Principle Microbiology

GFP-Photo reaction:

· The GFP absorps mainly at 396 nm, followed by loss of proton in excited state to form phenolate.

· The main emmision occurs at 508 nm. Small amount of GFP absorps at 475 nm and emits green light at 508 nm.

· The GFP as well as green fluoroscent gene can be modified. The gene can be modified by mutation.

· The fluoroscent may be partially or completely lost. The mutation gives some misfolding of protein that generally fails to produce fluoroscence, but sometimes also produce higher emission.

Green Fluoroscent Protein:

· The GFP was first discovered from the jellyfish Aequorea victoria. But it is also present in other organisms.

· The Aequorea victoria contains two luminescent proteins.

1) Aequorin

2) GFP, i.e. Green Fluoroscent Protein

· Aequorin, when interacts with Ca+2, it emits flashes of the blue light.

· The GFP aquires energy from Aequorin and emits green light.

Structure of GFP:

· The GFP from Aequorea victoria has an 11 stranded beta-barrel structure, with a alpha-helix running up the axis of the barrel.

· The chromophore is attached to alpha-helix in the center of barrel, few amino acids make the chromophore.

· The chromophore has Serine, Tyrosine and Glycine at the position 65; 66 and 67 respectively.

· 4-(p-hydroxybenzylidene)-imidazolidin-5-one attached to protein backbone through 1 and 2 position of the ring.

· Chromophore has hydrogen-bonding with amino acid residue and water molecules.

Vaccines are any preparation which are made up of either whole microorganisms, macromolecules of microorganisms, DNAs or Recombinant Vector which can induce immunity in the host animal when given to it.

Vaccines are produced in many ways, most traditional of which is the embryonated hen's egg technique. But nowadays the place of embryonated hen's egg is being taken by the use of animal cell lines. Animal cell lines have various advantages over the other traditional techniques. Moreover animal cell lines also overcome some of the traditional problems faced during older techniques. There are also some limitations of the use of animal cell lines.

The whole article on the Use of animal cell lines in vaccine production can be read here as a powerpoint presentation on Google documents.

http://docs.google.com/leaf?id=0B0c57-C3kwyuOTdkZGVmZjMtZDA2YS00YjU4LTk2ZmYtZDY1NDg0YjU0ZTkz&hl=en

It contains following points...

What is a Vaccine?
Historical notes
Types of vaccines
Why cell lines?
Potential advantages
Ideal characteristics of cell lines used in vaccine production
Considerations Related to Specific Cell Lines
Production steps

I hope this article and the powerpoint helps you in one or another way...

- TRUNAL YATINDRA GUPTE

Recently one of my friend Nilesh Vaghela prepared a seminar on the topic Acid rain. I've uploaded his powerpoint presentation on my blog. I hope it'll help you people in one or other way. The seminar has covered following topics about Acid rain.
- Introduction to presenter and guide
- Definition
- Acid deposition
- Formation of Acid rain
- Measurement of Acid rain
- Effects of Acid rain
- Control measure of Acid rain
- References
- Thank you

The link to the powerpoint present on web is given below...

http://docs.google.com/present/view?id=dg4jc3qb_948jb7ng6f

The magnetic stirrer is used in many biological labs, including microbiology labs also. A magnetic stirrer is a laboratory device consisting of either a rotating magnet or stationary electromagnets creating a rotating magnetic field. This device is used to cause a stir bar immersed in a liquid to spin very quickly, agitating or mixing the liquid. A magnetic stirrer often includes a provision for heating the liquid. Stirrers are often used in laboratories, especially in the field of biology and microbiology. They are preferred over gear-driven motorized stirrers because they are quieter, more efficient, and have no moving external parts to break or wear out (other than the simple bar magnet itself).

Due to its small size, a stirring bar is more easily cleaned and sterilized than other stirring devices. Magnetic stirrers avoid two major problems with motorized stirrers. Firstly, motorized stirrers use lubricants, which can contaminate the reaction vessel and the product. Secondly, in motorized stirrers, the sealing of the connection between the rotating shaft of the stirrer and the vessel can be problematic, especially if a closed system is needed.

Magnetic stirrers also have drawbacks. For example, the limited size of the stirring bar means it can only be used for relatively small (under 4 liters) experiments. In addition, viscous liquids or thick suspensions are extremely difficult to mix using this method, although there are some stirrers with special magnets to overcome this problem.

See the following pictures on the top of the post:
<1> The magnetic stirrer
<2> The bar magnets used in stirrer

PCR principles and procedure

PCR is used to amplify specific regions of a DNA strand (the DNA target). This can be a single gene, a part of a gene, or a non-coding sequence. Most PCR methods typically amplify DNA fragments of up to 10 kilo base pairs (kb), although some techniques allow for amplification of fragments up to 40 kb in size.

A basic PCR set up requires several components and reagents.

These components include:

* DNA template that contains the DNA region (target) to be amplified.

* Two primers that are complementary to the 3' (three prime) ends of each of the sense and anti-sense strand of the DNA target.

* Taq polymerase or another DNA polymerase with a temperature optimum at around 70 °C.

* Deoxynucleoside triphosphates (dNTPs; also very commonly and erroneously called deoxynucleotide triphosphates), the building blocks from which the DNA polymerases synthesizes a new DNA strand.

* Buffer solution, providing a suitable chemical environment for optimum activity and stability of the DNA polymerase.

* Divalent cations, magnesium or manganese ions; generally Mg2+ is used, but Mn2+ can be utilized for PCR-mediated DNA mutagenesis, as higher Mn2+ concentration increases the error rate during DNA synthesis.

* Monovalent cation potassium ions.


The PCR is commonly carried out in a reaction volume of 10–200 μl in small reaction tubes (0.2–0.5 ml volumes) in a thermal cycler. The thermal cycler heats and cools the reaction tubes to achieve the temperatures required at each step of the reaction (see below). Many modern thermal cyclers make use of the Peltier effect which permits both heating and cooling of the block holding the PCR tubes simply by reversing the electric current. Thin-walled reaction tubes permit favorable thermal conductivity to allow for rapid thermal equilibration. Most thermal cyclers have heated lids to prevent condensation at the top of the reaction tube. Older thermocyclers lacking a heated lid require a layer of oil on top of the reaction mixture or a ball of wax inside the tube.

Mary Mallon (September 23, 1869 – November 11, 1938), also known as Typhoid Mary, was the first person in the United States to be identified as a healthy carrier of typhoid fever. Over the course of her career as a cook, she is known to have infected 53 people, three of whom died from the disease. Her notoriety is in part due to her vehement denial of her own role in spreading the disease, together with her refusal to cease working as a cook. She was forcibly quarantined twice by public health authorities and died in quarantine. It is possible that she was born with the disease, as her mother had typhoid fever during her pregnancy.

Mallon was born in 1869 in County Tyrone, Northern Ireland, and emigrated to the United States in 1884. She worked as a cook in the New York City area between 1900 and 1907. She had been working in a house in Mamaroneck, New York for less than two weeks when the residents came down with typhoid. She moved to Manhattan in 1901 and members of the family for whom she worked developed fevers and diarrhea and the laundress died. She then went to work for a lawyer until seven of the eight household members developed typhoid. Mary spent months helping to care for the people she made sick, but her care further spread the disease through the household. In 1906, she took a position in Long Island. Within two weeks, six out of eleven family members were hospitalized with typhoid. She changed employment again and three more households were infected.

People catch typhoid fever after ingesting food or water which has been contaminated during handling by a human carrier. The human carrier is usually a healthy person who has survived a previous episode of typhoid fever but in whom the typhoid bacteria have been able to survive without causing further symptoms. Carriers continue to excrete the bacteria in their feces and urine. It takes vigorous scrubbing and thorough disinfection with soap and hot water to remove the bacteria from the hands. When typhoid researcher George Soper approached Mallon with the news she was possibly spreading typhoid, she adamantly rejected his request for urine and stool samples to ascertain whether she was a typhoid carrier. Soper left and later published his findings in the June 15, 1906 issue of the Journal of the American Medical Association.[1] On his next contact with her, he brought a doctor with him, but was again turned away. Mallon's denials that she was a carrier were based in part on the diagnosis of a reputable chemist who had found she was not harboring the bacteria. It is possible she was in temporary remission when tested. Moreover, when Soper first told her she was a carrier, the concept that a person could spread disease and remain healthy was not well known. During a later encounter in the hospital, he told Mary he would write a book about her and give her all the royalties; she angrily rejected his proposal and locked herself in the lavatory until he left.

Swine influenza (also called swine flu, hog flu and pig flu) refers to influenza caused by those strains of influenza virus, called swine influenza virus (SIV), that usually infect (is endemic in) pigs. As of 2009 these strains are all found in Influenza C virus and the subtypes of Influenza A virus known as H1N1, H1N2, H3N1, H3N2, and H2N3. Swine influenza is common in pigs in the midwestern United States (and occasionally in other states), Mexico, Canada, South America, Europe (including the United Kingdom, Sweden, and Italy), Kenya, Mainland China, Taiwan, Japan and other parts of eastern Asia. Transmission of swine influenza virus from pigs to humans is not common and does not always cause human influenza, often only resulting in the production of antibodies in the blood. The meat of the animal poses no risk of transmitting the virus when properly cooked. If transmission does cause human influenza, it is called zoonotic swine flu. People who work with pigs, especially people with intense exposures, are at increased risk of catching swine flu. In the mid-20th century, identification of influenza subtypes became possible, this allows accurate diagnosis of transmission to humans. Since then, fifty confirmed transmissions have been recorded, Rarely, these strains of swine flu can pass from human to human. In humans, the symptoms of swine flu are similar to those of influenza and of influenza-like illness in general, namely chills, fever, sore throat, muscle pains, severe headache, coughing, weakness and general discomfort. The 2009 flu outbreak in humans, known as "swine flu", is due to a new strain of influenza A virus subtype H1N1 that contains genes closely related to swine influenza. The origin of this new strain is unknown. However, the World Organization for Animal Health (OIE) reports that this strain has not been isolated in pigs. This strain can be transmitted from human to human, and causes the normal symptoms of influenza. Pigs can become infected with human influenza, and this appears to have happened during the 1918 flu pandemic and the 2009 flu outbreak.

The main raw materials used in beer production are as follows...


Malt
Malt adjuncts
Hops
Water

Malt :- The malt is prepared from carefully selected barley. This barley is first cleaned and then steeped in water for period up to two days. The excess water is then drained and the soaked barley is further incubated for periods of approximately four to six days to allow formation of a short rootlet and acrospire. This germination step allows the formation of highly viscous α - amylase, β - amylase and proteolytic enzymes, as well as flavor and color components.

At the end of incubation, the temperature is just raised to stop the germintaion without harming any enzyme, although high temperatures can be employed to obtain dark colored stout and bock beer fermentation.

Hence the green malt produced is carefully dried and stored. The preparation of good malt is an exacting task and it requires careful selection of barley and close supervision of malting process.
So most of the beer producing companies do not produce their own malt. But they rely on other companies who specialize in this art.

The other raw materials needed in beer production well be described soon.

The differential count gives us information on the number of different types of leukocytes in per 100 leukocytes.

The smear is made on the slide. First of all a drop is put on a slide. Then with the help of another slide the smear is prepared. The cells are counted one by one. No special slide like counting chamber or dilution fluids.

The cells are differentiated on the basis of nuclear lobes they contain. And the staining capacities of granules present in the cytoplasm.