DNA content through the cell cycle

The stages of the cell cycle can be assessed experimentally by measuring the DNA content in the cell. As discussed in the module on Genome Structure, a genome of a cell can be described in terms of the ploidy (copies of the genome) as well as the number of chromosomes in the genome. For example, human cells are described as 2n=46, where 2 indicates a diploid genome, and 46 indicates the total number of chromosomes. If 2n = 46, then simple math tells us n=23: the human genome has 23 pairs of chromosomes. While this textbook most often focuses on diploid (2n) cells and organisms, it is important to remember that some organisms have other ploidy: haploid (1n), triploid (3n), tetraploid (4n) and other ploidies exist as well, and are common in some plants.

At the start of the cell cycle in G1 phase, a human cell has DNA chromosomes described as 2n (diploid), each chromosome consists of one DNA molecule, and DNA content described as 2C. But by the end of S phase, DNA has been replicated, and each chromosome consists of two DNA molecules/chromatids (Figure 3). By the end of S phase, content is described as 4C: the number of chromosomes (2n) hasn’t changed, but the total DNA content has doubled because each chromosome now consists of two chromatids. The DNA content for cells in each phase of the cell cycle is shown in Figure 4.

Graphic depicting the cell cycle as a colorful circle, with M phase at the top and G1, S, and G2 arranged clockwise. At the end of each stage, a cell is shown with chromosomes. After G1 phase, cell has 2n = 2C = 4 chromosomes. After S phase and after G2 phase, the cell has 2n = 4C = 4 chromosomes. After M phase, two cells are produced, each with 2n = 2C = 4 chromosomes.
Figure 4 The DNA content of a cell changes with the stage of the cell cycle. After replication, a cell has twice as much DNA content (C) as in G1, although the total number of chromosomes (n) has not changed.

DNA content can be measured experimentally by using techniques like flow cytometry. Flow cytometry can measure the DNA content of single cells by labeling them with a fluorescent dye that binds to DNA: the cells are passed through a sensor that detects fluorescence, and the signal in each cell is measured. “Cyto” refers to cell, so “flow cytometry” refers to the measurement of the cells as they flow through the sensor.

Typically, an experiment measuring DNA content will present data from a population of cells, as shown in Figure 5, plotting the number of cells counted vs the fluorescent signal. The x-axis is often presented as DNA content rather than fluorescence, since the fluorescent signal is proportional to the amount of DNA in a cell.

Graph illustrating sample DNA content analysis via flow cytometry. On the x-axis is DNA content, and on the y-axis is number of cells. A two-peaked line is seen on the graph, with the first peak higher than the second. The first peak corresponds to cells in G1 phase. The second peak corresponds to cells in G2 phase. And the connecting region corresponds to cells in S phase.
Figure 5 Flow cytometry to measure DNA content.
Cells are stained with a dye that fluoresces when in contact with DNA. A flow cytometer measures the fluorescence in one cell at a time. The data are plotted as the number of cells observed vs their DNA content. An asynchronous population of cells typically appears like this, with a large peak representing cells in G1, a smaller peak or cells in G2, and cells in the middle representing S phase (with an intermediate DNA content). Not drawn to scale.

When an asynchronous population of diploid cells is measured for DNA content, a two-peak pattern like that seen in Figure 5 is observed: the greatest number of cells typically have a DNA content of 2C and are presumed to be in G1 phase of the cell cycle. A smaller number of cells have a DNA content of 4C and are in G2 phase. And some cells have an intermediate amount of DNA content: these are in S phase of the cell cycle and are in the process of making additional DNA. The abundance of cells with each DNA content is proportional to the length of each stage of the cell cycle: if about 40% of the cell cycle is spent in G1 phase, about 40% of cells in an asynchronous population will be in G1 phase. This technique is often used to measure the effects of experimental manipulation on the cell cycle. It also can be used to measure the ploidy of a cell.

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