![]() By requiring only venom to obtain venom protein cDNAs, the approach detailed here will provide access to cDNA-based protein sequences from commercial and other venom sources, facilitating study of snake venom protein composition and evolution. Venom protein transcripts offer important insights into potential snakebite envenomation profiles and the molecular evolution of venom protein multigene families. The application of this technique will help to provide venom protein sequences for many snake species, including understudied rear-fanged snakes. These sequences aid in the identification and characterization of venom proteome profiles, allowing for the identification of peptide sequences, specific isoforms, and novel venom proteins. Transcripts for all major venom protein families (metalloproteinases, serine proteases, C-type lectins, phospholipases A 2 and three-finger toxins) responsible for clinically significant snakebite symptoms were obtained from venoms. Full-length transcripts were obtained from venom samples that were fresh, newly lyophilized, old, field desiccated or commercially prepared, representing a significant advance over previous attempts which produced only partial sequence transcripts. This work demonstrates that full-length venom protein messenger RNAs are present in secreted venoms and can be used to acquire full-length protein sequences of toxins from both front-fanged (Elapidae, Viperidae) and rear-fanged (Colubridae) snake venoms, eliminating the need to use venom glands. This approach, requiring only venom, provides access to cDNA sequences in the absence of living specimens, even from commercial venom sources, to evaluate important regional differences in venom composition and to study snake venom protein evolution. Venom-derived sequences can also be used to aid in characterizing venoms that lack proteomic profiles and identify sequence characteristics indicating specific envenomation profiles. These cDNA sequences from venom were then used to explore potential activities derived from protein sequence similarities and evolutionary histories within these large multigene superfamilies. Phospholipase A 2 sequences were also recovered from the venoms of several rattlesnakes and an elapid snake ( Pseudechis porphyriacus), and three-finger toxin sequences were recovered from multiple rear-fanged snake species, demonstrating that the three major clades of advanced snakes (Elapidae, Viperidae, Colubridae) have stable mRNA present in their venoms. This technique was used to assemble a partial venom gland transcriptome for the Middle American Rattlesnake ( Crotalus simus tzabcan) by amplifying sequences for phospholipases A 2, serine proteases, C-lectins, and metalloproteinases from within venom. These unknown venom protein cDNAs are obtained by the use of primers designed from conserved signal peptide sequences within each venom protein superfamily. This study provides evidence that unknown full-length venom protein transcripts can be obtained from the venoms of multiple species from all major venomous snake families. ![]() However, toxin transcripts are also present in venoms, offering the possibility of obtaining cDNA sequences directly from venom. To compare venom complexity and protein sequences, venom gland transcriptomes are assembled, which usually requires the sacrifice of snakes for tissue. Venomous snakes and their venoms also provide models for the study of molecular evolutionary processes leading to adaptation and genotype-phenotype relationships. Envenomation of humans by snakes is a complex and continuously evolving medical emergency, and treatment is made that much more difficult by the diverse biochemical composition of many venoms. ![]()
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