Log in
Biology
1
answer
0
watching
88
views

I need help with these questions about Genetics Laboratory techniques involving DNA extraction please:

1.) We add RNase solution to the extraction cocktail (tissue and lysis buffer) in order to:

a. Degrade RNase to remove remaining RNase

b. Prevent degradation of DNA to increase yield of DNA

c. Remove any RNA from the extraction cocktail to ensure isolation of deoxynucleic acid molecules

d. All of the above

2.) After precipitation of DNA and centrifugation at maximum speed, a “supernatant” is produced. The “supernatant” is the portion that contains DNA but not the pellet.

a. True

b. False

3.) After precipitation of DNA and centrifugation at maximum speed, the supernatant is discarded.

a. True

b. False

4.) ____________ precipitation of DNA and centrifugation at maximum speed and washing the pellet using ethanol, _________________ is added to the pellet to elute the DNA in the solution.

a. After - DNA rehydration buffer

b. Before – isopropanol

c. After – lysis buffer

d. Before – ethanol

Watch
1
answer
0
watching
88
views

For unlimited access to Homework Help, a Homework+ subscription is required.

Jamar Ferry
Jamar FerryLv2
28 Sep 2019

Unlock all answers

Get 1 free homework help answer.
Already have an account? Log in
Share

Related textbook solutions

Molecular Cell Biology
8 Edition,
Lodish
ISBN: 9781464183393
Biology: Science for Life with Physiology
6 Edition,
Belk
ISBN: 9780134555430
Concepts of Biology
OER Edition,
Openstax
ISBN: 9781938168116
Biology
2nd Edition, 2013
Openstax
ISBN: 9781947172517
Essentials of Biology
5 Edition,
Mader
ISBN: 9781259660269

Related questions

I need help with these questions about Genetics Laboratory techniques involving DNA extraction please:

1.) After precipitation of DNA and centrifugation at maximum speed, supernatant is discarded and ethanol is added to the remainder to wash the pellet.

a. True

b. False

2.) ____________ precipitation of DNA and centrifugation at maximum speed and washing the pellet using ethanol, _________________ is added to the pellet to elute the DNA in the solution.

a. After - DNA rehydration buffer

b. Before – isopropanol

c. After – lysis buffer

d. Before – ethanol

3.) Using a heat block after adding DNA rehydration buffer to the pellet will accelerate the process of elution.

a. True

b. False

4.) In an electrophoresis gel, DNA moves from the negative electrode towards the positive electrode because: (1) the molecule of DNA is negatively charged and (2) the loading dye moves from the positive electrode towards the negative electrode.

a. True

b. False

Please answer questions at the end.

In this lab, you will add different volumes of “cell-free supernatant” (CFS) to fresh cultures of V. harveyi and observe bioluminescence. The CFS is prepared by culturing V. harveyi in liquid medium overnight, then separating the cells from the supernatant by centrifugation.

MATERIALS • overnight culture of V. harveyi in AB medium • fresh AB medium • 1 microcentrifuge tube • 3 glass culture tubes

PROCEDURE

1. This experiment will be performed with your lab group. Label the microcentrifuge tube and 3 glass cultures tubes with your group number. Also, label the 3 glass culture tubes as follows: “Media only,” “Cells+fresh media,” and “Cells+CFS.”

2. Using aseptic technique, pipet different volumes of fresh AB medium into the glass tubes as follows: 1 mL in the “Media only” tube, 800 ul in the “Cells+fresh media” tube, and 500 ul in the “Cells+CFS” tube.

3. Pipet 500 ul of the overnight culture into a microcentrifuge tube and centrifuge at maximum speed (>13,000 x g) for 5 min to pellet the cells.

4. After centrifugation is complete, transfer 300 ul of the cell free supernatant (CFS) from the microfuge tube to the glass tube labeled “Cells+CFS”. Be careful to avoid the cell pellet.

5. Remove the remaining supernatant from the microfuge tube using a micropipette and discard as culture waste.

6. Add 500 ul of fresh AB medium to the cell pellet in the microfuge tube and pipet up and down to resuspend until no clumps are visible.

7. Add 200 ul of the cell suspension to the “Cells+fresh media” tube and 200 ul of the cell suspension to the “Cells+CFS” tube. Note the time.

8. Place the 2 glass tubes containing cells in the 30 degree incubator/shaker.

9. After 1 hour and 2 hours, observe your tubes next to the “Media only” tube in a dark room. You will need to let your eyes adjust for at least 30 seconds (count to 30 slowly in the dark) before you can evaluate whether the cultures are luminescent or not. Also, make sure you try swirling the tubes in the darkroom and observe the tubes for a minute after you have stopped swirling. After the observation at the 1 hour time point, put your tubes back in the incubator shaker until the 2 hour time point.

10. Note your observations. At the end of the lab, place your tubes in the tube waste.

Questions

1. What were the expected results? Explain the rationale of the experiment. If you didn’t obtain the expected results, hypothesize why not and describe how the experiment could be improved.

2. What are differences between luminescence and fluorescence?

3. Why do V. harveyi cultures glow more brightly when swirled vigorously? Be specific.

4. Imagine you have a mutant V. harveyi strain that can’t synthesize autoinducer and another mutant V. harveyi strain that makes a defective luciferase. If you streaked both strains near (but not touching) each other on the same plate, would you expect to see light produced? If so, by which strain, where, and why?

5. You have two identical tubes of wild-type V. harveyi (with the same cell concentration in each tube) in fresh media. To one tube, you add CFS from a light-producing V. fischeri culture, and to the other tube you add an equal volume of fresh media. If both tubes of V. harveyi have the same light production, what can you conclude? Explain.

6. Many biomedical research labs study quorum sensing in V. fischeri or V. harveyi. How is studying light production in marine bacteria relevant to human health? List at least 3 different examples.

7. Why do you think it is important for bacteria to be able to control the expression of some genes based on the number of individuals present?

can anyone paraphrase the following procedure for me please. I prefer to be typed because the handwriting sometimes is not clear

First experiment: Bacterial Transformation and Affinity Purification of Protein:

Transformation procedure:

Bring 2 test tubes and label them +pGLO and –pGLO,

Transfer 250 ul of transformation solution to each tube and then place them in ice.

Pick a single colony of bacteria using a sterile loop from the bacterial plate and immerse the loop in +PGLO tube and start spinning the loop to mix, repeat the same step for –pGLO and place both tubes in ice again.

Then, immerse a new sterile loop in the pGLO DNA plasmid stock tube and withdraw a loopful of the plasmid and immerse it in the +pGLO tube to load it and place the tube in ice again. No pGLO plasmid will be added to the –pGLO sample as labeled.

Incubate the tubes in ice for 10 minutes and take the following labeled plates from the instructor: LB/amp + pGLO, LB/amp/ara + pGLO, LB/amp – pGLO, and LB – pGLO. There will be a total of four plates.

Heat hock will start by transforming both tubes to 42C water bath for 50 second and then returning them to ice again.

Add 250 ul of LB nutrient broth to both tubes and incubate at room temperature for 10 minutes.

Using a new sterile pipet for each plate, transfer 100 ul of the –pGLO solution to the, LB/amp – pGLO and LB – pGLO agar plates and 100 ul of +pGLO solution to the LB/amp + pGLO, and LB/amp/ara + pGLO plates.

Spread the suspension using sterile loops and place your plates upside down in the 37C incubator overnight.

Bacterial growth will occur and result can be analyzed the day after.

* For data analysis, observe the result in the plates under normal light conditions and then under UV light and right your observations. This will include observing the number of colonies, colonies color and growth. Transformed bacteria will look green in color under UV with the presence of arabinose sugar in the media and will look white in color if arabinose is absent. No growth will be shown if the bacteria are not transformed.

Second experiment: Affinity purification of green fluorescent protein:

Affinity purification procedure:

Re-observe you plates from previous experiment under natural light and UV light. Identify several green colonies on the LB/amp/ara plate and with the use of a loop scoop up some cells and immerse the loop in a new culture tube labeled (+), the same procedure is done for the LB/amp plate in a tube labeled (-). Disperse the colonies in the culture tubes and shake vigorously. And place in an incubator for 24 hours.

Transfer the entire culture to a 2-milliliter microtube labeled +. Centrifuge for 5 minutes and remove the supernatant.

Re suspend the pellet in 250 ul of TE solution and then add 1 drop of lysozyme and mix then place the tube in the freezer to help rupture the cells.

For bacterial lysis, thaw the microtube and centrifuge for 10 minutes at maximum speed. And prepare the chromatography column.

Before performing the chromatography, add 2 milliliters of Equilibration Buffer to the column, and Drain the buffer from the column until it reaches the 1 milliliter.

After centrifugation, transfer 250 μl of the supernatant into the new microtube and transfer 250 μl of Binding Buffer to the microtube and Place in the refrigerator.

Label 3 collection tubes 1, 2 and 3. Load 250 μl of the supernatant in Binding Buffer into the top of the column and Let the entire volume of supernatant flow into tube 1.

Transfer the column to collection tube 2. Add 250 μl of Wash Buffer and let the entire volume flow into tube 2.

Transfer the column to tube 3. Add 750 μl of TE buffer and let the entire volume flow into tube 3.

Examine all collection tubes using the UV lamp and note any differences in color between the tubes.

Weekly leaderboard

Home
Homework Help 3,900,000
Biology 460,000
Start filling in the gaps now
Log in