Molecular and ultrastructural detection of plastids in Juniperus ( Cupressaceae ) pollen

Transmission electron microscopy (TEM), PCR and sequencing studies can provide important evidence about the presence of plastids in pollen grains. Their presence is critical if one proposes to develop plastid primers for the amplification of DNA from pollen for species identification. Differential interference contrast microscopy (DIC) and TEM were used to investigate the presence of plastids in pollen from Juniperus species, a major cause of airborne allergies in North America. Standard PCR techniques were used to amplify DNA from the pollen using universal primers targeting known plastid DNA genes including the rbcL and species-specific primers targeting the matK coding regions. Studies using TEM confirmed the presence of plastids in Juniperus pollen at three increasing stages of hydration. The rbcL and matK genes were successfully amplified using DNA extracted from pollen of J. ashei, J. pinchotii and J. virginiana. The results open a wide range of possibilities for using plastid primers in general pollen research especially when the interspecific variation among the standard set of nuclear genes (for e.g., ITS-1, ITS-2) is minimal. TEM results coupled with PCR results confirmed that plastid primers can be used to amplify DNA from pollen of Juniperus. This can be used to study the role of plastid DNA in the quantification of airborne, allergenic pollen from various Juniperus sources. Published on-line www.phytologia.org Phytologia 98(4): 298-310 (Oct 6, 2016). ISSN 030319430.