Translator Disclaimer
1 March 2004 Phylogenetic Placement of the Spider Genus Nephila (Araneae: Araneoidea) Inferred from rRNA and MaSp1 Gene Sequences
Author Affiliations +
Abstract

The family status of the genus Nephila, which belongs to Tetragnathidae currently but Araneidae formerly, was reexamined based on molecular phylogenetic analyses. In the present study, 12S and 18S rRNA gene fragments of eight species of spiders were amplified and sequenced. In addition, 3′-end partial cDNA of major ampullate spidroin-1 (MaSp1) gene of Argiope amoena was cloned and sequenced, and the 3′-end non-repetitive region's cDNA sequence of MaSp1 gene and the predicted amino acid sequence of C-terminal non-repetitive region of MaSp1 were aligned with some previously known sequences. The resulting phylogeny showed that Araneidae and Tetragnathidae are not a sister group in the superfamily Araneoidea, and the genus Nephila is closer to the genera of the family Araneidae rather than to those of Tetragnathidae. We suggest that the genus Nephila should be transferred back to Araneidae. Or the subfamily Nephilinae might be elevated to family level after it was redefined and redelimited. Furthermore, the study showed that 3′-end non-repetitive region's cDNA sequence of MaSp1 gene and C-terminal non-repetitive region's amino acid sequence of MaSp1 are useful molecular markers for phylogenetic analysis of spiders.

INTRODUCTION

The phylogenetic placement of the genus Nephila Leach has been changed several times together with the subfamily Nephilinae since it was established in 1815. Generally, the following placement of the genus has been appeared in different classifications. The Nephilinae was placed in the family Argiopidae along with the subfamilies Argiopinae, Araneinae, Tetragnathinae etc. (Simon, 1894; Pocock, 1900; Petrunkevitch, 1928), or in the family Araneidae along with the subfamilies Argiopinae, Araneinae etc., excluding Tetragnathinae (Roewer, 1942; Brignoli, l983). In the later case Tetragnathinae was elevated to family level as Tetragnathidae. In 1980, Levi placed the genus Nephila in the subfamily Metinae and placed Metinae, Araneinae and Tetragnathinae in the family Araneidae. Meanwhile, in the paper he also mentioned the genus might belong to a separate subgroup or subfamily (Levi, 1980). Later, Levi & Eickstedt (1989) pointed out that the genus Nephila have certain apomorphic tetragnathid characters in their palpal structures, and suggested that the placement of Nephila under Araneidae was incorrect. Subsequently, their suggestions were also supported by some morphological and behavioral studies (Coddington, 1990; Hormiga et al., 1995; Griswold et al., 1998). In most current papers Nephila has been treated as a genus of Nephilinae in the Tetragnathidae along with Tetragnathinae, and/or Metinae, Leucauginae etc. (Coddington and Levi, 1991; Platnick, 1989, 1997, 2003; Dippenaar-Schoeman and Jocqué, 1997).

To date, whether nephilines should be placed in Tetragnathidae or Araneidae is still a problem remains to be unsolved. The purpose of the present study was to test the two different placements of the genus Nephila by means of molecular phylogenetic analysis. In addition, the use of the 3′-end non-repetitive region's DNA sequence of MaSp1 gene and the C-terminal non-repetitive region's amino acid sequences of MaSp1 as molecular markers for phylogenetic analysis of spiders is also discussed.

MATERIALS AND METHODS

Samples

A total of nine species of spiders representing six families according to current classification were collected from China and India (Table 1). Samples were preserved in 99.9% ethanol.

Table 1.

Specimens used in this study1)

i0289-0003-21-3-343-t01.gif

DNA extraction, amplification and sequencing of 12S and 18S rRNA gene fragments

Total genomic DNA was extracted from leg tissue following standard proteinase digestion and phenol-chloroform extraction procedures as described by Sambrook et al. (1989). A fragment of the mitochondral 12S rRNA gene was amplified by using primers 12St-L: 5′-GGTGGCATTTTATTTTATTAGAGG-3′ (Croom et al., 1991) and 12Sbi-H: 5′-AAGAGCGACGGGCGATGTGT-3′ (Simon et al., 1994), and a fragment of the nuclear 18S rRNA gene was also amplified by using primers 18S-ai: 5′-CCTGAGAAACGGCTACCACATC-3′ and 18S-b0.5: 5′-GTTTCAGCTTTGCAACCAT-3′ (Tautz et al., 1988). The polymerase chain reaction (PCR) was performed on PE 2400 (Perkin Elmer) thermocycler system in 30 μl volumes containing 1–3 μl of genomic DNA, 10 pM of each primer, each of four deoxynucleoside triphosphates at 250 μM, 1.5 mM MgCl2, and 2 units of Taq Polymerase. PCR was carried out under the following conditions: hot-start at 95°C for 5 min; repeated 30 cycles; denaturation at 95°C for 40 s, primer annealing at 55°C for 50 s and primer extension at 72°C for 2 min; final extension at 72°C for 10 min. PCR products were separated on 2% agarose gel, and purified with DNA gel extraction kit (Vitagene). The purified DNA was sequenced using the BigDyeTM Terminator Cycle Sequencing Ready Reaction Kit on an ABI PRISM 310 Genetic Analyzer. An additional sequence of 12S rDNA was retrieved from GenBank (Table 1).

Cloning and sequencing of the 3′-end cDNA fragment of MaSp1 gene

Silk glands were dissected from Argiope amoena abdomens. The entire gland was removed and frozen immediately in liquid nitrogen. The gland was then crushed in a mortar and pestle under liquid nitrogen. All solutions, instruments and glassware were treated to inhibit RNAase activity. Total RNA was isolated by using Trizol®Reagent (Gibco), mRNA was isolated from total RNA with Oligotex™ mRNA Purification Kit (Qiagen). Single-strand cDNA was synthesized directly from mRNA using SMART™ cDNA Library Construction Kit (Clontech). Double-strand cDNA was synthesized by long-distance PCR using two adaptor primers offered by SMART™ cDNA Library Construction Kit (3′ PCR Primer: 5′-TTCTAGAGGCCGAGGCGGCC-3′; 5′ PCR Primer: 5′-AGCAGTGGTATCAACGCAGAGT-3).

Based on repetitive region's amino acid sequence (GGQG-GYGGL) of MaSp1 in spiders (Xu and Lewis, 1990), one primer (PMaSp1: 5′-GGAGGACAAGGTGGATATGGCGGATTAGG-3′) was designed and synthesized. Double-strand cDNA of silk gland in Argiope amoena was used as a template for 3′ RACE. The 3′ end cDNA fragment of MaSp1 gene was amplified by the primer pair 3′ PCR Primer and PMaSp1. A total of 30 μl PCR reaction mixture was composed of 3 μl of 1:100 dilution of double-strand cDNA of silk gland, 10 pM of each primer, each of four deoxynucleoside triphosphates at 250 μM, 1.5 mM MgCl2, and 2 units of Taq Polymerase. Amplification was carried out on PE2400 (Perkin Elmer) thermocycler system under the following conditions: hot-start at 95°C for 5 min; repeated 30 cycles: denaturation at 95°C for 40 s, primer annealing at 50°C for 40 s and primer extension at 72°C for 2 min; final extension at 72°C for 10 min.

PCR products were separated on 2% agarose gel, purified with DNA gel extraction kit (Vitagene), and ligated into PMD18-T vector using T/A Cloning Kit (TaKaRa). The cloned DNA was transformed and replicated in Escherichia coli (JM109), plasmid DNA was purified using Plasmid Mini Kit (Watson). After identified by PCR using M13 and M13 Reserve as primers, positive clones were sequenced on LI-COR DNA sequencer (LI-COR) also using M13 and M13 Reserve as primers. A further eight sequences were retrieved from GenBank (Table 1).

Data analysis

The predicted C-terminal non-repetitive region's amino acid sequences of MaSp1 were obtained from the 3′-end cDNA fragment of MaSp1 gene by using DNAclub. The DNA sequences of 12S and 18S rRNA gene fragments, MaSp1 gene fragments and predicted C-terminal non-repetitive region's amino acid sequences of MaSp1 were aligned respectively with CLUSTAL × 1.8 (Thompson et al., 1997). The neighbor-joining (NJ) method was applied to infer relationships among taxa on the basis of a pairwise matrix of the distance from Kimura's two-parameter model, using MEGA 2.1 (Kumar et al., 2000). The maximum-parsimony (MP) analyses were conducted using heuristic search option of PAUP 4.0b (Swofford, 2000). And the Maximum-likelihood (ML) analyses were performed by use of DnaSP 3.5 and PHYLIP vers. 3.5c computer packages (Felsenstein, 1993).

RESULTS

12S rDNA data set

The aligned sequences of 12S rDNA fragments for nine species of spiders consisted of a total of 269 sites, including 191 variable and 102 parsimony informative sites. The fragments have a high average A/T content of about 75.5%; this result is consistent with the statement that arthropod mitochondrial genomes in general tend to be highly A+T biased (Crozier and Croizer, 1993). The majority of transition values were lower than transversion ones. Sequence differences (transition + transversion) among these spiders ranged from 0.6% to 33.9%, with an average of 24.7%. It is noteworthy that average sequence difference (23.0%) between the genus Nephila and other tetragnathid genera was somewhat higher than that between Nephila and the araneid genera (21.4%) (Table 2).

Table 2.

Percent difference of base substitution (upper triangle) and values of transitions/transver-sions (lower triangle) for 12S rRNA gene fragment among 9 species of spiders

i0289-0003-21-3-343-t02.gif

NJ, MP and ML trees of the 12S rDNA fragments with Ornithoctonus huwena as outgroup indicate that the Araneidae, Nephila, Linyphiidae and Tetragnathidae clustered together, while the theridiid genus (Achaearanea) formed another single clade (Fig. 1). In NJ and ML trees, two species of Nephila clustered with the araneid clade, the linyphiid genus (Hylyphantes) and two tetragnathid genera (Tetragnatha and Leucauge) formed another monophyletic clade; but in the MP tree, the linyphiid genus clustered with the group which consists of the araneid genera (Gasteracantha, Argiope) and Nephila.

Fig. 1.

1Neighbor-joining (NJ), maximum parsimony (MP) and maximum-likelihood (ML) trees based on 12S rDNA sequence data of nine species of spiders. Bar equals 0.05 (NJ) and 10 (ML) units of Kimura's two-parameter distance. Numbers above branches in these trees are Bootstrap values at least 50% of the 1,000 bootstrap replications.

i0289-0003-21-3-343-f01.gif

18S rDNA data set

The aligned sequences of 18S rDNA fragments for eight species of spiders consisted of 738 total sites, including 190 variable and 58 parsimony informative sites. In contrast to the 12S rDNA fragments, the 18S fragments have a lower average A/T content about 49.6%, and the majority of transition values were higher than transversion ones (Table 3).

Table 3.

Percent difference of base substitution (upper triangle) and values of transitions/transversions (lower triangle) for 18S rRNA gene fragment among 8 species of spiders

i0289-0003-21-3-343-t03.gif

NJ, MP and ML trees of the 18S rDNA fragments with Heptathela hangzhouensis and Ornithoctonus huwena as outgroups generated very similar tree topologies one another (Fig. 2). The theridiid genus (Achaearanea) clustered with the group which consists of Nephila and the araneid genera (Gasteracantha, Argiope), while the tetragnathid genus (Tetragnatha) and the linyphiid genus (Hylyphantes) formed another monophyletic clade.

Fig. 2.

Neighbor-joining (NJ), maximum parsimony (MP) and maximum-likelihood (ML) trees based on 18S rDNA sequence data of eight species of spiders. Bar equals 0.02 (NJ) and 10 (ML) units of Kimura's two-parameter distance. Numbers above branches in these trees are Bootstrap values at least 50% of the 1,000 bootstrap replications.

i0289-0003-21-3-343-f02.gif

MaSp1 gene data set

The sequences of the cDNA fragment (GenBank Accession no.  AY263390) of MaSp1 gene can be divided into two regions: (1) a repetitive region that codes for an alternating alanine-rich and glycine-rich domain and (2) a non-repetitive coding region. The predicted C-terminal non-repetitive region of MaSp1 consisted of predicted 102 amino acids, its sequence was rather homologous to published amino acid sequences of other eight species of spiders, for example, the amino acid sequence of Argiope amoena had a homology of 72.4% to that of Nephila clavipes (Fig. 3).

Fig. 3.

Allignment of the C-terminal non-repetitive regions predicted Amino acid sequences of MaSp1 of Argiope amoena and other eight species of spiders. The mark (#) indicates where amino acid sequence has not revealed, the short line (–) indicates where an amino acid is deleted.

i0289-0003-21-3-343-f03.gif

The aligned 3′-end non-repetitive region's cDNA sequences of MaSp1 gene for nine species of spiders consisted of 310 total sites, including 215 variable and 149 parsimony informative sites. The MaSp1 gene fragments have a lower average A/T content about 54.6%, and the majority of transition values were higher than transversion ones. The average sequence difference (18.1%) between the genus Nephila and other tetragnathid genera was markedly higher than that between Nephila and the araneid genera (13.2%) (Table 4).

Table 4.

Percent difference of base substitution (upper triangle) and values of transitions/transversions (lower triangle) for 3′-end non-repetitive region's cDNA sequence of MaSp1 gene among 9 species of spiders

i0289-0003-21-3-343-t04.gif

NJ and MP trees based on the 3′-end non-repetitive region's cDNA sequences of MaSp1 gene or the predicted amino acid sequences of C-terminal non-repetitive region of MaSp1 with Dolomedes tenebrosus as an outgroup are also in agreement with the traditional view that genus Nephila should be placed in the family Araneidae rather than Tetragnathidae (Fig. 4). Three species of the genus Nephila is sister to other two araneid genera Araneus and Argiope. And then the clade is sister to genus Tetragnatha of the family Tetragnathidae.

Fig. 4.

The neighbour-joining (NJ) and Maximum parsimony (MP) trees resulting from analysis of the 3′-end non-repetitive regions cDNA sequences of MaSp1 gene and C-terminal non-repetitive regions amino acid sequences of MaSp1 of nine species of spiders. Bootstrap values at least 50% of the 1,000 bootstrap replications are shown above branches in these trees.

i0289-0003-21-3-343-f04.gif

DISCUSSION

The phylogenetic position of the genus Nephila

Although the genus Nephila possesses morphological characters that are believed to be synapomorphies of the family Tetragnathidae (Roewer, 1942; Brignoli, 1983; Song et al., 2001), it also has many characters that are not shared by other tetragnathid genera. These characters include the labium longer than wide, transverse grooves on the book lung covers, and the metatarsi and tarsi together longer than the patellae and tibiae (Levi and Eickstedt, 1989). On the other hand, it also bears many characters in common with the family Araneidae, such as a distinct epigynum, characteristic integral structure of male palpal organ and striking sexual size dimorphism (female giantism). Hence, it seems to be difficult to judge the phylogenetic position of the genus Nephila by means of morphological methods alone.

In the present study, the NJ, MP and ML trees based on the sequences of 12S rRNA or 18S rRNA gene fragments, and the NJ and MP trees based on the 3′-end non-repetitive region's cDNA sequences of MaSp1 gene or the C-terminal non-repetitive region's amino acid sequences of MaSp1, revealed that the phylogenetic position of the genus Nephila is closer to the araneid genera than to tetragnathid. It is notable that the NJ, MP and ML trees of the 12S and 18S rDNA fragments corroborated the statements that Araneidae and Tetragnathidae do not form a sister group in the superfamily Araneoidea (Coddington and Levi, 1991; Hausdorf, 1999). Although a linyphiid genus or a theriddiid genus included in the analyses intervened between Araneidae and Tetragnathidae in those reconstructed phylogenetic trees (Figs. 1 and 2), the genus Nephila was always connected directly with the araneid genera. Our results showed that Nephila should be transferred back to the family Araneidae, or the genus and its subfamily might be recognized as a separate clade in the superfamily Araneoidea and should be removed from either Araneidae or Tetragnathidae. In the later case, the subfamily Nephilinae might be elevated to family level after being redefined and redelimited.

The use of the C-terminal non-repetitive region of MaSp1 as a molecular marker

As the web frame and lifelines of spiders, dragline silk plays an important role in spider's life. Dragline silk is a two-protein fiber and, MaSp1 (major ampullate spidroin-1) and MaSp2 (major ampullate spidroin-2) genes which are active in the major ampullate gland are thought to be responsible for the dragline silk. MaSp1 gene encodes a kind of protein molecule that contains the repetitive region and C-terminal non-repetitive region (Xu and Lewis, 1990; Van Beek et al., 2002). The repetitive region exhibits a pattern of alternating alanine-rich, crystal-forming blocks that impart the silk's unmatched strength in the natural world and glycine-rich amorphous blocks implicated in providing elasticity in the silk filament. In contrast to the repetitive region, the C-terminal non-repetitive region is hydrophilic, thus it proved to be necessary to maintain the soluble or liquid crystalline state of silk molecules along the gland duct before being secreted from silk gland (Anthoula et al., 2002). The alignment of predicted amino acid sequences of C-terminal non-repetitive region of MaSp1 of nine species of spiders shows this region has considerably conserved amino acid sequences (Fig. 3).

The results from the NJ and MP trees based on the 3′-end non-repetitive region's cDNA sequences of MaSp1 gene or the C-terminal non-repetitive region's amino acid sequences (Fig. 4) agree with the phylogenetic relationships among these spiders in traditional classifications. They are also similar to the results based on sequences of 12S and 18S rRNA gene fragments (Figs. 1 and 2) on the phylogenetic placement of Nephila. At least, it indicates that both the 3′-end non-repetitive region's cDNA sequences of MaSp1 gene and the C-terminal non-repetitive region's amino acid sequence of MaSp1 have a certain value in phylogenetic analysis of spiders. In addition, the MaSp1 gene has no intron, and consists of a single exon alone (Beckwitt et al, 1998; Hayashi and Lewis, 2000). Therefore, amplification of the 3′-end partial DNA of MaSp1 gene directly from genomic DNA of spider is feasible. That is to say, not via complicated reverse transcription from mRNA, the 3′-end non-repetitive region's DNA sequences of MaSp1 gene and the C-terminal non-repetitive region's amino acid sequence of MaSp1 can be obtained conveniently and economically.

Acknowledgments

We thank Prof. An. Subramanian (Centre of Advanced Study in Marine Biology, Annamalai University, India) for his help in collecting some spiders of India. Thanks are also due to Dr. Zhongquan Liu (Department of Biology, Yancheng teacher's college, China) and Dr. Changfa Zhou (College of life Sciences, Nanjing Normal University, China) for their valuable discussions and technical assistance. This work was supported by a key project of the National Natural Science Foundation of China (No. 30130040).

REFERENCES

1.

L. Anthoula, S. Arcidiacono, Y. Huang, J. F. Zhou, F. Duguay, N. Chretien, E. A. Welsh, J. W. Soares, and C. N. Karatzas . 2002. Spider silk fibers spun from soluble recombinant silk produced in mammalian cells. Science 295:472–476. Google Scholar

2.

R. Beckwitt, S. Arcidiacono, and R. Stote . 1998. Evolution of repetitive proteins: spider silks from Nephila clavipes (Tetragnathidae) and Araneus bicentenarius (Araneidae). Insect Biochem Mol Biol 28:121–130. Google Scholar

3.

P. M. Brignoli 1983. A Catalogue of the Araneae Described between 1940 and 1981. Manchester University Press. Manchester. pp. 755. Google Scholar

4.

J. A. Coddington 1990. Ontogeny and homology in the male palpus of orb-weaving spiders and their relatives, with comments on phylogeny (Araneoclada: Araneoidea, Deinopoidea). Smithsonian Contribution to Zoology 496:1–52. Google Scholar

5.

J. A. Coddington and H. W. Levi . 1991. Systematics and evolution of spiders (Araneae). Annu Rev Ecol Syst 22:565–592. Google Scholar

6.

H. B. Croom, R. G. Gillesple, and S. R. Palumbi . 1991. Mitochondrial DNA sequences coding for a portion of the RNA of the small ribosomal subunits of Tetragnath mandibulata and Tetragnatha hawaiensis (Araneae, Tetragnathidae). J Arachnol 19:210–214. Google Scholar

7.

R. H. Crozier and Y. C. Crozier . 1993. The mitochondrial genome of the honyebee Apis mellifera: Complete sequence and genome organization. Genetics 133:97–117. Google Scholar

8.

A. S. Dippenaar-Schoeman and R. Jocqué . 1997. African Spiders, An Identification Manual. Isteg Scientific Publications. Irene. pp. 392. Google Scholar

9.

J. Felsenstein 1993. PHYLIP (phylogeny inference package) version 3.5c. Distributed by the author. Dept Genet Univ Washington. Seattle. Google Scholar

10.

C. E. Griswold, J. A. Coddington, G. Hormiga, and N. Scharff . 1998. Phylogeny of the orb-web building spiders (Araneae, Orbiculariae: Deinopoidea, Araneoidea). Zool J Linn Soc 123:1–99. Google Scholar

11.

B. Hausdorf 1999. Molecular phylogeny of araneomorph spiders. J Evol Biol 12:980–985. Google Scholar

12.

C. Y. Hayashi and R. V. Lewis . 2000. Molecular architecture and evolution of a modular spider silk protein gene. Science 287:1477–1479. Google Scholar

13.

G. Hormiga, W. G. Eberhard, and J. A. Coddington . 1995. Web-construction behaviour in Australian Phonognatha and the phylogeny of nephiline and tetragnathid spiders (Araneae: Tetragnathidae). Aust J Zool 43:313–364. Google Scholar

14.

S. Kumar, K. Tamura, I. Jakobsen, and M. Nei . 2000. MEGA2: molecular evolutionary genetic analysis. Arizona State University. Tempe. Google Scholar

15.

H. W. Levi 1980. The orb-weaver genus Mecynogea, the subfamily Metinae and the genera Pachygnatha, Glenognatha and Azilia of the subfamily Tetragnathinae north of Mexico (Araneae: Araneidae). Bull Mus Comp Zool 149:1–74. Google Scholar

16.

H. W. Levi and V. R. D. von Eickstedt . 1989. The Nephilinae spiders of the neotropics. Mems Inst Butantan 51:43–56. Google Scholar

17.

A. Petrunkevitch 1928. Systema Aranearum. Trans Connecticut Acad Sci, Yale University Press 29:1–270. Google Scholar

18.

N. I. Platnick 1989. Advances in Spider Taxonomy 1981–1987. Manchester University Press. Manchester. pp. 1–673. Google Scholar

19.

N. I. Platnick 1997. Advances in Spider Taxonomy 1992–1995. New York Entomological Society. New York. pp. 1–976. Google Scholar

20.

N. I. Platnick 2003. The world spider catalog, version 3.5. American Museum of Natural History online at  http://research.amnh.org/entomology/spiders/catalog81-87/index.htmlGoogle Scholar

21.

R. I. Pocock 1900. The Fauna of British India, Including Ceylon and Phylogenetic Placement of Genus Nephila 351 Burma. Arachnida. London. pp. 1–279. Google Scholar

22.

C. F. Roewer 1942. Katalog der Araneae 1758–1940. Kommissions-Verlag von “Natura”. Bremen. pp. 733–971. Google Scholar

23.

J. E. Sambrook, F. Fitch, and T. Maniatis . 1989. Molecular cloning, a laboratory manual. 2nd edition. Cold Spring Harbour Laboratory Press. Cold Spring Harbor, New York. Google Scholar

24.

E. Simon 1894. Histoire Naturelle des Araignées. Paris Libraire Encydo pédique de Roret, Tome 1, Fascicule 3. pp. 489–760. Google Scholar

25.

C. Simon, F. Frati, A. Bechenbach, B. Crespi, H. Liu, and P. Flook . 1994. Evolution, weighting and phylogenetic utility of mitochondrial gene sequences and a compilation of conserved polymerase chain reaction primers. Ann Entomol Soc Am 87:651–710. Google Scholar

26.

D. X. Song, M. S. Zhu, and J. Chen . 2001. The Spiders of China. Hebei Science and Technology Press. Sijiazhuan. pp. 1–32. Google Scholar

27.

D. L. Swofford 2000. PAUP: phylogenetic analysis using parsimony. Version 4. Sinaner Associates. Sunderland, Mass. Google Scholar

28.

D. Tautz, J. M. Hancock, D. A. Webb, C. Tautz, and G. A. Dover . 1988. Complete sequences of the rRNA genes of Drosophila melanogaster. Mol Biol Evol 5:366–376. Google Scholar

29.

J. D. Thompson, T. J. Gibson, and F. Plewniak . 1997. Clustal X windows interface: flexible strategies multiple sequence alignment aided by quality analysis tools. Nucl Acid Res 24:4876–4882. Google Scholar

30.

J. D. Van Beek, S. Hess, F. Vollrath, and B. H. Merier . 2002. The molecular structure of spider dragline silk: folding and orientation of the protein backbone. Proc Natl Acad Sci USA 99:10266–10271. Google Scholar

31.

M. Xu and R. V. Lewis . 1990. Structure of a protein superfiber: spider dragline silk. Proc Natl Acad Sci USA 87:7120–7124. Google Scholar
Pan Hong-Chun, Zhou Kai-Ya, Song Da-Xiang, and Qiu Yang "Phylogenetic Placement of the Spider Genus Nephila (Araneae: Araneoidea) Inferred from rRNA and MaSp1 Gene Sequences," Zoological Science 21(3), 343-351, (1 March 2004). https://doi.org/10.2108/zsj.21.343
Received: 12 May 2003; Accepted: 1 November 2003; Published: 1 March 2004
JOURNAL ARTICLE
9 PAGES


SHARE
ARTICLE IMPACT
Back to Top