Carbon stable isotope composition of DNA isolated from an incipient paleosol A. Hope Jahren* Department of Earth and Planetary Sciences, Johns Hopkins University, Baltimore, Maryland 21218, USA Kellie Kelm Beverly Wendland Department of Biology, Johns Hopkins University, Baltimore, Maryland 21218, USA Gitte Petersen Ole Seberg Botanical Institute, Department of Evolutionary Botany, University of Copenhagen, Gothersgade 140, DK-1123 Copenhagen K, Denmark ABSTRACT We determined the carbon isotope (␦13C) value of double-stranded DNA (dsDNA) iso- lated from the organic horizons of a Delaware soil that is actively being covered by an encroaching sand dune. The soil belongs to a Nymphaea odorata Ait. (water lily) wetland, and we regard its active acquisition of a thick (ϳ24 cm) surface mantle to embody the process of paleopedogenesis; therefore, we have termed it an ‘‘incipient paleosol.’’ In this study, we compared the ␦13C value of paleosol dsDNA to the bulk ␦13C value of N. odorata, as well as to the ␦13C value of plants that had colonized the surface mantle. The isotopic 13 13 offset between paleosol ␦ CdsDNA and N. odorata ␦ Ctissue was identical to the relationship 13 13 between ␦ CdsDNA and ␦ Ctissue for tracheophytes, which we had previously determined. 13 13 In contrast, the isotopic offset between paleosol ␦ CdsDNA and the ␦ Ctissue of plants colonizing the surface mantle differed from this relationship by as much as 4‰. Similarly, the ␦13C value of bulk paleosol organic matter was extremely heterogeneous and varied across 6‰. All paleosol DNA polymerase chain reaction (PCR) products produced clear, sharp, 350 base-pair (bp) fragments of rbcL, a gene shared by all photosynthetic organ- isms. These results open the exciting possibility that stable isotope analysis of dsDNA isolated from paleosol organic matter can be used to infer the ␦13C value of the plant that dominated the nucleic acid contribution. Keywords: paleosol, DNA, carbon, stable isotope. INTRODUCTION pared the carbon stable isotope offset between the inclusion of many herbs in addition to Paleosols preserve a myriad of characteris- plant nucleic acids isolated from paleosol or- trees (Table 1). Because all sampled plants tics that reflect the climate and environment ganic horizons and the paleovegetation sug- were under cultivation, they were largely of their time (e.g., Retallack, 2001). The car- gested by the geological context and palyno- shielded from stress, and many of these plants bon isotope (␦13C) value of paleosol bulk or- logical examination. We then compared the were growing within greenhouses. ganic matter is central to many characteriza- offset to our revised and taxonomically ex- The paleosol field site is located within ␦13 tions of paleoclimate; however, its isotopic panded data set of tracheophyte CDNA. Cape Henlopen State Park, Delaware (mean signal is extensively modified by processes of annual air temperature ϭ 13.3 ЊC; mean an- decomposition (e.g., Bowen and Beerling, nual precipitation ϭ 112.5 cm), at the frontal BOTANICAL GARDEN SAMPLING 2004), leading workers to increasingly focus margin of a large eolian sand dune that is ac- AND PALEOSOL SITE DESCRIPTION on the isolation and analysis of specific com- tively migrating westward. As it migrates, the Plant leaf tissues were collected from 12 pounds from bulk organics. Isolation of ge- dune encroaches on a freshwater wetland, cov- species selected in order to highlight mono- netic material from paleosol organic matter ering the organic-rich soils that formed in the cots and other groups not included in our pre- has successfully resulted in reconstructions of moist environment. The wetland is dominated vious work (Table 1; Jahren et al., 2004). The plant taxonomic status (Mitchell et al., 2005; by Nymphaea odorata Ait. (water lily); in Willerslev et al., 2003). Because soil organic total data set now contains four gymnosperms contrast, the encroaching sand dunes bring carbon is overwhelmingly dominated by con- (all conifers) and twenty angiosperms (five Oxydendrum arboretum (L.) DC. (swamp tributions from vegetation (e.g., Jobba´gy and monocots and fifteen dicots). Within the an- cranberry), Pinus rigida P. Mill (pitch pine), Jackson, 2000), we set out to test the potential giosperms, the magnoliid group and many and Acer rubrum L. (red maple) plants and of paleosol nucleic acids to act as an isotopic core eudicots were included, effectively sam- seedlings in large numbers. proxy for the dominant vegetation that was pling across the dicots. Plant species reported We excavated a profile set back from the growing in the soil. Toward this end, we com- here strategically augment the previously re- leading edge of the dune where 24 cm of sand ported sample set (Jahren et al., 2004), not had buried the wetland soil; below this surface *E-mail: [email protected]. only by the inclusion of monocots, but also by mantle we exposed three distinct soil horizons ᭧ 2006 Geological Society of America. For permission to copy, contact Copyright Permissions, GSA, or [email protected]. Geology; May 2006; v. 34; no. 5; p. 381–384; doi: 10.1130/G22408.1; 3 figures; 2 tables; Data Repository item 2006076. 381 TABLE 1. ␦13C VALUES OF PLANT SPECIES AND EXTRACTED NUCLEIC ACIDS ␦13 ␦13 Taxonomic Site of Latin name Common name A260/A280 Cof Cof group collection* bulk plant nucleic acids tissue Ϯ␴(‰)§ Ϯ␴(‰)§ Conifers ATL Cryptomeria japonica (L.f.) D. Don Japanese cedar 1.462 Ϫ27.44 Ϯ 0.06 Ϫ26.42 Ϯ 0.15 STR Cupressus sempervirens L.† Italian cypress 1.380 Ϫ29.68 Ϯ 0.07 Ϫ28.54 Ϯ 0.59 UCD Pinus canariensis C. Smith† Canary Island pine 1.270 Ϫ27.40 Ϯ 0.14 Ϫ26.11 Ϯ 0.40 UCR Pinus halepensis P. Mill.† Aleppo pine 1.600 Ϫ28.97 Ϯ 0.06 Ϫ27.90 Ϯ 0.29 Angiosperms Magnoliids UKB Atriplex littoralis L. Grassleaf orache 1.422 Ϫ28.02 Ϯ 0.04 Ϫ27.45 Ϯ 0.20 ATL Magnolia virginiana L.† Umbrella tree 1.610 Ϫ29.19 Ϯ 0.11 Ϫ27.38 Ϯ 0.06 Monocots UKB Acorus calamus L. Calamus 1.416 Ϫ30.11 Ϯ 0.04 Ϫ28.41 Ϯ 0.20 UKB Dietes grandiflora N.E. Br. Wild iris 1.418 Ϫ28.13 Ϯ 0.06 Ϫ27.26 Ϯ 0.01 UKB Dypsis lutescens (H. Wendl.) Yellow butterfly palm 1.453 Ϫ30.40 Ϯ 0.04 Ϫ28.85 Ϯ 0.22 UKB Lilium regale E.E. Wilson Regal lily 1.433 Ϫ27.67 Ϯ 0.05 Ϫ26.27 Ϯ 0.01 UKB Phalaris arundinacea L. Reed canarygrass 1.414 Ϫ28.02 Ϯ 0.06 Ϫ27.07 Ϯ 0.04 Rosids ATL Acer campestre L.† Hedge maple 1.710 Ϫ31.31 Ϯ 0.05 Ϫ29.25 Ϯ 0.08 ATL Aesculus sylvatica Bartr.† Painted buckeye 1.590 Ϫ31.85 Ϯ 0.14 Ϫ29.79 Ϯ 0.07 ATL Carya alba L.† Mockernut hickory 1.600 Ϫ28.15 Ϯ 0.04 Ϫ26.29 Ϯ 0.10 UCR Cneoridium dumosum Nutt.† Bush rue 1.470 Ϫ30.74 Ϯ 0.06 Ϫ29.41 Ϯ 0.24 UCD Eucalyptus robusta Sm. Swampmahogany 1.472 Ϫ28.66 Ϯ 0.11 Ϫ27.30 Ϯ 0.12 Asterids ATL Camellia sasanqua Thunb. Sasanqua camellia 1.870 Ϫ30.74 Ϯ 0.60 Ϫ30.03 Ϯ 0.16 STR Clethra mexicana L. Sweetpepperbush 1.444 Ϫ30.85 Ϯ 0.01 Ϫ29.86 Ϯ 0.33 ATL Ilex decidua Walt Possumhaw 1.425 Ϫ30.24 Ϯ 0.03 Ϫ28.62 Ϯ 0.39 ATL Kalmia latifolia L.† Mountain laurel 1.260 Ϫ29.83 Ϯ 0.11 Ϫ28.91 Ϯ 0.04 UCR Laurus nobilis L.† Sweet bay 1.150 Ϫ28.25 Ϯ 0.17 Ϫ27.67 Ϯ 0.06 FTG Manilkara zapota (L.) van Royen† Sapodilla 1.600 Ϫ30.25 Ϯ 0.13 Ϫ29.25 Ϯ 0.04 UCR Umbellaria californica L. Oregon myrtle 1.550 Ϫ29.25 Ϯ 0.17 Ϫ27.52 Ϯ 0.06 Other Eudicots ATL Hamamelis virginiana L.† American witch hazel 1.470 Ϫ30.26 Ϯ 0.06 Ϫ28.75 Ϯ 0.47 *Samples were collected at the following arboreta and botanical gardens during 2001 and 2003: Atlanta Botanical Gardens (ATL), University of Copenhagen Botanical Gardens (UKB), Fairchild Tropical Gardens (FTG), Strybing Arboretum and Botanical Gardens (STR), University of California at Davis Arboretum (UCD), University of California at Riverside Botanical Garden (UCR). †Reported in Jahren et al. (2004). §␴ is the standard deviation seen in three replicates of the sample. (Table 2). The uppermost horizon (1Oe) was (1Oe) and second (1A) horizons to comprise An examination of pollen isolated from hori- 18 cm thick and was composed of primarily a buried Typic Endoaquent. Radiocarbon dat- zons 1Oe and 1A revealed the presence of N. hemic organic materials. Mixed with the or- ing of fine-grained organic (Ͻ0.1 ␮m parti- odorata to the exclusion of other species. The ganic material was medium-grained quartz cles) yielded an age of 1900–present (Table third horizon (2Oe) was 13 cm thick and had sand, accounting for 20% of the bulk material 2), which is in keeping with maps of the mi- the highest organic content of the three hori- by mass. The second horizon (1A) was 11 cm gration of the shoreline (Division of Parks and zons; however, there was evidence of exten- thick and classified as a mineral horizon, with Recreation, Dover, Delaware, 2005, personal sive bioturbation within this horizon, includ- 80% of the bulk material as medium-grained commun.). Based on these features, the buried ing evidence of fluctuating water table. For quartz sand and the remainder made up of he- horizons comprise an incipient paleosol—a this study, we performed nucleic acid extrac- mic organic material. We interpret the first model for the inception of paleopedogenesis. tions from the 1Oe and 1A horizons because TABLE 2.