Unlike other species, passenger pigeons (Ectopistes migratorius) faced a recent extinction. This extinction took place at the turn of the 20th century, the last individual died in captivity in 1914. Based on morphology it was assumed that they belonged in the genus Zenaida (Fulton 2012). This genus consists of mourning doves. When the mitochondrial DNA was analyzed it was found that they are more closely related to Patagioenas which are the New World pigeons. They used mitochondrial DNA because there is a high number of copies of this DNA in the cells when you compare this to the 2 copies of DNA that are in the nucleus. For this reason, it is common to use this for the analysis of ancient DNA.
The DNA was extracted from a toe pad sample. The toe pad sample came from a museum. This was done at Pennsylvania State University in a special DNA lab. They used ancient DNA protocols in order to reduce potential contamination of the sample. They extracted the DNA using the Qiagen DNeasy Tissue Kit. The lysis step was extended for 4 days and extra proteinase K was added. They sequenced intron 7 of the FGB gene. They used PCR to amplify this section. Using the TOPO TA cloning kit the samples were also cloned and then sequenced. This allowed the researchers to look for damage and contamination.
To make the phylogeny itself, the researchers used the ATP8, 12s rRNA, and cytb genes in addition to the FGB gene. Data for other Columbiformes came from Gen Bank. They aligned locus using the MAFFT v.6 online moderately accurate automatic strategy. They used data from 122 Columbiformes and 6 outgroups. They used jModelTest to create the best fit model. It was found that there is a high statistical probability that the passenger pigeon belongs within the typical pigeon and dove group. The data shows that there also was a single origin of the New World pigeons.
Fulton, T. L., Wagner, S. M., Fisher, C. Shapiro, B. 2012. Nuclear DNA from the extinct Passenger Pigeon (Ectopistes migratorius) confirms a single origin of New World pigeons. Annals of Anatomy. 194, 52- 57.