BIOL123

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Lab 4 Selective Media & Agar BIO250L”

Student Name: Click here to enter text. Access Code (located on the underside of the lid of your lab kit):

“Pre-Lab Questions”

1. What is the difference between chemically defined and chemically complex media? Give either a

clinical or environmental research example for which each media type would be the most

appropriate choice for culturing microorganisms.

In chemically defined media, the ingredients that make up the chemical media are know

and can be identified and quantity known such a medium is use to determine the least

amount of nutrients that a microbe needs to survive In chemically complex media the

chemical composition is unknown.the ingredients in this media have an organic origin for

instance, blood, and animal or human tissue can be added to a culture to enable a

pathogen to grow. The reason these organic ingredients are added is because the

pathogen is already know to grow in an animal or a human being

2. Why is differential media typically inoculated with isolated colonies that have been previously

cultured on general growth media?

Differential media is used in previously isolated colonies to identify specific microbes on

basis of the characteristics of that mircorogranism. One example is how agar can

differentiate between hemolytic and non hemolytic bacteria-

3. Use a textbook or a reputable internet source (such as www.cdc.gov) to research and describe a

scenario in a lab or clinical setting in which a selective and/or differential test would be

necessary.

In the case where a microbe is resistant to an antibiotic, selective test can and have

been used on a culture that is already resistant to anti-biotic.this allows for the cells that

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Lab 4 Selective Media & Agar BIO250L” have already grown a resistance to the antibiotic to be identified and marked. This was

done to cells that were resistant to neomycin. In a differential test, pin points the

presenceof a certain organism this makes it possible to eliminate this organism as soon

as it has been spoted https://bio.libretexts.org/Bookshelves/Microbiology/Book

%3A_Microbiology_(Boundless)/6%3A_Culturing_Microorganisms/6.3%3A_Culturing_B

acteria/6.3C%3A_Selective_and_Differential_Media

“EXPERIMENT 1: Bioprospecting for Starch Degrading Bacteria

Data Tables Table 3: Starch Agar and Gram Iodine Results

Sample Growth on Starch Agar? (Yes or No)

Clear area around colonies?

Do these bacteria contain amylase?

A Yes no y

B Yes yes- yes-

C Yes y- -yes

D No no no

Post-Lab Questions 1. Why is cow manure used as a potential source of starch-degrading bacteria? (If you are not

familiar with the process of digestion in cows, use a reputable internet source to inform your

answer.)

Because of the presence of Amylase which is needed to digest starch in the mouth

2. What are some other potential sources of starch-degrading bacteria?

potential sources of starch degrading bacteria can be found in fermenting food like hops

in the process of beer making, kombucha, and milk kefir

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Lab 4 Selective Media & Agar BIO250L” 3. What component makes starch agar selective for starch-degrading bacteria?

Because starch agar usually has in it the bacteria that degrades starch, it makes a good

selective media because it is will act as a selective nutrient to promote the growth of this

degrading bacteria

4. Why were each of the following steps performed in this experiment?

a. Serial dilution:

To act as a dilutant of microbes so that they it is possible to count them The

stock solution has too many microbes to be effective in the other process of

microbe identification thought isolation.-

b. Growth on the nutrient agar plates:

this was for purpose of bacteria isolation then transfer a single type of bacteria

hence being able to identify them

c. Streak on the starch agar plates:

to introduce a different media that would identify the presence of starch

degrading bacteria

Insert a photo of your plates. Include your name and access code handwritten in the background of

your photo.

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Lab 4 Selective Media & Agar BIO250L”

“EXPERIMENT 2: Selection and Differentiation of Body Inhabiting Gram-Positive Bacteria

Data Tables Table 4: Experiment 2 MSA Growth Observations

Surface Tested

Growth (Yes or No) –

Nutrient Agar

Growth (Yes or No) – MSA Agar

MSA color around colonies (Red or

Yellow)

Other Observations

Face Yes Yes Yellow Cfogged up –

nose Yes Yes Yellow Thick and droppy-

Controller Yes Yes Yellow Many organisms

–

Control No No Select one: clear –

Post-Lab Questions 1. What chemical in MSA confers selectivity? How?

The HIGH concentration of sodium chloride allows for the growth of microgranisms like

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Lab 4 Selective Media & Agar BIO250L” staphylococcus which while other types of this organism cannot ferment MSA, The

staphylococcus does

2. What chemical makes MSA differential? How?

Sugar mannitol is the differential chemical presence of organisms that can use sugar

mannitol as a source of nutrition will ferment it causing to become acidic which in turn

change the ph .This change in ph accounts for color change when this kind of bacteria is

present.-

3. Why are the nutrient agar plates used in this experiment?

The agar plates are serve to help compare the growth rates in MSA and Agar

4. Was there any growth on you MSA plates? Did any of the colonies change the color of the MSA?

What does this tell you about the bacteria taken from each area of your body?

yes there was a change in color which serves as confirmation that there were bacteria

that allowed for MSA to ferment hence making the set up acidic. The acidity is what

made the co

Insert a photo of your plates. Include your name and access code handwritten in the background of

your photo.

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Lab 4 Selective Media & Agar BIO250L”

EXPERIMENT 3: Selection and Differentiation of Body Inhabiting Gram-Negative Bacteria from

Liquid Samples

Data Tables Table 5: MacConkey Agar Results

Sample Growth?

(Yes or No) Colony Color (Clear or Red)

Analysis

CTap water- Yes Select one: Cpretty clean-

Spring water – Yes Select one: C pure and seems extremely safe –

Pond water – Yes Select one: extremely dirty-

Experiment 3 Post-Lab Questions 1. What types of bacteria are inhibited on MacConkey agar? What ingredient(s) in MacConkey agar

selects against those bacteria?

MacConkey agar are inhibited by Gram positive bacteria the ingredients

that allow for for selection are crystal violet and salts .

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Lab 4 Selective Media & Agar BIO250L” 2. What ingredient(s) makes MacConkey agar differential?

MacCoknkey agar is a differential Because it contains lactose. When bacteria works on

lactose to break it down, there is a release of acidic waste which changes the ph and

gives it the red-pink look

3. Using a textbook and a reputable online source (such as the CDC), research and describe some

potentially pathogenic members of the intestinal bacteria family Enterobacteriaceae. Which

pathogenic species are lactose fermenters that will grow on MacConkey agar?

Salmonella typhimurium- i will ferment lactose that is on the MacConkey agar.E-coli is

also a lactose fermenting bacteria.

4. Use a reputable internet source to research and describe some potentially pathogenic intestinal

bacteria that do not ferment lactose that will grow on MacConkey agar.

C-Legionella, Bordetella, Helicobacter are non lactose fermenting bacteria

5. Use a reputable internet source to research and describe what variations of MacConkey agar can

be used to detect other species of bacteria.

there is a variation of MacConkey Agar without Crystal Violet it detects Gram-Positive

cocci.- There there is MacConkey Agar that has Violet it is used to control t bacteria

like Proteus which interfere with other results.:

6. How would you verify that the colonies that grew on a MacConkey agar plate were Gram-

negative?

Crystal violet and bile salts usage helps identify gram negative colonies because they

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Lab 4 Selective Media & Agar BIO250L” stop the growth of gram-positive bacteria thus all that is left in the sample are gram

negative bacteria

7. Look up the formula for MacConkey agar either in a microbiology textbook or online. Is this a

chemically defined or a chemically complex media? Why is that important?

MacConkey agar is a complex chemically defined media because within it there

ingredients that propagate the growth of some bacteria while other ingredients inhibit the

growth of othes.

Insert a photo of your plates. Include your name and access code handwritten in the background of

your photo.

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Lab 4 Selective Media & Agar BIO250L”

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Biotechnology

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for manipulating the genomes of cells are:

1. to eliminate undesirable phenotypic traits in humans, animals, plants and microbes.

2. to combine the beneficial traits of two or more organisms to create a new more valuable organism.

3. to create cells that synthesize products humans need.

Recombinant DNA technology employs a number to tools and techniques to isolate genes and insert them into cells grown in culture. The following are five tools of recombinant DNA technology and examples of application of the tools.

Mutagens are physical and chemical agents that produce mutation.  Some mutations are beneficial, so creating a large number of mutations increases the probability of discovering a beneficial mutation. For example the fungus Penicillium may be mutated to syntheisize a more effective antibiotic. Mutagens include uv radiation and ionizing radiation. Mueller first demonstrated the effects of X rays on Drosophilia, establishing cause and effect between radiation and genetic changes manifested in phenotypic changes.  Chemical mutagens in bacteria may be carcinogenic in humans.  The Ames test utilizes bacteria to test for mutagenic activity, and identify chemicals that may induce cell changes that lead to cancer. Ethidium bromide which may be used to visualize DNA on gel, is also a mutagen because it disrupts base pairing.

1. Explain and describe one physical and one chemical mutagen and its application.

Reverse transcriptase  synthesizes cDNA from mRNA, which is the reverse of information flow as depicted in the central dogma of molecular biology, i.e.  DNA is transcribed to RNA which is translated to peptides. Cells may have millions of copies of mRNA, so being able to synthesize the complementary DNA reveals the gene that is being expressed. In addition, cDNA contains no introns because of processing in the eukaryotic transcription, so that cDNA may be inserted into a prokaryotic cell and may be translated into the corresponding peptide.

2. Explain how reverse transcriptase differs from RNA polymerase.  Give one example of an application of reverse transcriptase in recombinant DNA technology.

Synthetic nucleic acids are synthesized in vitro using enzymes from cells. Currently there are machines that syntheized DNA or RNA by having the sequence of bases entered tthrough a keyborad.

3.  If each letter of the keyboard represents a base in DNA, how many keys does the keyboard require? How does this means of DNA synthesis differ from DNA replication in the cell?

Synthetic nucleic acids were used to elucidate the genetic code, create genes for specific proteins, synthesize probes to locate genes in a genome, to synthesize antisense nucleic acids and to make PCR primers.

4. What is the genetic code, how might a synthetic nucleotide be used to determine which amino acid corrresponds to which codon?

5. Give an example of a probe with a flourescent tag that will be used to locate a gene associated with an aggressive form of breast cancer.

Restiction enzymes cut dsDNA at restriction sites, or palindromes. Examples of restriction enzymes are EcoRI and HindIII.  There are hundreds of restriction enzymes isolated from bacteria and used to cut DNA at predictable sites.  One type of restriction enzyme cuts dsDNA to make blunt ends and the other type creates sticky ends.

6. Compare and contrast blunt and sticky ends, and give one example of each type of restriction enzyme.

Vectors insert DNA into a new cell so that the cell acquires a new phenotype.   Vectors are pieces of DNA that are small enough to manipulate in a lab, survive inside the new cell, contain a recognizalbe genetic marker and carry the gene regions necessary to ensure the gene is transcribed and translated.   Examples of vectors are plasmids, viruses and transposons.

7. Explain one risk of a viral vector that inserts itself into a necessary gene and causes a mutation.

Gene library is a collection of cells or viruses, in which each member carries a portion of a given organism’s genome.  Gene libraries provide a ready

Techniques of recombinant DNA technology include the polymerase chaing reaction (PCR)  developed by Cary Mullis.  PCR has three steps that are repeated while the same sample of primers, target gene, taq polymerase and nucleotides are cycled through three different temperatures in a microfuge tube. Gel electrophoresis separated fragments of molecules by size, shape and charge.  DNA microarrays are able to monitor thousands of genes on one plate. Applications of these techniques include diagnostics or vaccine design.

8. Explain how a subunit vaccine is designed using  genes from a pathogen and a viral vector? p502-504.

One proposed way to stop the spread of arboviruses by mosquitoes is to vaccinate the mosquito.  This type of vaccine is a transmission blocking vaccine (TBV).  One typeof TBV vaccinates the human so that a blood meal from a vaccinated human would prevent the mosquito from being infected.  Another strategy is to use the gene drive known as Crispr-Cas9 which would edit the mosquito genome to confer resistance to diseases mosquitoes transmit to humans, and is very targeted.  This requires wild type mosquitoes to mate with lab mosquitoes that have been genetically modified.  Some data would indicate the lab mosquitoes are not competitve with wild type males.

9. Explain one advantage and one disadvantage of the TBV approach for controlling the spread of arboviruses.

10. Does the public have any control over the use of gene drives to alter species in the ecosystem for public health concerns?

 

 
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Mendelian Genetics

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Part I: Mendelian Genetics

In this assignment, you will use a useful tool, the Punnett square, to predict the probabilities of offspring gender and genotypes and phenotypes of different matings based on parental genetic makeup. Please answer all of the bulleted questions and tasks as you read through this assignment and submit them as a Word document to the assignment drop box titled “Punnett Squares Assignment.”

Each person has two copies of each chromosome, one from each parent. Your genome is exactly one-half your father’s genome and one-half your mother’s genome. After sperm and egg meet, the baby carries both copies of each gene in every cell for the rest of his or her life…except when eggs or sperm are produced. The eggs or sperm receive only one copy of each chromosome and the cycle starts all over again. Exactly which half will the baby get? That is the random part.

Sexual reproduction relies on chance to determine what type of offspring will result. A couple anxiously awaits a boy or girl and a dog breeder anxiously awaits the colors and markings of the puppies to be born. Although there is a random element involved, offspring from a mating will follow mathematical laws of probability based on the genetic makeup of the mother and father.

Watch this video to learn about Punnett squares. Please recall that dominant alleles mask recessive alleles and each baby has two copies of each gene, one from each parent.

 

http://www.clemson.edu/glimpse/?p=1175

  • Complete a Punnett square for the cross Bb x bb, where B is brown eyes and b is blue eyes.
  • What percentage of offpring will be BB? Bb? bb?
  • What percentage will have blue eyes and what percentage will have brown eyes?

Part II: Sex Determination

A male carries an X chromosome and a Y chromosome. A female carries two X chromosomes.

After meiosis, sperm and egg are produced. Sperm have a 50% chance of carrying and X chromosome and a 50% chance of carrying a Y chromosome. Eggs have a 100% chance of carrying an X chromosome, because females ONLY carry X chromosomes.

Sperm:                                             Eggs:

(X)       (X)           (Y)                            (X)                 (X)           (X)

(Y)          (Y)             (X)                                (X)           (X)            (X)

We can represent four potential scenarios if these two parents produce offspring neatly in a Punnett square. The possible sperm are placed above the top of the Punnett Square (circled in blue) and the possible eggs are placed along the left side beside the Punnett Square (circled in red). Each box is then filled in with the letter of each column and row.

punnett square gender

  • From the Punnett Square above, what is the % chance that offspring will be male? What is the percent chance that offspring will be female?

Part III: Codominance, Multiple Alleles and Blood typing

Review the embedded Amoeba Sisters video before completing the following questions.

 

Multiple alleles means that there are more than 2 alleles which can be inherited in a population.  Remember though, each individual only receives 2 alleles, one from each parent.

ABO blood typing uses 3 alleles. From your course notes answer the following two questions:

  • Which two are dominant? _________  Write the two dominant allele genotypes, using the capital letter I, and then a superscript for each of them. ______________________
  • Which allele is recessive? __________  Write the genotype (letters) for the recessive trait, beginning with the lower case i. ____________.

Complete a Punnett square for the following scenario:

Mom is heterozygous for blood type B and Dad is blood type AB. (make sure to put the parent alleles in the correct places outside of the square, and fill in each of the boxes within the square)

  • Place the Punnett square in your word document and answer the following questions:
  • What are all the possible blood types of their offspring? __________________________
  • What is the probability of their child having blood type AB? ________  Blood type O? _____________

Part IV: Deleterious Recessive Traits

A recessive trait is one where the individual must have two recessive alleles for the phenotype of the trait to be visible.  The recessive trait can be carried from generation to generation through heterozygous individuals. A person who is heterozygous for a harmful or deleterious trait is said to be a “carrier” of the trait.

If an individual receives 2 recessive alleles for a harmful or deleterious trait, let us say for our problem below, they will have the “illness”.

Solve the problem of the parents provided below.  Perform the Punnett Square and answer the questions below.  Only the answers are required in the answer sheet that you upload.

Mom and Dad are both heterozygous for the harmful or deleterious recessive trait.

  • What is the probability of their offspring having the illness?  ____________%
  • What is the probability of their offspring being a “carrier”? _____________%
 
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Micro HW 4

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Question 1

Future drugs and vaccines for the use against Ebola

 

Pick an experimental drug, vaccine or other treatment that is being considered in the fight against Ebola. http://www.cdc.gov/vhf/ebola/outbreaks/2014-west-africa/qa-experimental-treatments.html

 

Write a response to the following discussion question in the Discussion forum:

 

How exactly does the experimental treatment you have chosen work?

Provide a brief history of where we are now with the potential treatment.

How is it produced?

When do the developers of the treatment believe they will be able to begin experimental treatments?

Initial answers to the discussion question must be substantive and in the range of 350–500 words. Any references used should be properly cited following APA formatting guidelines.

 

Question 2

CDC’s New Superbug: CRE

 

CRE, which stands for Carbapenem-resistant Enterobacteriaceae, are a part of Enterobacteriaceae that are difficult to treat because they are resistant to commonly used antibiotics. Occasionally CRE are completely resistant to all available antibiotics. CRE have become a nightmare to infection control in health care facilities. http://www.usatoday.com/story/news/nation/2013/03/05/superbugs-infections-hospitals/1965133/ http://www.cdc.gov/hai/organisms/cre/

 

Write a response to the following discussion question in the Discussion forum:

 

Describe one of the new superbugs such as CRE, MRSA, Clostridium difficile, etc. that are causing havoc in our health care facilities.

What strategies are being employed to combat these new threats to our public health?

Initial answers to the discussion question must be substantive and in the range of 350–500 words. Any references used should be properly cited following APA formatting guidelines.

 

Question 3

2nd New Hampshire resident dies of Eastern Equine Encephalitis (EEE)

 

New Hampshire public health officials say a Manchester [Hillsborough County] resident is the 2nd person to die from eastern equine encephalitis (EEE) virus infection this season, summer and fall 2014. Public Health Director Dr. Jose Montero says the individual was exposed to the virus at the end of August and died in mid-September 2014. He says testing was complicated, and the virus was not confirmed until after the patient died. In the 1st fatal case, someone who became sick in the Conway, Carroll County, area in August died last month September 2014. A 3rd person also has been diagnosed with EEE, which is spread by the bite of infected mosquitoes and can lead to seizures and coma. Symptoms can include high fever, severe headache, stiff neck, and sore throat. These cases in New Hampshire are not surprising since they occurred within the geographical range of EEE virus in the eastern USA, where human and equine cases occur sporadically. As is usual, there are many more equine than human cases. There have been recent human cases in New Hampshire and an equine case involving a mule. The virus is maintained in passerine birds and transmitted among them by the black-tailed mosquito, Culiseta melanura. The species occasionally feeds on mammals and may have been the bridge transmitter in the above case. Clinical cases in humans are very serious, with a case fatality rate of 33 per cent and significant neurological sequelae in survivors. It is prudent to follow the recommendations above to avoid mosquito bites, particularly in areas where equine cases have been reported. http://www.cdc.gov/EasternEquineEncephalitis/

 

The first symptoms of EEE are fever (often 103º to 106ºF), stiff neck, headache, and lack of energy. These symptoms show up three to ten days after a bite from an infected mosquito. Inflammation and swelling of the brain, called encephalitis, is the most dangerous and common serious complication. The disease generally worsens quickly, and some patients may go into a coma within a week. People have an important role to play in protecting themselves and their loved ones from illnesses caused by mosquitoes. The group of viruses is known as arthropod borne viruses or Arboviruses and EEE is one of the deadliest.

 

Write a response to the following discussion question in the Discussion forum:

 

Discuss one of the mosquito borne viruses such as there are four main virus agents of encephalitis in the United States: eastern equine encephalitis (EEE), western equine encephalitis (WEE), St. Louis encephalitis (SLE) and La Crosse (LAC) encephalitis, all of which are transmitted by mosquitoes.

What recent outbreaks did you find on the Internet?

What strategies are being employed to combat these threats to our public health?

Initial answers to the discussion question must be substantive and in the range of 350–500 words. Any references used should be properly cited following APA formatting guidelines.

 

Question 4

Hantavirus discovery now 20 years old

 

The New Mexico Health Department confirmed on April 19th 2013 that a 45-year-old McKinley County woman has contracted the 1st case of a hantavirus this year in the state. In 1993, a massive investigation by public health officials from federal, state, and local agencies quickly discovered that a previously unknown hantavirus named Sin Nombre virus was the cause of the illnesses, and the deer mouse Peromyscus maniculatus was mainly responsible for excreting the virus in its droppings and urine. The virus killed 3 Yosemite visitors and sickened 6 more last summer. http://www.kob.com/article/stories/S3010686.shtml?cat=504

 

Write a response to the following discussion question in the Discussion forum:

 

Describe one of the outbreaks over the past 20 years.

What measures were used to prevent transmission of the virus?

Initial answers to the discussion question must be substantive and in the range of 350–500 words. Any references used should be properly cited following APA formatting guidelines.

 

 

 

 

Case study

At the Mount Union hospital, a 5-year old white male child in good general health and physical condition was presented at the Saturday walk-in clinic by his mother. He was brought in because he had a fever, was cranky and had complained of a sore throat for about 24 hours. On physical examination by the attending resident, the patient had a fever of 39.3° C, and he had considerable swelling and drainage of the pharynx and in the conjunctivae. His tonsils were enlarged and coated with a white patchy exudate. He had a red throat and swollen anterior cervical lymph nodes. His ears were clear. His chest sounded clear and he had no additional remarkable findings on routine examination.

Directions: Answer the following questions in your own words.

1. What would be your presumptive diagnosis for this child? Why?

 

 

 

 

 

2. What diagnostic testing would be indicated to follow this exam?

 

 

 

 

 

3. What is the most likely treatment for this illness? Why is it important?

 

 

 

 

 

4. What factors of this case allowed you to make a presumptive diagnosis?

 

 

Any references used should be properly cited following APA formatting guidelines.

 

 

Case study 2

 

A 62-year old diabetic black man presents in the emergency room with a swollen left leg with areas of blanching and blue mottling. A “foul odor” is coming from a dressed wound. The physicians remove the dressing and a brownish fluid is seeping from a wounded area. The fluid contains what appear to be small bits of the tissue. No pus appears to be present. The wound has a strong “rotten” odor.

Five days earlier, while at his work as a farmer, he caught the leg in his manure spreader, sustaining a deep, crushing, grossly dirty injury. His wife cleaned the wound as well as she could with soap and water, dressed it with clean gauze, and wrapped it tightly with an elastic bandage to stop the bleeding. The second day they redressed the wound and applied triple antibiotic ointment. The patient treated his pain with ibuprofen (Advil). He reported the pain was not very bad for the first 72 hours. In the past 24 hours, the leg swelled and the mottling began to appear. A foul odor and severe pain accompanied the swelling. His wife convinced him to come to the emergency room even though they did not have medical insurance.

Directions: Answer the following questions in your own words.

1. What is your diagnosis in this case?

 

 

 

 

 

2. How should this wound be treated?

 

 

 

 

 

3. Is this a life-threatening condition?

 

 

 

 

 

4. Is it likely that the patient’s diabetes contributed to the problem as presented?

 

 

 

 

 

Any references used should be properly cited following APA formatting guidelines.

 
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