Editing Molecular phylogenetics (section)

==Theoretical background==
Early attempts at molecular systematics were also termed [[chemotaxonomy]] and made use of proteins, [[enzyme]]s, [[carbohydrate]]s, and other molecules that were separated and characterized using techniques such as [[chromatography]]. These have been replaced in recent times largely by [[DNA sequencing]], which produces the exact sequences of [[nucleotides]] or ''bases'' in either DNA or RNA segments extracted using different techniques. In general, these are considered superior for evolutionary studies, since the actions of evolution are ultimately reflected in the genetic sequences. At present, it is still a long and expensive process to sequence the entire DNA of an organism (its [[genome]]). However, it is quite feasible to determine the sequence of a defined area of a particular [[chromosome]]. Typical molecular systematic analyses require the sequencing of around 1000 [[base pair]]s. At any location within such a sequence, the bases found in a given position may vary between organisms. The particular sequence found in a given organism is referred to as its [[haplotype]]. In principle, since there are four base types, with 1000 base pairs, we could have 4<sup>1000</sup> distinct haplotypes. However, for organisms within a particular species or in a group of related species, it has been found empirically that only a minority of sites show any variation at all, and most of the variations that are found are correlated, so that the number of distinct haplotypes that are found is relatively small.<ref>{{Cite book|title=Molecular evolution: a phylogenetic approach |last1=Page |first1=Roderic D. M. |first2=Edward C. |last2=Holmes |author-link2=Edward C. Holmes |date=1998 |publisher=[[Blackwell Science]] |isbn=9780865428898 |location=Oxford |oclc=47011609}}</ref>

[[File:Clade in Phylogenetic Tree.png|400x220px|thumb|In a phylogenetic tree, numerous groupings (clades) exist. A clade may be defined as a group of organisms having a common ancestor throughout evolution. This figure illustrates how a clade in a phylogenetic tree may be expressed.]]

In a molecular systematic analysis, the haplotypes are determined for a defined area of [[genetic material]]; a substantial sample of individuals of the target [[species]] or other [[taxon]] is used; however, many current studies are based on single individuals. Haplotypes of individuals of closely related, yet different, taxa are also determined. Finally, haplotypes from a smaller number of individuals from a definitely different taxon are determined: these are referred to as an [[Outgroup (cladistics)|outgroup]]. The base sequences for the haplotypes are then compared. In the simplest case, the difference between two haplotypes is assessed by counting the number of locations where they have different bases: this is referred to as the number of ''substitutions'' (other kinds of differences between haplotypes can also occur, for example, the ''insertion'' of a section of [[nucleic acid]] in one haplotype that is not present in another). The difference between organisms is usually re-expressed as a ''percentage divergence'', by dividing the number of substitutions by the number of base pairs analysed: the hope is that this measure will be independent of the location and length of the section of DNA that is sequenced.

An older and superseded approach was to determine the divergences between the [[genotype]]s of individuals by [[DNA–DNA hybridization]]. The advantage claimed for using hybridization rather than gene sequencing was that it was based on the entire genotype, rather than on particular sections of DNA. Modern sequence comparison techniques overcome this objection by the use of multiple sequences.

Once the divergences between all pairs of samples have been determined, the resulting [[triangular matrix]] of differences is submitted to some form of statistical [[cluster analysis]], and the resulting [[dendrogram]] is examined in order to see whether the samples cluster in the way that would be expected from current ideas about the taxonomy of the group. Any group of haplotypes that are all more similar to one another than any of them is to any other haplotype may be said to constitute a [[clade]], which may be visually represented as the figure displayed on the right demonstrates. [[Statistical]] techniques such as [[Bootstrapping (statistics)|bootstrapping]] and [[Resampling (statistics)#Jackknife|jackknifing]] help in providing reliability estimates for the positions of haplotypes within the evolutionary trees.