Relative dating (A was before B) is often good enough for studying processes of evolution, but this has also been difficult, because of the problems involved in matching up rocks of the same age across different continents, particularly around the internationally-defined Precambrian/Cambrian boundary section.^[1] (the most common technique uses widespread but short-lived fossil species to identify rocks of similar ages)

So any dates or descriptions of sequences of events should be regarded with caution until better data become available.

Body fossils preserve significant parts of organisms and are therefore the most informative type of evidence. Unfortunately they are increasingly rare as one looks further back in time, among other reasons because the rocks in which they are buried are usually covered by more recent rocks and because they may have been eroded before being covered by later rocks. One recent study concluded that “parts of the fossil record are clearly incomplete, but they can be regarded as adequate to illustrate the broad patterns of the history of life.”^[2] But there is evidence that some types of animals or parts of animals are relatively likely to be preserved as fossils in some environments and times, and extremely unlikely to be preserved in other environments and times. Part of this is due to changes in the chemistry of the oceans, which were partly caused by the on-going evolution of life, and these changes were most significant before the start of the Cambrian – for example any increase in the marine biomass would reduce the concentration of carbon, and the appearance of sponges reduced the concentration of silicon.^[3]

Another limitation in the discovery and use of body fossils is the lack of preservation of large portions of the body. In most cases the sole anatomical features that are fossilized are the highly mineralised body parts containing high proportions of silica (sponges' skeletons), calcium carbonate (the shells of bivalves, gastropods and ammonites and exoskeletons of most trilobites and some crustaceans) or calcium phosphate (the bones of vertebrates). The majority of animal species living now are unlikely ever to leave fossils, since they are soft-bodied invertebrates such as worms and slugs. Of the more than 30 phyla of living animals, two-thirds of these have never been found as fossils.^[4]

The Cambrian fossil record includes an unusually high number of lagerstätten which preserved the fossils' soft tissues in extremely fine detail, allowing a very informative study of animals that normally would not have left fossils. The fine detail of the deposits has allowed paleontologists to examine the internal workings of animals which in other sediments are only represented by shells, spines, claws, etc. The most significant Cambrian lagerstätten are: the early Cambrian Maotianshan shale beds of Chengjiang (Yunnan, China) and Sirius Passet (Greenland)^[5]; the middle Cambrian Burgess Shale (British Columbia, Canada)^[6]; and the Upper Cambrian Orsten (Sweden) fossil beds.

While lagerstätten are superior to most fossil beds in preserving fine anatomical detail, they are far from perfect. The majority of then-living animals are probably not represented because lagerstätten are restricted to a narrow range of environments (e.g. where soft-bodied organisms can be preserved very quickly such as by mudslides), and the exceptional events that cause quick burial make it difficult to study the normal environments of the animals.^[7] In addition, the known lagerstätten cover only a very limited period of time within the Cambrian, and none covers the crucial period just before the start of the Cambrian. Because normal fossil beds are very rare and lagerstätten even rarer, both are very unlikely to show the first occurrence of any type of organism.^[8]

Trace fossils consist mainly of tracks and burrows on and under what was then the seabed.

Trace fossils are particularly significant because they represent a data source that is not limited to animals with easily-fossilized hard parts. Also many traces date from significantly earlier than the body fossils of animals that are thought to have been capable of making them.^[9] Whilst exact assignment of trace fossils to their makers is generally impossible, traces may provide the earliest physical evidence of the appearance of moderately complex animals (comparable to earthworms).

The ratios of three major isotopes, ⁸⁷Sr / ⁸⁶Sr, ³⁴S / ³²S and ¹³C / ¹²C, undergo dramatic fluctuations around the beginning of the Cambrian.^[10] This chemical signature in the rocks of the Precambrian/Cambrian boundary is difficult to interpret, and may be related to continental break-up, the end of a “global glaciation”, or a catastrophic drop in productivity caused by a mass extinction just before the beginning of the Cambrian.

Carbon has 2 stable isotopes, carbon-12 (¹²C) and carbon-13 (¹³C). Causes often suggested for changes in the ratio of ¹³C to ¹²C found in rocks include:^[11]

• A mass extinction. Chemistry is largely driven by electro-magnetic forces, and lighter isotopes such as ¹²C respond to these more quickly than heavier ones such as ¹³C. So living organisms generally contain a disproportionate amount of ¹²C. A mass extinction would increase the amount of ¹²C available to be included in rocks and therefore reduce the ratio of ¹³C to ¹²C.
• A methane “burp”. In permafrosts and continental shelves methane produced by bacteria gets trapped in “cages” of water molecules, forming a mixture called a clathrate. This methane is very rich in ¹²C because it has been produced by organisms. Clathrates may dissociate (break up) suddenly if the temperature rises or the pressure on them drops. Such dissociations release the ¹²C-rich methane and thus reduce the ratio of ¹³C to ¹²C as this carbon is gradually incorporated into rocks (methane in the atmosphere breaks down into carbon dioxide and water; carbon dioxide reacts with minerals to form carbonate rocks).

Cladistics is a technique for working out the “family tree” of a set of organisms, and has most often applied to evidence from comparative anatomy (features of the bodies of organisms). In this kind of analysis it is possible to include both living and fossilized organisms and work out their evolutionary relationships. Sometimes one can conclude that group A must have evolved before groups B and C, because B and C have more similarities to each other than either has to A. On its own this method can say nothing about when A evolved, but if there are fossils of B or C dating from X million years ago, then A must have evolved more than X million years ago.

Molecular phylogenetics attempts to reconstruct the relationships between organisms by comparing details of their biochemistry, such as their DNA. In other words, it applies the analysis techniques of cladistics to biochemical rather than anatomical features. It provides an alternative line of evidence about evolution in the Cambrian and Precambrian, although the need for calibration against the fossil record means it is not entirely independent. Further, since the “clocks” measure molecular evolution, a period of rapid evolution is indistinguishable from a longer period of slow change, so it is unwise to rely on molecular phylogeny for estimates of dates^[12].

Although this rapidly developing science must be treated with a degree of caution,^[13] it has yielded some useful results. For example, it provides evidence that the three major animal groups diverged some time before the Cambrian, then independently underwent a rapid Cambrian diversification^[14] – although the reliability and implications of this apparent finding are still being debated.^[15] The current state of molecular phylogenetics seems not to support the Cambrian Explosion theory, but rather a considerably earlier evolutionary radiation.^[16]