Reviews and Previews: the Future of Molecular Biology

Palaeontologia Electronica http://palaeo-electronica.org

THE FUTURE OF MOLECULAR PALEONTOLOGY

Molecular paleontology as a field ucts of their interactions with geochemi- engenders much controversy, particularly cals, should also be considered aspects of with respect to the recovery and analyses molecular paleontology. Molecular paleon- of truly ancient molecules, i.e. those more tology, then, could be defined as the study than tens of thousands of years old. In part of all biomolecules or their degradation this is due to empirical hypotheses regard- products that can be traced to their source ing the ultimate durability and survivability and that can shed light on the molecular of molecules in the fossil record, and in diagenetic history of an organism. part due to the problems of differentiating One assumption that has been perva- between endogenous molecules and sive in paleontological thought since the exogenous contamination. Molecular and inception of the science is that the organic chemical analytical methods are expen- constituents of an organism, namely soft sive and destructive to rare fossil material, tissues, cells, or the proteins and nucleic and usually, those fossils most amenable acids which were produced by its living to molecular analyses and most likely to cells, were either destroyed in the process yield positive results are those preserved of fossilization (Allison 1990), or rendered in an exceptional manner, thus making uninformative by the diagenetic changes them too valuable for destructive analy- accumulated during geological time (Curry ses. The value of molecular paleontology 1990). With advances in the fields of ana- and the techniques and methods it lytical biochemistry, molecular biology, and applies, then, is often called into question. geochemistry, it is becoming increasingly Molecular paleontology, like molecu- evident that this is not always the case, lar biology, has come to refer to the recov- and that there may be a wealth of informa- ery, analyses and characterization of DNA. tion to be gained through the study of However, the molecular record of an molecular fragments preserved in the fos- organism is certainly retained in molecules sil record. Examination of such molecules other than DNA, although these other life may strengthen the objectivity of the sci- molecules are variably informative with entific discipline of paleontology, as well as respect to phylogenies, evolutionary his- providing an independent means of testing tory and other characteristics. Analyses of phylogenetic hypotheses. proteins, carbohydrates, and lipids, as well Molecular paleontology in the modern as their degradation products and prod- sense probably began with the report by

Schweitzer, Mary Higby, 2003. Reviews and Previews: The Future of Molecular Biology. Palaeontologia Electronica, vol. 5, issue 2, editorial 2: 11pp., 177KB; http://palaeo-electronica.org. SCHWEITZER, MARY HIGBY: REVIEWS AND PREVIEWS: THE FUTURE OF MOLECULAR BIOLOGY

Abelson (1956) of the recovery of protein- and Griffen 1987), and a method for verify- aceous components of fossils. As technol- ing the endogeneity of proteinaceous ogy expanded and increased in accuracy, material using both racemization analyses sensitivity, and reliability, new analytical and stable isotope geochemistry has been methods began to be applied to fossil proposed (Macko and Engel 1991). material. Further attempts to identify endoge- In 1974, deJong et al. demonstrated nous molecules in fossil materials were the retention of the antigenic components undertaken by Lowenstein (1980, 1981, of proteins preserved within 70 Ma mol- 1985), who demonstrated that chemical lusk shells by precipitation reactions with extractions of fossil bone were amenable antisera. These results were supported by to immunological analyses in the form of immunogenic reactivity in other Creta- solid phase radio-immunoassays. He ceous fossil shells (Weiner et al. 1976; demonstrated antibody binding to extracts Westbroek et al. 1979). Amino acid analy- of fossil material from a variety of bone ses undertaken by Armstrong et al. (1983) samples, including human, which dated to showed the presence of amino acids in a two million years BP. Based upon his variety of bone samples, and in 1991, Gur- results, he proposed utilizing immunologi- ley et al. reported the isolation and identifi- cal methods to elucidate phylogenetic cation of amino acids in the bony tissues relationships of extinct organisms (Lowen- of the sauropod dinosaur Seisomosaurus. stein 1985, 1988). It is now widely Identification of amino acids within accepted that DNA and proteins may be fossil materials does not necessarily imply, retained in recent fossils or subfossils, however, that those amino acids are although there is much skepticism regard- derived from original and ancient proteins, ing such preservation in fossils tens of mil- as this method does not differentiate lions of years old. between endogenous molecules and Early work seeking to identify proteins those that may have accumulated at any preserved in fossil material focused on the point during diagenesis. Because all identification of collagen, because the organisms on this planet use only the L presence of collagen can be verified by form of amino acids to build proteins, and electron microscopy, owing to its unique these L-amino acids racemize to an equi- cross-banded pattern (Van der Rest librium mixture of D and L isomers after 1991). However, it was also shown that death, chiral analyses of amino acids has even preservation at this level of micro- been suggested as a means of ruling out structure does not necessarily indicate the the possibility of modern contamination, as presence of endogenous molecules, as a preponderance of L-amino acids may be collagen-specific amino acids hydroxypro- indicative of an extant or recent origin of line and hydroxylysine were not identified protein fragments (Schroeder and Bada in samples in which collagen cross-band- 1976). In addition, since each amino acid ing could be visualized (Towe and racemizes to completion at a different rate, Urbanek 1972). it was hypothesized that the degradation While demonstrating the presence of of amino acids to a racemic mixture of amino acids or identifying structural pro- their D/L isomers may be linked to the age teins was the goal of early attempts at of specimen (Schroeder and Bada 1976; molecular paleontology, the recognition Bada 1985). This latter proposal has been that some molecules were very durable met with some controversy (e.g., Kimber and that some may have better survival

2 SCHWEITZER, MARY HIGBY: REVIEWS AND PREVIEWS: THE FUTURE OF MOLECULAR BIOLOGY

potential than others (e.g., Runnegar 1990, Dolittle et al. 1996). These methods 1986) led to the search for other, perhaps provide the investigator with a means to more informative proteins, as well as non- establish the endogeneity of ancient DNA proteinaceous material. In addition to col- by placing recovered sequences in correct lagen (e.g., Baird and Rowley 1990), pro- phylogenetic contexts. teins such as IgG and albumin (Tuross The prevailing scientific opinion has 1989; Cattaneo et al. 1992) have been long held that DNA is unstable and easily shown to be preserved in fossil bone, and degraded; therefore its presence in tissue the vertebrate-specific protein osteocalcin samples much older than a hundred thou- has been identified in both tooth and bone sand years is highly suspect (Curry 1990; samples (Ulrich et al. 1987) including Lindahl 1993). As our understanding of the those of dinosaurs (Muyzer et al. 1992). chemical nature of this molecule Hemoglobin, the protein involved in oxy- increases, it is becoming evident that cer- gen transport, has also shown potential for tain factors act to stabilize DNA, thus sig- preservation in the fossil record, having nificantly increasing its longevity. been identified in association with stone Desiccation, protection from oxidative tools (Loy 1983, 1987; Loy and Wood damage through rapid burial, and pres- 1989) as well as ancient bone samples ence of a mineral substrate to which the (Ascenzi 1985; Smith and Wilson 1990; molecule may adsorb and thus become Cattaneo et al. 1990). Hemoglobin is stabilized, all enhance the preservation important both as an indicator of physiol- potential of DNA (Eglington and Logan ogy (Dickerson and Geis 1983) and for 1991; Tuross 1994). More efficient means studies in phylogenetic divergence (Perutz of extraction (Hoss and Pääbo 1993), cou- 1983; Nikinmaa 1990; Gorr 1998), and the pled with the use of chemical agents to possibility of its presence in dinosaur bone free DNA from complexes of degradation (Schweitzer et al. 1999) may shed light products (Poinar et al. 1998) also increase upon questions of metabolic rates, as well our chances of success in the identifica- as the relationship of these animals to tion and recovery of endogenous mole- modern taxa. cules from the fossil record. The advent of Despite these intriguing results, how- the polymerase chain reaction (PCR) ever, the ultimate success of molecular opened the door to the possibility that paleontology is viewed by some to be the DNA may indeed be recovered from very identification and recovery of DNA old fossils because this reaction makes it sequences from extinct taxa. Of all the bio- possible to amplify small and degraded or molecules produced by an animal, DNA altered DNA fragments that perhaps would contains the most phylogenetic informa- not be suitable for cloning. However, the tion in its sequences. Data bases now sensitivity of PCR creates problems in the exist that allow comparison of sequences analysis of ancient specimens, the most obtained from fossil specimens with those notable of which are the ease with which of extant taxa (e.g., Handt et al. 1994; modern contaminating molecules are Cooper 1994; Erlich et al. 1991; Pääbo et amplified and the misleading results due al. 1989) to test phylogenetic hypotheses to template damage in ancient samples (e.g., Felsenstein 1981, 1993; Kumar and (Pääbo et al. 1990; DeSalle et al. 1993; Hedges 1998) and to infer evolutionary Handt et al. 1994). distance (Lewontin 1989; Hedges et al.

3 SCHWEITZER, MARY HIGBY: REVIEWS AND PREVIEWS: THE FUTURE OF MOLECULAR BIOLOGY

Molecular preservation Microbes produce extracellular polymeric Mechanisms for the preservation of substances that trap minerals, thus con- organic compounds such as DNA or pro- tributing to cementation and their own fos- tein over the course of geological time silization as well as the mineralization or remain to be elucidated. However, it has "fossilization" of components in the envi- been proposed that these compounds, in ronment. Microbial influences have been the process of degradation and bond invoked in the formation of early diage- breakage, may co-react to form complex netic concretions around biological speci- biopolymers which resist further degrada- mens, which can lead to extraordinary tion (Curry 1990). preservation of macro- and microstructure, A second mechanism which has been occasionally including soft tissues (Can- proposed for preservation of biomolecular field and Raiswell 1991). materials is the stabilization of these mole- Finally, it is noted in the literature that cules through complex interactions with a primary factor in preserving both pro- organic breakdown products of the sur- teins and nucleic acids over geological rounding soils, in particular humic or fulvic time may be the association of these pro- acids (Tuross 1994). These associations, teins and/or nucleic acids to a mineral sub- while an important factor in the preserva- strate, such as is found in bone (Runnegar tion of biomolecules, are also deleterious 1986; Tuross et al. 1989; Ambler and from an analytical standpoint. Separating Daniel 1991; Logan et al. 1991). Adsor- out the endogenous components from the bance of biomolecules to minerals may be rest of the aggregation in order to perform among the most important of mechanisms various analyses has proven difficult, involved in biomolecular preservation. although the compound N-phenacylthiazo- Preservation potential is enhanced in lium bromide (PTB; Poinar et al. 1998) has biomineralized tissues because there is a been demonstrated to be effective in component of protein that is encased cleaving glycosidic bonds involved in within the mineral crystals, creating a these molecular aggregates, freeing the closed system (Weiner et al. 1989; Sykes components of interest. In addition, humic et al. 1995). acids fluoresce at the wavelengths of No doubt, the preservation of biomole- some proteins, amino acids, or nucleic cules over the course of geological time is acids (Tuross and Stathoplos 1993), and enhanced by a combination of the above may therefore interfere with or mask indig- mechanisms to varying degrees and, most enous biomolecular signals. Finally, these likely, there are other interactions involved breakdown products inhibit the action of in molecular preservation that have yet to some enzymes that may be used to iden- be identified. However, there is little exper- tify organic remains (Tuross 1994), such imental evidence for a temporal limit to as digestive enzymes or the polymerase preservation enhanced by such mecha- enzymes used in PCR. nisms. Another factor contributing to the pres- The fossil record is capricious in its ervation of endogenous biomolecules is preservation. Whereas most fossils are the early cementation of surrounding or well permineralized, individual specimens entombing sediments, often facilitated by can show little evidence of permineraliza- microbes. This cementation creates a vir- tion, which may be an indication of mini- tually closed system that greatly reduces mal water infiltration. Additionally, exogenous degradation processes. surprisingly delicate structures, such as

4 SCHWEITZER, MARY HIGBY: REVIEWS AND PREVIEWS: THE FUTURE OF MOLECULAR BIOLOGY

feather barbules or embryonic tissues, can table the question "will advances in molec- sometimes be seen, and, in fossil Lager- ular paleontology ever allow us to statten such as the Messel Shale (Schaal resurrect the dinosaurs"? This is a ques- and Ziegler 1988), pigment, hair, and indi- tion that paleontologists surely face often vidual sarcomeres of muscle fibers have since the release of the movie series, and been preserved. Such intricately pre- it is to the advantage of students of dino- served specimens can reveal additional saur paleontology to understand the details which contribute to our understand- issues involved, and to have a clear and ing of how extinct organisms lived, looked concise answer ready when asked. and functioned. For example, the impres- While no one can predict the future or sions of feathers in the burial sediments the directions in which advances in tech- surrounding Archaeopteryx led to its nology will lead us, my answer to the placement in the bird lineage, while the question of dinosaur cloning is a definitive presence of feathers and other integumen- "no". There are several reasons beyond tary structures in exceptionally preserved technological problems that lead to this dinosaurs (e.g., Chen et al. 1998; Quiang conclusion. First, the successful cloning of et al. 1998; Mayr et al. 2002) not only sup- a dinosaur requires the recovery of DNA. port the phylogenetic link between dino- Proteins, lipids or carbohydrates are insuf- saurs and birds (e.g., Gauthier 1986; ficient to direct the ontogeny of a living Sereno 1997), but may also suggest being, as they are simply the indirect or increased metabolic strategies in this direct products of information stored in the group of dinosaurs (Schweitzer and Mar- base sequence of DNA. A full complement shall 2001), consistent with the hypothesis of DNA is needed to produce a functioning that the origin of birds lies within the Dino- being. In humans, there are more than sauria. The discovery of oviraptor eggs 108 base pairs of DNA, coding for more (Norell et al. 1994) and sauropod eggs than 30,000 genes, all arranged in a spe- (Chiappe et al. 1998) containing delicate cific order and distributed among 46 chro- embryonic tissues may help to illuminate mosomes, and each one is necessary to parental behaviors among dinosaurs, and produce a functioning human, orders of may shed light on aspects of ontogeny in magnitude more than the 200–500 bases these taxa. However, little has been done of endogenous DNA that has been recov- until lately to examine the possibilities of ered from fossil material. Additionally, if preservation of the molecules that consti- these genes become rearranged, or if a tuted the fossil organisms. This may be chromosome is lost, or if something else due in part to the rarity of appropriate fos- happens to alter the ORDER of base sil finds, which precludes destructive anal- pairs, genes, or chromosomes, it is usually yses, and in part to the fact that lethal, and almost always severely delete- adaptations of technologies developed for rious, greatly affecting the survivability of the field of molecular biology have only the organism. recently been applied to fossil specimens. We have no way of knowing how many genes or chromosomes each dino- Where do we go from here? saur taxon had, and we cannot deduce the The popularity of the "Jurassic Park" order in which genes were arranged upon series of books and movies, and the individual chromosomes. The order of enduring magical fascination dinosaurs arrangement is absolutely critical to the holds for the general public, makes inevi- development of an organism from a single

5 SCHWEITZER, MARY HIGBY: REVIEWS AND PREVIEWS: THE FUTURE OF MOLECULAR BIOLOGY

fertilized cell to a multicellular functioning dilia, gender is determined by the temper- being. ature of incubation (Bull 1983; Woodward While the idea of filling in this "miss- and Murray 1993). Eggs laid closer to the ing" genetic information with genes from perimeter of a nest are subjected to colder living, related organisms, as presented in temperatures, and are usually of a differ- the movie "Jurassic Park", is intriguing, to ent gender than those incubated within the be successful these genes would have to middle of the nest (Woodward and Murray be almost identical to those of the original 1993). In all living birds, on the other hand, dinosaurs, containing the same informa- sex of offspring is determined genetically, tion and dictating the same functions. We and usually, though not always, the genes do not possess the information needed to determining gender are located on specific deduce that, and by far the greatest likeli- chromosomes. We do not know which hood is that we would end up with a mess reproductive strategy the dinosaurs pos- of genetic "soup" that would be utterly sessed. non-functional. Additionally, we do not know the spe- However, suppose that we could cific hormones involved, their timing, or recover, either from the fossil record, or by the amounts needed to turn on and off the piecing together the information from living genes of development. The divergence of taxa, the total DNA to encode a dinosaur, the dinosaur–bird lineage from the ances- and that it was arranged in the proper tors of today’s crocodiles is estimated to order, upon the exact number of chromo- have taken place about 230–250 Ma in the somes. Would we have what we needed past (Carroll 1988). That means that there then to "grow" a dinosaur? No, it is much would be at least 460 million years of inde- more complicated than that, as we would pendent evolution between a living croco- still need an environment in which this dile and extant birds, with dinosaurs genetic information could develop. diverging at some point along this contin- In all living animals, development is uum (see Figure 1). There is certainly no directly influenced by hormonal cues from way of knowing if today’s crocodiles are the mother. In egg-laying animals, these adequate hosts, at the molecular level, for cues are contained in the yolk and mem- the development of a resurrected dino- branes deposited with the embryo. In ani- saur. Similarly, birds diverged from dino- mals that develop internally, the cues are saurs, it is assumed, at least 150 million provided as a continuous flux through the years ago. Can we possibly reconstruct circulatory system, delivered over time to the molecular environments in which a the developing embryo. To "grow" a dino- dinosaur embryo developed? saur, we would have to take the packet of But suppose we could overcome complete genetic information and insert it these hurdles, and get a viable dinosaur to into the enucleated egg of a closely hatch. What would it eat? The enzymes in related taxon, such as a bird or crocodile, its digestive tract would have evolved for then place that egg within the host ani- specific foodstuffs that in all likelihood no mal’s reproductive tract so that it could longer exist and have not left any living access the necessary external cues descendants. If it is a carnivorous dino- needed to develop. However, the hor- saur, could its digestive enzymes break monal and environmental cues for devel- down the components of mammalian tis- opment vary greatly within living members sues? Or would byproducts of digestion be of the closest dinosaur relatives. In Croco- toxic? If it is an herbivore, would today’s

6 SCHWEITZER, MARY HIGBY: REVIEWS AND PREVIEWS: THE FUTURE OF MOLECULAR BIOLOGY

Figure 1. From the point of divergence, based upon the first appearance of a "true" dinosaur in the Triassic, extant birds and crocodiles have undergone approximately 460 million years of independent evolution.

plants give it sufficient nutrition, or would it fragment of DNA (e.g., the hemoglobin possess the enzymes needed to extract gene) with 40 informative sites could be the nutrients from the plant tissues? recovered from exceptionally preserved These enormous hurdles would have bone tissues of a Velociraptor, it would be to be dealt with in attempts at resurrecting possible to align the dinosaur gene region any extinct animal. They are much easier with the comparable region of extant croc- to address, however, with taxa like the odiles and birds, and to identify the types mammoth, having extant relatives, living of changes between gene sequences (Fig- elephants, that are extremely closely ure 2). In addition, by comparing small related, and where not much time has regions of genes for changes, one could elapsed since their divergence and/or infer the closeness of relationships extinction. The farther back in time, the between extinct taxa and their extant more difficult these problems become. descendants (e.g., Cooper 1994). Therefore, if one’s goal in the study of Another advantage to studying molec- molecular paleontology is to resurrect ani- ular fragments preserved within fossil tis- mals that have become extinct, the future sues would be to date absolutely the of the science is bleak. timing and direction of genetic changes within taxa, because it would be rooted by Beyond cloning the absolute date for the fossil (e.g., van Even if the likelihood of building a Tuinen and Hedges 2001). This would "real" Jurassic park is virtually non-exis- give us an idea of how long it took genetic tent, there are many important and inter- changes to accumulate in a lineage, as esting questions that can be addressed by well as allowing us to infer the number of applying molecular techniques to fossil individual evolutionary events (Lewontin specimens. For appropriately preserved 1989). fossil material that still retains usable The study of remnant molecules in molecular information, it may be possible fossils also allows us to understand the to compare fragments of molecules with processes of molecular diagenesis, or those of close living relatives to estimate changes that accumulate in the molecule the rate and direction of evolutionary as the result of degradation, modification, change. For example, if a 200 base-pair and interaction with geochemical residues,

7 SCHWEITZER, MARY HIGBY: REVIEWS AND PREVIEWS: THE FUTURE OF MOLECULAR BIOLOGY

Figure 2. Alignment of a small region of the hemoglobin gene from chicken and crocodiles, compared with a hypo- thetical region of recovered dinosaur material. Asterisks represent base changes in the crocodile and dinosaur rela- tive to the chicken.

and factors within the depositional envi- evidence is accumulating that biomole- ronment that contribute to the preservation cules, fragments of molecules, or their of these same molecules. degradation products can indeed be pre- All biomolecules break down over served over geological time scales. Better time, owing to the action of autolytic understanding of the processes of molecu- enzymes, microbial influences, oxidation, lar degradation and fossilization, as well hydrolytic damage, or intra- or intermolec- as the processes of biomineralization at ular crosslinking. In addition, DNA mole- the molecular level, are shedding light on cules can become depurinated or more efficient means of extracting mole- deaminated, or the sugar–phosphate cules from fossils (Poinar et al. 1998), backbone can be cleaved, leaving frag- which types of molecules may be the best ments (Curry 1990). Likewise, proteins targets for molecular investigations, and can be denatured to primary structure. on which fossils in which environments Once this occurs, original amino acids can may be most appropriate for molecular convert to others, combine and form investigations. cross-links, or lose R-groups completely, In short, molecular paleontology is a leaving any amino acid altered to glycine. new field, the potential of which is only Amino acids can also undergo polycon- beginning to be realized. As technologies densations, through processes such as advance, we will no doubt be able to Amadori rearrangements or Maillard reac- recover more and more information from tions, leaving insoluble residues contain- the physical remains of animals long ing parts of the original molecules within extinct. The pursuit of this knowledge is complexes containing other organic solu- valuable, and will aid in our understanding tions. If one knows the starting molecules, of evolutionary processes, as well as the these chemical changes may be able to be processes of fossilization, particularly at elucidated and quantified through the the molecular level. Additionally, such recovery of molecules from fossils. In studies will clarify our understanding of the addition, understanding the type of stages in the breakdown and modification changes molecules undergo will allow us of molecules over time, thus allowing us to to predict the chances of molecular recov- link preserved molecular markers in the ery of fossils in various environ- fossil record with their source molecules. ments.Summary Finally, understanding molecular diagene- While we will almost certainly never sis across geological time scales and rec- clone a Tyrannosaurus rex, swim with a ognizing preserved biomarkers from the giant plesiosaur, or breathe new life into fossil record may aid in our search for evi- the pterosaurs, the field of molecular pale- dence of life on other planets. The search ontology has much to offer. Despite the for extraterrestrial life rests on three possi- technical problems inherent in dealing with bilities, namely, life may never have ancient biomolecules and their derivatives, existed, life may have existed for a short

8 SCHWEITZER, MARY HIGBY: REVIEWS AND PREVIEWS: THE FUTURE OF MOLECULAR BIOLOGY time, then gone extinct, or life may be cur- Cattaneo, C., Gelsthorpe, K., Phillips, P., and Sokol, R.J. rently thriving. If the second situation 1990. Blood in ancient human bone. Nature, 347:339. occurs, all that may be left as evidence are Cattaneo, C., Gelsthorpe, K., Phillips, P., and Sokol, R.J. resistant molecular markers that are 1992. Detection of blood proteins in ancient human unique to life. We must be able to recog- bone using ELISA: a comparative study of the survival of IgG and albumin. International Journal of nize the range of diagenetic alteration of Osteoarchaeology, 2:103-107. biomolecules across time on this planet in Chen, P.-J., Dong, Z., and Zhen, S.-N. 1998. An excep- order to detect them on other planets, tionally well-preserved theropod dinosaur from the where life may have gotten a tenuous Yixian Formation of China. Nature, 391:147-152. Chiappe, L.M., Coria, R.A., Dingus, L., Jackson, F., Chin- start, and then became extinct. Molecular samy, A. and Fox, M. 1998. Sauropod dinosaur paleontology has much to contribute to the embryos from the Late Cretaceous of Patagonia. search for life on other planets, in addition Nature, 396:258-261. to addition to our understanding of the Curry, G.B. 1990. Molecular palaeontology. p. 95-100. In Briggs, D.E.G. and Crowther, P.R. (eds), Palaeobiol- evolution and extinction of life on this one. ogy: A Synthesis. Blackwell Scientific Publications, Oxford. REFERENCES deJong, E.W., Westbroek, P., Westbroek, J.F., and Brun- ing, J.W. 1974. Preservation of antigenic properties Abelson, P.H. 1956. Paleobiochemistry. Scientific Ameri- of macromolecules over 70 Myr. Nature, 252:63-64. can, 195:83-92 DeSalle, R., Barcia, M., and Wray, C. 1993. PCR jump- Allison, P.A. 1990. Decay Processes, p. 213-216. In ing in clones of 30-million-year-old DNA fragments Briggs, D.E.G. and Crowther, P.R. (eds), Palaeobiol- from amber preserved in termites (Mastotermes elec- ogy: A Synthesis. Blackwell Scientific Publications, trodominicus). Experientia, 49:906-909. Oxford. Dickerson, R.E., and Geis, I. 1983. Hemoglobin: Struc- Ambler, R.P., and Daniel, M. 1991. Proteins and molecu- ture, Function, Evolution, and Pathology. Benjamin/ lar palaeontology. Philosophical Transactions of the Cummings, Menlo Park, CA. Royal Society of London B, 333:381-389. Dolittle, R.F., Feng, D.-F., Tsang, S., Cho G., and Little, Armstrong, W.G., Halstead, L.B., Reed, F.B. ,and Wood, E. 1996. Determining divergence times of the major L. 1983. Fossil proteins in vertebrate calcified tis- kingdoms of living organisms with a protein clock. sues. Philosophical Transactions of the Royal Soci- Science, 271:470-477. ety of London B, 301:301-343. Eglinton, G., and Logan, G.A. 1991. Molecular preserva- Ascenzi, A., Brunori, M., Citro, G., and Zito, R. 1985. tion. Philosophical Transactions of the Royal Society Immunological detection of hemoglobin in bones of of London B, 333:315-328. ancient Roman times and of Iron and Eneolithic Erlich, H.A., Gelfand, D., and Sninsky, J.J. 1991. Recent Ages. Proceedings of the National Acadademy of advances in the polymerase chain reaction. Science, Sciences USA, 82:7170-7172. 252:1643-1651. Bada, J.L. 1985. Amino acid racemization dating of fossil Felsenstein, J. 1981. Evolutionary trees from DNA bones. Annual Revues Earth and Planetary Sci- sequences: a maximum likelihood approach. Journal ences, 13:241-268. of Molecular Evolution, 17:368-376. Baird, R.F., and Rowley, M.J. 1990. Preservation of Felsenstein, J. 1993. Phylip: Phylogeny Inference Pack- avian collagen in Australian Quaternary cave depos- age. (University of Washington, Seattle.) its. Palaeontology, 33(2):447-451. Gauthier, J. 1986. Saurischian monophyly and the origin Bull, J.J. 1983. Evolution of Sex Determining Mecha- of birds, p. 1–55. In Padian, K. (ed.), The Origin of nisms. Benjamin/Cummings, Menlo Park, CA. Birds and the Evolution of Flight. California Academy Canfield, D.E., and Raiswell, R. 1991. Carbonate precip- of Sciences, Memoirs, No. 8.Gorr, T.A., and Klein- itation and dissolution, p. 411-453. In Allison, P.A. schmidt, T. 1993. Evolutionary relationships of the and Briggs, D.E.G. (eds), Taphonomy: Releasing the coelacanth. American Scientist, 81:72-82. Data Locked in the Fossil Record. Plenum Press, NY. Gurley, L., Valdez, J.G, Spall, W.D., Smith, B.F., and Carroll, R.L. 1988. Vertebrate Paleontology and Evolu- Gillette, D.D. 1991. Proteins in the fossil bone of the tion. W.H. Freeman, New York. dinosaur, Seismosaurus. Journal of Protein Chemis- Cooper, A. 1994. Ancient DNA sequences reveal unsus- try, 10(1):75-90. pected phylogenetic relationships within New Handt, O., Hoss, M., Krings, M., and Pääbo, S. 1994. Zealand wrens (Acanthisittidae). Experientia, 50:558- Ancient DNA: methodological challenges. Experien- 563. tia, 50:524-528. Hedges, S.B., Moberg, K.D., and Maxson, L.R. 1990. Tetrapod phylogeny inferred from 18s and 28s ribo-

9 SCHWEITZER, MARY HIGBY: REVIEWS AND PREVIEWS: THE FUTURE OF MOLECULAR BIOLOGY

somal RNA sequences and a review of the evidence Norell, M.A, Clark, J.M., Chiappe, L.M., and Dashzeveg for amniotic relationships. Mol. Biol. Evol. 7:607-633. D. 1994. A nesting dinosaur. Nature, 378:774-776. Hoss, M., and Pääbo, S. 1993. DNA extraction from Pääbo, S., Higuchi, R.G., and Wilson, A.C. 1989. Ancient Pleistocene bones by a silica-based purification DNA and the polymerase chain reaction. Journal of method. Nucleic Acids Research, 21(16):3913-3914. Biological Chemistry, 264(7):9709-9712. Kimber, R.W.L. Griffen, C.V. 1987. Further evidence of Pääbo, S., Irwin, D.A., and Wilson, A.C. 1990. DNA the complexity of the racemization process in fossil damage promotes jumping between templates during shells with implications for amino acid racemization enzymatic amplification. Journal of Biological Chem- dating. Geochimica et Cosmochimica Acta, 51:839- istry, 265(8):4718-4721. 846. Perutz, M.F. 1983. Species adaptation in a protein mole- Kumar, S., and Hedges, S.B. 1998. A molecular times- cule. Molecular Biology and Evolution, 1(1):1-28. cale for vertebrate evolution. Nature, 392:917-920. Poinar H.N., Hofreiter M., Spaulding W.G., Martin P.S., Lewontin, R. 1989. Inferring the number of evolutionary Stankiewicz B.A., Bland H., Evershed R.P., Possnert, events from DNA coding sequence differences. G., and Pääbo S. 1998. Molecular coproscopy: dung Molecular Biolology and Evolution, 6(1):15-32. and diet of the extinct ground sloth Nothrotheriops Lindahl, T. 1993. Instability and decay of the primary shastensis. Science, 281:402-406 structure of DNA. Nature, 362:709-715. Qiang, J., Currie, P.J., Norell, M.A., and Shu-An, J. 1998. Logan, G., Collins, M.J., and Eglinton, J. 1991. Preserva- Two feathered dinosaurs from northeastern China. tion of organic biomolecules, p. 1-24. In Allison, P.A. Nature, 393:753-761. and Briggs, D.E.G. (eds), Taphonomy: Releasing the Runnegar, B. 1986. Molecular palaeontology. Palaeon- Data Locked in the Fossil Record. Plenum Press, NY. tology, 29:1:1-24 Lowenstein, J.M. 1980. Species-specific proteins in fos- Sereno, P.C. 1997. The origin and evolution of dino- sils. Naturwissenschaften 67:343-346. saurs. Annual Revues Earth and Planetary Sciences, Lowenstein, J.M. 1981. Immunological reactions from 25:435-490. fossil material. Philosophical Transactions of the Schaal, S., and Ziegler, W., (eds.) 1988. Messel—ein Royal Society of London B, 292:143-149. Schaufenster in die Geschichte der Erde und des Leb- Lowenstein, J.M. 1985. Molecular approaches to the ens. Springer, Frankfurt.Schroeder, R.A., and Bada, J.L. identification of species. American Scientist, 73:541- 1976. A review of the geochemical applications of the 546. amino acid racemization reaction. Earth-Science Lowenstein, J.M. 1988. Immunological methods for Revues, 12:347-391. determining phylogenetic relationships, p. 12-19. In Schweitzer M.H., Watt, J.A., Avci, R., Knapp, L., Chi- Broadhead, T.W. (ed.), Molecular Evolution and the appe, L., Norell, M., and Marshall, M. 1999. Beta-ker- Fossil Record: Short Courses in Paleontology, Num- atin specific immunological reactivity in feather-like ber 1. The Paleontological Society. structures of the Cretaceous Alvarezsaurid, Shuv- Loy, T.H. 1983. Prehistoric blood residues: Detection on uuia deserti. Journal of Experimental Zoology (Mol. tool surfaces and identification of species of origin. Dev. Evol), 285:146-157. Science, 220:1269-1271. Smith, P.R., and Wilson, M.T. 1990. Detection of haemo- Loy, T.H., and Wood, A.R. 1989. Blood residue analysis globin in human skeletal remains by ELISA. Journal at Cayonu Tepesi, Turkey. Journal of Field Archaeol- of Archaeological Science, 17:255-268. ogy, 16:451-460. Sykes, G.A., Collins, M.J., Walton, D.I. 1995. The signifi- Loy, T.H. 1987. Recent advances in blood residue analy- cance of a geochemically isolated intracrystalline sis, p. 57-65. In Ambrose, W.R., and Mummer, J.M.J. organic fraction within biominerals. Organic (eds), Archaeometry: further Australasian Studies. Geochemistry, 23(11-12):1059-1065. Australian National University, Canberra. Towe, K.M., and Urbanek, A. 1972. Collagen-like struc- Macko, S.A., and Engel M.H. 1991. Assessment of indi- tures in Ordovician graptolite periderm. Nature, geneity in fossil organic matter: amino acids and sta- 237:443-445. ble isotopes. Philosophical Transactions of the Royal Tuross, N. 1989. Albumin preservation in the Taima- Society of London B, 333:367-374. taima mastodon skeleton. Applied Geochemistry, Mayr, G., Peters, D.S., Plodowski, G., and Vogel, O. 4:255-259. 2002. Bristle-like integumentary structures at the tail Tuross, N. 1994. The biochemistry of ancient DNA in of the horned dinosaur Psittacosaurus. Naturwissen- bone. Experientia, 50:530-535. schaften, 89:361-365. Tuross, N., and Stathoplos, L. 1993. Ancient proteins in Muyzer, G., Sandberg, P., Knapen, M.H.J., Vermeer, C., fossil bones. Methods in Enzymology, 224:121-129. Collins, M., and Westbroek, P. 1992. Preservation of Tuross, N. 1991. Recovery of bone and serum proteins the bone protein osteocalcin in dinosaurs. Geology, from human skeletal tissue: IgG, osteonectin, and 20:871-874. albumin, p. 51-54. In Ortner, D.J., and Aufderheide, Nikinmaa, M. 1990. Vertebrate Red Blood Cells. A.C. (eds), Human Paleopathology: Current Synthe- Springer, Berlin.

10 SCHWEITZER, MARY HIGBY: REVIEWS AND PREVIEWS: THE FUTURE OF MOLECULAR BIOLOGY

ses and Future Options. Smithsonian Institution Weiner, S., Traub, W., Elster, H., and DeNiro, M.J. 1989. Press, Washington, DC. The molecular structure of bone and its relation to Ulrich, M.M.W., Perizonius, W.R.K., Spoor, C.F., Sand- diagenesis. Applied Geochemistry, 4:231-232. berg, P., and Vermeer, C. 1987. Extraction of osteo- Westbroek, P., Van der Meide, P.H., van der Wey-Klop- calcin from fossil bones and teeth. Biochemical and pers, J.S., van der Sluis, R.J., de Leeuw, J.W., and Biophysical Research Communications, 149(2):712- de Jong, E.W. 1979. Fossil macromolecules from 719. cephalopod shells: characterization, immunological Van der Rest, M. 1991. The collagens of bone. In Hale, response and diagenesis. Paleobiology, 5(2):151- B.K. (ed.), Bone Matrix and Bone Specific Products. 167. Bone. 3:187-237. CRC Press, Boca Raton. Woodward, D.E., and Murray, J.D. 1993. On the effect of van Tuinen M., and Hedges S.B. 2001. Calibration of temperature-dependent sex determination on sex avian molecular clocks. Molecular Biology and Evolu- ratio and survivorship in crocodilians. Proceedings of tion, 18(2):206-13 the Royal Society of London, 252:149-155. Weiner, S, Lowenstam, HA , Hood, L, 1976. Character- ization of 80-million-year-old mollusk shell proteins. Mary Higby Schweitzer Proceedings of the National Acadademy of Sciences USA, 73(8):2541-2545.