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Chromosome Structure

Page history last edited by Derek Weber 12 years, 3 months ago

A.  Learning Objectives

In this lab, students will:

• prepare and view a slide of chromosomes from HeLa cells.

 

 

 B.  Chromosome Structure

During interphase, when the cell is actively engaged in its metabolic processes, its DNA is unwound into a thin strand referred to as chromatin. In this form it is available for transcription (the production of RNA from a DNA template). When the cell prepares to enter mitosis, the chromosomes (which were replicated during S phase) are condensed into a very tight coil. The two identical replicated chromosomes are called sister chromatids and are connected to each other at the centromere (see Figure below).

   

It is possible to view actual chromosomes from a line of cells called HeLa cells. These cells originated in a woman named Henrietta Lacks. She died from cervical cancer in 1951. Prior to her death, some of her cancerous cells were removed from her body and cultured. In this artificial environment, the cells continued to grow and repeatedly divide.  Up until that time, the viability of cells in the laboratory environment was limited.  This was a major scientific breakthrough as it allowed scientists to study a cell line that was immortal.   These cells have been used to study HIV, to cultivate vaccines, to study the behavior of cancer cells and for biology students, to observe chromosomes.

 

Being cancerous cells that have lost the normal checks and balances on growth and chromosome number, these cells tend to produce cells with more than the normal 23 pairs (46 total chromosomes) of chromosomes.  In this exercise, you will prepare a slide of HeLa cells, stain it, and identify and count the number of chromosomes in the cells.  

 

Procedure

Part I:  Prepare the specimen

1. Cover your work tray with paper towels.

2. Rest against the side of the work tray at a 45° angle so that short edge is resting on the paper towel.

3. Resuspend the HeLa cells in the culture tube by placing the tip of the pipet near the bottom of the culture tube and squeezing the bulb to expel air.

4. Remove a small sample with the pipet.

5. Hold the pipet 18-36 cm (use the ruler to determine this height) above the slide and gently

squeeze the bulb so that 8-10 drops of the culture “splats” onto the slide. Aim for the upper edge of the slide so the drops splatter down the slide.

6. Allow the slide to air dry completely.

 

Part II: Staining

7. Dip your slide in stain 1 three times for 1 second each time. Timing is critical; count “one thousand and one” to determine 1 second.

8. Blot the excess stain off the bottom edge of your slide with paper towel.

9. Immediately dip your slide in stain 2 three times for 1 second each time.

10. Blot the excess stain off the bottom edge of the slide.

11. Next, rinse your slide by gently swishing it back and forth in dH2O.

12. Allow the slide to air dry completely.

13. Make a wet mount of your slide using dH20 and a coverslip.

 

Part III: Viewing the Chromosomes

14. Place your slide on the microscope and first use the medium-power objective and then switch to high power to locate an area where the cells appear to have burst and the chromosomes are spread out.

15. Once you have identified a good specimen, alert the instructor to assist in using oil-immersion to view the chromosomes.

16. Closely observe the structure of the chromosomes and upload your observations in the space below.  Label sister chromatids and centromeres.  Count the number of chromosomes.  Do they look abnormal?  How?  Explain.

 

Insert labelled images here. 

 

   

 

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