AS Biology 9700: Calculating Mitotic Index From a Root Tip Squash

A student prepared a stained root tip squash from an onion (Allium cepa) and examined it under a light microscope. In one field of view, the student counted 250 cells in total. Of these, 20 cells had visibly condensed chromosomes, and the remaining cells showed no visible individual chromosomes.

(a) Define the term mitotic index. [1]

(b) Calculate the mitotic index for this field of view, showing your working. [2]

(c) The 230 cells with no visible individual chromosomes could be in the G1, S or G2 phase of interphase. Explain why a cell in any of these three phases looks the same under a light microscope. [2]

(d) State two visible features that would allow the student to identify a cell as being in metaphase, rather than a cell in interphase. [2]

Part (a): Defining mitotic index

The mitotic index is the proportion of cells in a sample that are in mitosis (rather than in interphase) at the moment the sample is examined, expressed as a fraction or a percentage of the total number of cells counted.

Part (b): Calculating the mitotic index

The student counted 20 cells with visibly condensed chromosomes (i.e. cells in mitosis) out of a total of 250 cells in the field of view.

Mitotic index = (number of cells in mitosis / total number of cells counted) x 100 Mitotic index = (20 / 250) x 100 = 8%

So the mitotic index for this field of view is 8%.

Part (c): Why interphase sub-phases look identical

During G1, S and G2, the chromosomes have not yet condensed: they exist as long, thin, loosely coiled chromatin spread throughout the nucleus, which is far too fine and extended for individual chromosomes to be resolved as separate structures using a light microscope. This is true regardless of what is happening biochemically. A cell in G1 is growing and making proteins and organelles, a cell in S phase is actively replicating its DNA, and a cell in G2 is continuing to grow and preparing for mitosis, but none of these processes changes the loosely coiled appearance of the chromatin enough to be visibly distinguished. It is only when the chromosomes condense (supercoil) during prophase, at the start of mitosis, that they become visible as distinct, individual structures, which is why a light microscope alone cannot tell G1, S and G2 apart.

Part (d): Identifying a cell in metaphase

A cell in metaphase can be distinguished from a cell in interphase by several visible features. The nuclear envelope, present around the chromatin in interphase, has completely broken down by metaphase. The chromosomes, invisible as individual structures in interphase, are now condensed and clearly visible, each one still made of two sister chromatids joined at a centromere. Rather than being scattered through the nucleus as in interphase, these condensed chromosomes are arranged in a single line along the equator of the cell (the metaphase plate). Spindle fibres, which are absent in interphase, can also be seen spanning the cell between the two poles, attached to the centromere of each chromosome.

Final answers

(a) Mitotic index: the proportion of cells in a sample that are undergoing mitosis, out of the total cells counted.
(b) Mitotic index = 20/250 x 100 = 8%.
(c) Chromosomes in G1, S and G2 are uncondensed, thin chromatin that cannot be resolved into distinct chromosomes with a light microscope, regardless of which sub-phase the cell is in.
(d) Any two of: no nuclear envelope; visible condensed chromosomes (as sister chromatid pairs); chromosomes aligned along the equator; visible spindle fibres.