Journal of Foraminiferal Research, v. 2 I , no.

2, p, 159- 167, April 1991

TOWARD A CLASSIFICATION OF PLANKTONIC FORAMINIFERA BASED ON
BIOCHEMICAL, GEOCHEMICAL, AND MORPHOLOGICAL CRITERIA
LISAL. ROBBINS’AND NANCYHEALY-WILLIAMS~

organism because their amino acid sequence is encoded by DNA (the genotype). Genetic information may
be discerned from the protein make-up of the organic
matrix found in the CaCO, test of planktonic foraminifera (Abelson, 1955; Hare, 1969; King and Hare,
1972). Biochemical analyses by Robbins (1 987) and
Robbins and Brew (1990) have also shown that the
tests contain preserved proteins. This paper represents
a first attempt to integrate several new approaches for
obtaining genetic information from Recent planktonic
foraminifera.
Planktonic foraminifera, with their nearly continuous record of preservation in deep sea sediments, exhibit morphologic variations through time (Malmgren
and Kennett, 198I , 1982; Gary and Healy-Williams,
1987) and are ideal fossils for testing theories about
the tempo and mode of evolution. By analyzing discrete, preserved proteins and polypeptides from the
shells of fossil planktonic foraminifera and integrating
these data with quantitative morphology and stable
isotope data, we hope to clarify taxonomic distinctions.
The ultimate goal of this more sophisticated taxonomical approach is to help refine the biostratrigraphic
framework based on planktonic foraminifera.
Understanding the environmental parameters responsible for phenotypic variation of planktonic foraminifera is another area where biochemistry could
make an important contribution (Bolli, 1950; Kennett,
1968; Malmgren and Kennett, 1972; B6 and others,
1973, among others). The use of morphology as a paleoecologic indicator assumes that planktonic foraminifera are generally panmictic, Le., without isolated
populations. Recent work by Healy-Williams and others (1 985) and Healy-Williams ( 1 988) questions this
assumption by revealing significant stable oxygen and
carbon isotopic differences between morphotypes of
Globorotalia truncatulinoides and Neogloboquadrina
pachyderma. Recent work suggest that the “morphotype effect” may be the result of genetic differentiation
within populations, although this does not rule out the
possibility that the isotopes may be measuring environmental differencesas well (Healy-Williams and others, 1985; Robbins, 1987; Williams and others, 1988a).
Biochemical analysis of the morphotypes in terms of
protein differences could supply the necessary information to test the hypothesis that these populations
are discrete.

ABSTRACT

New data on the protein composition of the organic
matrix from tests of six species of planktonic foraminifera demonstrate that the amino acid composition of
single proteins from fossil shell matrices provides information on genetic relatedness. We present here preliminary results which utilize an integrated approach
to decipher intraspecific variation of planktonic foraminifera. Protein biochemical data from two morphotypes of Globigerinoides ruber provide an example of
how quantitative morphology, stable isotope analysis,
and biochemical analysis can provide the basis for an
integrated approach to foraminiferal taxonomy. This
combined approach could lead to a classification framework that more closely reflects genotypic relationships
than does the standard morphologic approach.

INTRODUCTION
Taxonomic ambiguities are the bane of paleontology
and impede our understanding of evolutionary processes, paleobiological relationships, and paleoceanographic problems. In the taxonomy of Foraminiferida,
one of the dominant microfossil groups in marine sediments, artificial classifications typically are erected on
the basis ofcommon morphological features (Saito and
others, 1981). Whereas the morphological features may
appear to be unifying or differentiating, in actuality
they result from a combination of genotypic and environmental influences. Thus, morphology alone is
sometimes a poor approximation of true genetic associations and taxonomic affinities.
Planktonic foraminifera are greatly influenced by environmental changes in the upper oceanic waters, as
demonstrated by their wide rage of morphologic variability (see Kennett, 1976, for a review). Other independent techniques are therefore needed to clarify taxonomic problems and evaluate subtle differences among
species of planktonic foraminifera. Ideally, a combination of both ecophenotypic and genotypic variation
should be considered in order to construct natural classification schemes and decipher evolutionary trends in
the fossil record.
A major problem in achieving this goal is the lack
of basic genetic information needed to evaluate genotypic features in fossil foraminifera. In living organisms a genetic signature can be isolated from morphologic variation through the use of molecular
information, such as that found in DNA or proteins.
Proteins in general reflect the genetic make-up of an

METHODS
PROTEINBIOGEOCHEMISTRY
To determine species specificity of shell matrix proteins, monospecific samples of Recent planktonic foraminifera were analyzed from the top 2 cm of three
low-latitude cores from the western Pacific, Atlantic,

Department of Geology, University of South Florida, Tampa,
FL 33620.
2 Marine Science Program and Department of Geological Sciences,
University of South Carolina, Columbia, SC 29208.
I

159

and Bandy) and Globorotalia tumida (Brady).000-8. The two dimensional outline of each specimen was captured via video digitizer. 1981). Here. Details of the biochemical methods are presented elsewhere (Robbins. 1970). 1984. 1987). 198 1) (ECEQ) to determine the morphotypic composition of the assemblages of each species. Berger and others. Twenty-four harmonics or shape components were computed for each digitized specimen. Age estimates for the coretop samples range from 2. The extraction procedures used in the isotopic analysis are discussed in detail in Williams and others (1 988b).000 yr. unpublished data). Specimens were mounted on standard micropaleontology slides in a predetermined standard orientation. White and pink G. Approximately 0.159 3. and (4) Orbulina universa (d'orbigny).354 1. 3. Le. 1978. and the amino acids were then monitored by UV absorption at 254 nm after phenylisothiocyanate (PITC) precolumn derivatization (Robbins. 1990. These specimens were crushed. 1990). Four planktonic foraminiferal genera were studied: (1) Globigerinoides ruber (&Orbigny) and Globigerinoides sacculifer (Brady). Jones. and vacuum desiccated.3-0.190 Caribbean Eastern Pacific *P6304-4-4 P6702.1-2 P6702-2-4 Tongue of the Ocean. ruber were isotopically analyzed separately.888 571 2.. 1987).692 2.643 2. The morphotypes were removed from the samples based on the ECEQ determined morphotypes. as discussed in Healy-Williams and Williams (1 98 1). The retained protein material was freeze dried and separated on the basis of hydrophobicity (charge) by reverse phase high performance liquid chromatography (RPHPLC) (Robbins. Droxler. The isotopic results presented here are relative to PDB and have a precision of O.Robbins and Donachy.6 17 QUANTITATIVE MORPHOLQGY 4.000 daltons. The outlines of all specimens were digitized in a ventral orientation. (2) Pulleniatina obliquiloculata (Parker and Jones). (3) Globorotalia menardii (Parker.5 grams of individual species (several thousand monospecific specimens) were isolated from the >250 pm fraction with a vacuum picker.049 * Refers to cores which were used in the biochemical part of this study.136 3.200 specimens of each species were picked from the top 2 cm of 16 widely spaced tropical cores (Table 1). Robbins and Brew. ruber was picked from the sediment samples of Box 1. core x Location Western Pacific *ERDC Box 92 ERDC Box I12 Atlantic *P7008. BP based on sediment accumulation rates determined by biostratigraphy and oxygen isotope stratigraphy (Emiliani. The supernatent was exhaustively dialyzed against distilled water in a membrane with an upper molecular weight limit of 6. Reproducibility of area response of the chromatographs averages 1. ultrasonically cleaned in distilled water.000 to 4.168 the eluent was monitored at 220 nm for proteins and peptides. STABLEISOTOPIC ANALYSIS An average of five specimens of each morphotype of G. specimens were removed according to their harmonic 2 amplitude. Individual protein peaks collected from this procedure were first hydrolyzed. The absorbance of To quantify the morphological variability of the foraminifera. All specimens were selected from the 150-250 pm size fraction to avoid ontogenetic biases.4 V22. 198l). and ERDC 92 (Table l).10-2. and Caribbean (Table 1). a measure of test elongation (Healy-Williams and Williams. 1972. RESULTS AND DISCUSSION PROTEINBIOGEOCHEMISTRY The important first step in developing a procedure to ascertain a genetic signature from test material demonstrated that high molecular weight proteins and polypeptides could be isolated and partially characterized from the fossil organic shell matrices of single species of planktonic foraminifera (Robbins. Robbins and Brew. 1990). We used EXTENDED CABFAC and EXTENDED QMODEL vector analysis (Full and others. we will only present the morphologic results for the morphotypes of the species Globigerinoides ruber. P6304-4.160 ROBBINS AND HEALY-WILLIAMS TABLE1.000 1. 1987. and the outline coordinates were subjected to closed form Fourier series analysis (Ehrlich and Weinberg. All cores were used for the image analysis part of this study. Dried test material was decalcified in cold 50% formic acid and centrifuged. 1987.598 (m) 2. Robbins. . The monospecific specimens were picked from the 150250 pm fraction to minimize isotope effects due to size (Curry and Matthews.5%. Bahamas BX 1 BX8 Cariaco Trench P6603-3-4 Indian Ocean V 19-200 V 19-20 1 VI 9-204 Sulu Sea Circe 18PG RC14-78 RCI 2-357 Latitude/ longtude Depth 2"13'S 156'59'E 1'37's 159"14'E 12"59'N 44"20'W 692" 2 I"16'W 15"27'N 70'4 3'W 590" 103"oO'W 2'29" 103"oO'W 25"09'N 77"49'W 2592" 76"54'W 10'48" 64"47'W 4'1 3's 41"33'E 5"20'S 40'26'E lo"14'S 43'49'E 7"65'N 1 19"oO'E 8'00" 1 18"oO'E 9"oO'N 12095'E 1. Locations of cores sampled in this study. approximately 1.lo/Oo. and analyzed for stable oxygen and carbon isotopic composition with a VG SIRA 24 isotope ratio mass spectrometer.

and alanine (Robbins. These structural proteins may also be less prone to degradation than some other types of proteins because of their composition and fibrous structure (Robbins. 1987). One foraminiferal protein fraction. aspartic acid asparagine (Asx). serine. The results show that. Ala = alanine. This protein is dominated by the amino acids. The compositions of five major amino acids of polypeptide FP8 from the six species of planktonic foraminifera are shown in Figure 1. although the nonspinose group has slightly more glycine ( 1 7. The abbreviations used for amino acids are as follows: Asx = aspartic acid and asparagine. 1987). but do not distinguish spinose from non-spinose species as in the case of Asx. as expected. Ser = serine. Pulleniatina obliquiloculata. No significant interspecific or intergeneric differences were observed in the amino acids serine and alanine (Fig. >93 the proteins are not related. the species we analyzed are related to each other in varying degrees (Table 2). 1987. glycine (Gly). The RPHPLC separation of proteins shows that proteins from the six species of core-top planktonic foraminifera have comparable chromatographic profiles. In general. Robbins and Brew.5% f 1. and Globorotalia menardii.obliquiloculata G. l). sacculifer (spinose) and P. We applied the statistical method of Cornish-Bowden ( 1 983) using 13amino acids to determine if specific differences exist in the FP8 and to quantify the degree of relatedness of this protein.5% f 0.glutamic acid + glutamine (Glx). The percentages of Glx in G.161 BIOCHEMICAL. Globorotalia tumida. serine (Ser). expressed as percent of total. confirms that the protein composition is indeed similar from species to species. was found to be common to all species examined (Rob- bins. I). This similarity indicates that the shell organic matrix from different species is composed of many of the same proteins. GEOCHEMICAL. this technique can also quantify the degree of relatedness within a category.tumida G.6% f 1. sacculifer 2o P. Histograms of relative abundances of 5 of the 17 amino acids. 1984)and mollusks (Wheeler and others. Globigerinoides sacculife. the percentage of Asx in spinose species (10. and alanine (Ala). The amino acid composition of equivalent chromatographic peaks.3% f 1. Although there are distinct numerical cut-offs for classes of relationships (<42 proteins are strongly related. Glx = glutamic acid and glutamine. These results suggest that structural proteins are responsible for the morphological differences between species and perhaps determine the morphotype of species. 1987).9).universa AMINO ACID COMPOSITION OF FP8 G. AND MORPHOLOGICAL CRITERIA IN FORAMINIFERA 0 G. obliquiloculata (non-spinose) are significantly higher than in the other four species (Fig. suggesting that some of the proteins are homologous. although some amino acid compositional differences were also demonstrated (Robbins. ruber ( p ) 0 0. + . menardii r Asx Glx Ser Amino Acids GlY Ala FIGURE1 . etc.2) is always higher than non-spinose species (4.O) (Fig. thought to represent single proteins.4) than the spinose group (14.). The amino acid composition of some of the more hydrophobic proteins resembles fibrous structural proteins. On the average. which are similar to other structural proteins of Recent benthic foraminifera (Weiner and Erez. found in the FP8 protein from six species of planktonic foraminifera: Globigerinoides ruber. 1987). having significant quantities of glycine. I). Orbulina universa. 1990). designated FP8. Gly = glycine.

which eliminates free amino acids. Also.3 54. rnenordii P . STABLE ISOTOPESAND BIOCHEMISTRY Both environmental and genetic influences are inherent in the phenotypic expression of foraminifera. tumidu Gt.7 57. For example.1 36. is actually an average of the heterogeneous collection of proteins within the shell because the organic matrix was found .9 56. However. ~~ ~~ FIGURE2. 1990).9 0 0 47. significant amounts of free amino acids (especially acidic ones. which has been used for taxonomic distinctions (King and Hare. menardii (36.5% (CornishBowden. More specific phylogenetic conclusions from the molecular data require sequencing of the protein or may be obtained through other molecular or immunological approaches (Robbins and others.4 69. G. sacculifer (3 1. universa results indicated anomalously high methionine which could have affected the outcome of the Cornish-Bowden statistics. I 69. RUBER USING MORPHOLOGY.3 48. Robbins and Brew.4 0 44. such as aspartic acid) and small humic acids with hydrolyzable amino acids are adsorbed onto the test as well (Jackson and Bishoff. individual proteins in fossil foraminifera may provide more phylogenetic information (King and Hare.000 daltons. 1989). the relatedness of 0. and that which re- . Therefore. A cluster analysis using all of the amino acids in FP8 demonstrated a phylogenetic relationship similar to that presented by Kennett and Srinivasan (1983) (Fig.3 47.4 49.3 50. universa and G.6 8. obliquiloculnta Nhpr 0 19. 1971.0 G. 1972.8 31.5 44. universa and G.3 49. G. s'l<mlf<r 0. however.3 49.I62 ROBBINS AND HEALY-WILLIAMS TABLE 2. since the structural proteins.7 36. I x < 42 proteins strongly related 93 > x > 42 weaker indication of relatedness x > 93 proteins not related a biochemical basis exists for the distinction of genera based on spinose vs. Dendrogram of results of hierarchical cluster analysis performed on the amino acid composition of FF'8 from six species of planktonic foraminifera. Haugen and others. 1972. King.1 0 27. 1990). such as FP8. since the taxonomically useful aspartic acid and glutamic acid (and possible other important amino acid) signals are retained. Our approach of analyzing the amino acid composition of specific proteins uses a combination of dialysis. 1978. Table 2) suggests it has isologous variants that exhibit speciesspecific differences and that can provide additional phylogenetic information to the micropaleontologist. 1989).4 27. The only exception to this is the relatedness of Orbulina universa (spinose) to Globorotalia menardii (non-spinose).un.0 48.7 8. Haugen and others. In fact. 1) are not likely dissolution artifacts. DISTINGUISHING MORPHOTYPES OF G. For instance. abliquiloculoro 19. Therefore.7 49. and analysis of high molecular weight protein components. The interspecific compositional differences shown here for FP8 (such as differences in Asx) (Fig. 1987.ueno GI. 1987) than that previously obtained using bulk amino acid analyses. Relatedness of FP8 among 6 species of planktonic foraminifera based on statistical method of Cornish-Bowden (1983) as calculated using mole percent of 13 amino acids and a conservative estimate of molecular weight of 10. Such techniques virtually eliminate the possibility of analyzing contaminating free amino acids. the results presented here represent a more accurate analysis of the amino acid composition of a single protein. Bulk analysis. The nominal confidence level of this "strong" test is approximately 92. 1980. 1970). the compositions are unlikely to be influenced by other environmental factors such as food source differences or even secondary calcification. 1980). Robbins. ruber G. 1983) and could indicate a statistical or an inherent analytical error.3 56. to contain up to seven proteins (Robbins. 1972. Species-specific aspartic acid percentages from bulk amino acid analyses of spinose and non-spinose planktonic foraminifera and of benthic foraminifera were previously described (King and Hare. non-spinose morphology and species distinction based on overall amino acid composition of FP8. 1990). sacculifer 0. 1972. Carter and Mitterer. menmdii P.5 49. the 0.8 57.. on a relative scale.7).universa Gt.7 54. Robbins and Brew. However. Mitterer. 2). the tests of spinose species are more soluble to bottom waters that non-spinose species (Berger. These free amino acids could lead to false taxonomic classification. there is considerable difficulty in distinguishing between intraspecific morphologic variation that is purely ecophenotypic. This is particularly a cause for concern with aspartic acid.4) is not as strong as that between 0. are genetically predetermined. mwda GI.6 0 31. As a consequence. the amino acid compositional variability that FP8 displays from species to species (Fig. 1. yet the spinose species have higher percentages of this relatively unstable amino acid (Hoering.

environmental differences. Morphotypes were distinguished on the basis of chamber elongation (harmonic 2) as determined by EXTENDED CABFAC and EXTENDED QMODEL vector analysis. IOW < Elongation high A.75 0 G ruber-'normal' Ipl 0 G ruber-G elongatus G menardii (wl i .-2. 1987). Histograms of Tongue of the Ocean VOTO) sample shows that it is dominated by EM 1. A and B.PACIFIC G. Increasing elongation between the ultimate and penultimate chambers is shown from left to right. we undertook a detailed morphological. Also. 1974. and show wide variation in values between the groups. 1977). Hecht. the suggestion that these morphotypes are phenotypic variants (Kennett and Srinivasan. was examined via closed-form Fourier in a previous study of the 16 core tops listed in Table 1 (Robbins. Globorolnlia menardii. Histograms showing relative percentages of 5 of the total 17 amino acids comprising FPS protein from a morphotype representative of isotopic groups A and B. 21 16 20. TOTO G.0 -1. The morphologic variability of 1. 1974). B. Graph of carbon versus oxygen isotope values for six morphotypes from TOTO Box 1 shown in Figure 3A.2. 1826). stable isotopic. andisotopic separation.25 . (Parker. RUBER B. ruber that are 150-250 Fm in diameter. pyrumidulis (van den Broeck. cyclostomus (Galloway and Wissler.75 . W. 2. 1927).75 2 Del GI3 2. G. two groups. G. 1972. B. and 6 individuals. The abbreviations used for amino acids are as in Figure 1 and Thr = threonine and Met = methionine. ruber have been recognized as genetically distinct species. calor differences (p = pink versus w = white).75 -1.25 2. GEOCHEMICAL. a measure of shell elongation. Although a number of the morphotypes of G. Hecht and Savin. C. and biochemical study of the planktonic foraminiferal species Globigerinoides ruber in order to determine the genetic relatedness of the morphologic variants of this species. 1983) is at least partially supported by ecological studies (B6. Z 0 Y u P !-: I 20 I 1. MORPHOTYPES .25 1. included for comparison are compositional data from a morphologically unrelated species. West Pacific ERDC Box 92 shows high proportion of EM I and EM 6 individuals. ruber as quantified fmm 16 core tops from the tropics. and biochemical data for morphotypes of G. elongatus from group A were chosen on the hasis of shape difference (harmonic 2). Globigerinoides ruber exhibits considerable morphological variation.5 2. 1984) of the harmonic amplitude spectra indicated that harmonic 2.5 c. and G. RUEER MORPHOTYPES eo E ' 0 ~ 10' M 7 -2.200 specimens of G. The six morphotypes of G. A maximum entropy analysis (Full and others. Histograms of proportions of Globigerinoides ruber end members in a sample. sults from genetic differentiation either in concert with. elongatus (d'orbigny.I S? "10. contained the maximum amount of morphologic variability. 1876). Based on the results ofthe protein analysis. A. and it remains unresolved if the morphotypes represent separate species or environmental variants Amnlitude Amplitude FIGURE4. isotopic. or caused by.163 BIOCHEMICAL. AND MORPHOLOGICAL CRITERIA IN FORAMINIFERA A. An EXTENDED CABFAC EXTENDED QMODEL vector analysis of the frequency distributions of harmonic 2 indicated that six morphotypes account for more than . Sampleswerepicked from Atlantic core top P7008. Emiliani.5 "I 1. are delineated. As shown. Diagrams showing morphological. 12 8 4 0 As% Glx Met Thr Val Amino Acids FIGURE 3. ruber: A. Globigerinoides ruber "normal" from group B and C. 1962. for example.

in order of increasing elongation (harmonic 2).57~)is the typical 6I8O difference found between glacial-interglacial samples (Lynts and Judd. which significantly reduces the effect of vertical mixing. called end members (EM 1 through EM 6). Erez (1 979) determined an oxygen isotopic difference which averaged 0. 6I3C is not a function of temperature but rather a function of how each species incorporates the bicarbonate ion during calcification. 1981) and morphology (Spero and others. The oxygen isotopic difference found between extremes for the two groups (1. From the six identified morphotypes (EM 1 4 ) . represent increasing degrees of elongation that are schematically depicted in Figure 3A and Plate 1. 197 l). ruber. 1987). We therefore suggest that the 6I8O differences between morphotypes groups A and B are neither due to bioturbation nor purely ecological factors. These six morphotypes. The estimated oxygen isotopic value for equilibrium calcite at the TOTO site is . Erez. Average sedimentation rate for Box 1 is approximately 7-1 5 cm/yr3 (Droxler. A seasonality effect does not eliminate the possible genetic component to the differentiation of the morphotypes of G. Specimens of G. 1978).5700between G. 2. two general groups of end member associations can be distinguished on the basis of their oxygen and carbon isotopic composition. ruber as related to the isotopic and . 1984). 95% of the variance in the data array. To examine possible biochemicalhiological differentiation in G. the western Pacific is dominated by a bimodal distribution of end members 6 and 1. Caribbean. and other Atlantic cores (Robbins. Unlike the 6I8O of foraminifera. Scale bar = 50 pm. from left to right. 4 7 ~and . Globigerinoides ruber is known to have a seasonal component in both its oxygen isotopic composition (Williams and others. Similar isotopic/morphologic trends for this species were seen in the western Pacific. Not all end members are present in all samples examined. Such 6I3Cdifferences as those seen in G.1. 1979. 2 5 7 ~respectively. Group A contains the less elongate morphotypes EM 1-3. TOTO Bahamas.1 . 1977. 1971). Williams and others. Tongue of the Ocean. ruber. 3B).164 ROBBINS AND HEALY-WILLIAMS PLATE I Morphotypes of Globigerinoides ruber based on the results of closed form Fourier series analysis. It is unlikely that the range of values observed in the G. ruber morphotypes from Box 1. ruber morphotypes is due to bioturbation since the Tongue of the Ocean is a region of relatively high sedimentation rate (Lynts and Judd. including genetic variation (Weiner. ruber and G. elongatus from plankton tows. Bahamas (TOTO) was found to be dominated by end members 1.47~. which on the average are depleted in 6I8O and 6I3C by approximately .1 . but has no specimens typical of end member 4 (Fig. and 6 (Fig. For example. ruber morphotypes have often been attributed to the catchall term “vital effects” that may actually reflect a number of phenomena. were subsequently analyzed for their stable isotopic compositions (Fig. We interpret this to indicate geographic variability in the dispersal of these forms. EM 1 through EM 6. 4A). 4B). 1987). These morphotypes might indicate that these organisms are utilizing particular niches.close to the value of EM 5 G. compared to the more elongate morphotypes in Group B (EM 4-6). 1975. The range in isotopic values could be the result of seasonal temperature changes at the site.

a close approximation of the genotype. As a first step. During speciation. Figure 3C shows a partial amino acid composition of the individual FP8 proteins in the two G. These partial sequences may still allow phylogenetic relationships to be explored because of unique domains in the protein structure. 1976. Another obstacle is that sequencing the shell matrix pro- 165 teins is not a straightforward procedure: at least one of the proteins is blocked to Edman degradation (Robbins and Donachy. hundreds of thousands of specimens (approximately 5-1 0 grams) are needed. we determined the amino acid compositions of the protein FP8 from one morphotype in each of the two isotopic groups (EM 1 from Group A and EM 5 from Group B) (Fig. which contained these two morphotypes in abundance and had similar isotopic/morphologic trends. However. Until more is known about foraminiferal DNA (the genotype). In order to fully sequence the FP8 protein from the two morphotypes of G. While more data are needed to investigate the biochemical relationships between EM1 and EM5. One problem that presents itself is the amount of material needed to sequence proteins. ruber morphotypes. A small number of mutations (amino acids that have replaced each other) may be indicative of less genetic divergence of the proteins (and therefore of the species) as compared to larger differences seen in FP8 between species and genera. In addition. are some of the more conserved amino acids. For this analysis we used Atlantic core P7008. ruber morphotypes. Also notable is the fact that the amino acids. Furthmayr and Timple. 199 1). one can look toward technological improvements to eventually eliminate the problem of large sample size. which will provide information concerning degree of relatedness and give an indication of the time of branching events in the phylogenetic history of the lineage. The addition of a proxy genetic component . Further evaluation of the small compositional differences between morphotypes is currently in progress. The integration ofa biochemical signature with morphologic and isotopic data may aid in the refinement of taxonomic lineages. 1980). there may be more efficient ways to determine a genetic signal from living foraminifera through the analysis and sequence of DNA or through the use of monoclonal antibodies (Robbins and others. and insertions of amino acids that have occurred within the polypeptide chain. LIMITATIONS OF THE PRESENT BIOCHEMICAL RESULTS In order to fully understand the relationship between the planktonic foraminiferal morphotypes and differences in protein amino acid composition. 1991). These preliminary data show that only a few differences in the amino acid composition of FP8 exist between the two morphotypes of G. These two forms were chosen because they are morphologically distinct due to their degree of elongation (Plate 1. While obtaining large amounts of sediment may not be an issue. 3C). the differences in amino acid composition of the FP8 protein found in the two morphotypes are consistent with our interpretation of the morphologic and stable isotopic results indicating that there are two distinct groups of G. In a separate line of investigation. aspartic acid (Creighton. we show here that the protein FP8. Sequencedata will allow comparison of the amino acid sequence from one species to another. ruber examined in this study. shows compositional differences reflecting species differences. Lowenstein. In structural proteins of taxa from sponges to humans. new protein structures gradually emerge from old ones as a direct consequence of changes in genetic material along different lines of descent (Doolittle. and isotopic evidence might be created much in the same way that micropaleontologists now construct them based solely on morphology. 1979). morphologic. 3A). ruber. evolutionarily conservative proteins can be recognized by comparing the diverged sequences and the less conservative proteins can be used by taxonomists. In the next few years it can be anticipated that technological limitations will be reduced. will allow us to compare proteins within and between species and explore evolutionary histories of species. it appears likely that partial sequences can be obtained (Robbins and Donachy. 1970. ruber. Since slight modifications of morphology may be due to either environmental or to genetic influences. ruber morphotypes in terms of genetic rather than environmental causes. small differences in amino acid composition are unique and can be used taxonomically (Balazs. sequence data are required. Successful sequencing of the proteins will enable us to determine the substitutions. Deuser and Ross (1989) have also suggested a non-environmental distinction between the two varieties. and geochemical results support the distinction of G. morphological. Phylogenetic trees based on a combination of molecular. ruber and that a portion of these differences may be the result of genetic variation between morphotypes rather than ecologic variation. GEOCHEMICAL. If one compares the vast numbers of specimens needed by Emiliani (1 954) for his original stable isotope work to the few specimens required by present day mass spectrometers. Fietzek and Kuhn. morphologicdata alone are sometimes not sufficient to make taxonomic distinctions in micropaleontology. Fig. The combined biochemical. We have further shown that there are stable isotopic differences between the morphotypes of G. the limiting factor is the time involved in the picking of a monospecific sample. thereby improving biostratigraphy. They are relatively immutable during evolution as compared to the highly mutable amino acid. present in different species of core top planktonic foraminifera. 1991) and therefore limits the amount of protein that can actually be sequenced. IMPLICATIONSOF THISSTUDY The goal of this preliminary research is to provide the foundation from which we can eventually infer evolutionary relationships. In addition. AND MORPHOLOGICAL CRITERIA IN FORAMINIFERA morphological differences of groups A and B. which show variation between G. 1976. 1983). protein data. deletions.BIOCHEMICAL.

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