Major ampullate silk, also known as dragline silk, is one of the strongest biomaterials known. This silk is composed of two proteins, major ampullate spidroin 1 (MaSp1) and major ampullate spidroin 2 (MaSp2). Only partial cDNA sequences have been obtained for these proteins, and these sequences are toward the C-terminus. Thus, the N-terminal domains have never been characterized for either protein. Here we report the sequence of the N-terminal region of major ampullate silk proteins from three spider species: Argiope trifasciata, Latrodectus geometricus, and Nephila inaurata madagascariensis. The amino acid sequences are inferred from genomic DNA clones. Northern blotting experiments suggest that the predicted 5' end of the transcripts are present in fibroin mRNA. The presence of more than one Met codon in the N-terminal region indicates the possibility of translation of both a long and a short isoform. The size of the short isoform is consistent with the published, cDNA based, N-terminal sequence found in flagelliform silk. Analyses comparing the level of identity of all known spider silk N-termini show that the N-terminus is the most conserved part of silk proteins. Two DNA sequence motifs identified upstream of the putative transcription start site are potential silk fibroin promoter elements.