Greenness as an Indicator of DNA Quality in Herbarium Specimens

An experimental and comparative study of leaf greenness as an indicator of DNA barcode recovery in herbarium specimens. 

Herbarium collections are becoming increasingly important sources of DNA for plant research, yet the recovery of usable sequence data from preserved plant material remains difficult. The DNA contained in herbarium specimens is often degraded, fragmented, chemically damaged and has usually undergone different methods of preservation. This study tests whether leaf greenness can be used as an indicator of DNA quality and DNA barcode recovery success in herbarium specimens, using specimens from the Natural History Museum of Denmark (NHMD) supplemented with data from the Barcode UK project, 194 NHMD and 224 Barcode UK specimens were analysed. 

To assess the role of leaf greenness in DNA recovery success, specimens were categorized in six greenness stages and a range of age groups. Three standard barcode markers were used: rbcL, ITS2 and matK. DNA concentration was measured on all NHMD specimens before amplification.

There was a large variation between the recovery success of the markers after amplification, where rbcL showed the highest success overall, and matK performed worst. Specimen age was the most dominant factor and was a more consistent indicator of recovery than greenness. Greener specimens did generally have higher DNA concentrations and more favourable recovery outcomes, but the stage of greenness was not the sole indicator of being able to get a useful sequence.

The difficulty of getting a usable sequence between the different taxonomic orders, is also something to consider. This suggest that leaf greenness can provide useful information about the potential success of barcode recovery, but it should be considered alongside the specimen age, taxonomic grouping and choice of marker.