be difficult, particularly when genes act in concert (with varying expression with respect to gene activity). The length of time it takes to run a successful PCR and perform other techniques before additional studies can be done (protein expression, isolation, and purification, for example), makes biochemical research time-consuming and difficult. However, PCR, coupled with other biochemical techniques, allows us to analyze the very core of organisms and the processes by which they function. Common PCR protocols in labs today include knockout genotyping, fluorescence genotyping and mutant genotyping. Researchers can use PCR as a method of searching for genes by using primers that flank the target sequence of the gene along with all other necessary components for PCR. If the gene is present, the primers will bind and amplify the DNA, giving a band of amplified DNA on the agarose gel that will be run. If the gene is not present, the primers will not anneal and no amplification will occur.

The ability to identify specific genes to specific organisms has increased the use of PCR and has allowed it to be more specific and eliminate the possibility of cross contaminants. The identification of specific genes to specific organisms has important medical diagnostic value.

PCR is a reliable method to detect the presence of unwanted genetic materials, such as infections and bacteria in the clinical setting. It can even allow identification of an infectious agent without culturing. For example, in diagnosis of diseases like AIDS, PCR can be used to detect the small percentage of cells that are infected with HIV by utilizing primers that are specific for genes specialized to the HIV virus. PCR can reveal the presence of HIV in people who have not mounted an immune response to this pathogen, which may otherwise be missed with an antibody assay). Additionally, PCR is used for identifying bacterial species, such as Mycobacterium tuberculosis in tissue specimens. With the use of PCR, as few as 10 bacilli per million human cells can be readily detected. The bacilli are identified by using Mycobacterium tuberculosis specific genes.


Watch the video: Microbiology - Lawn (January 2022).