Identifying parasitism genes encoding proteins secreted from a nematode's esophageal gland cells and injected through its stylet into plant tissue is the key to understanding the molecular basis of nematode parasitism of plants. Meloidogyne incognita parasitism genes were cloned by microaspirating the cytoplasm from the esophageal gland cells of different parasitic stages to provide mRNA to create a gland cell-specific cDNA library by long-distance reverse-transcriptase polymerase chain reaction. Of 2,452 cDNA clones sequenced, deduced protein sequences of 185 cDNAs had a signal peptide for secretion and, thus, could have a role in root-knot nematode parasitism of plants. High-throughput in situ hybridization with cDNA clones encoding signal peptides resulted in probes of 37 unique clones specifically hybridizing to transcripts accumulating within the subventral (13 clones) or dorsal (24 clones) esophageal gland cells of M. incognita. In BLASTP analyses, 73% of the predicted proteins were novel proteins. Those with similarities to known proteins included a pectate lyase, acid phosphatase, and hypothetical proteins from other organisms. Our cell-specific analysis of genes encoding secretory proteins provided, for the first time, a profile of putative parasitism genes expressed in the M. incognita esophageal gland cells throughout the parasitic cycle.