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These unique peptides were found to be similar to the xyloglucan-specific fungal endoglucanase inhibitor protein (XEGIP) precursor in tomato (Lycopersicon esculentum) and its homolog in potato (Solanum tuberosum). A pair of oligonucleotide primers was designed based on the potato and tomato sequences that were used to clone a 1,018-bp internal piece of nec4 cDNA from a stage 6 nectary cDNA library. The remaining portions of the cDNA were subsequently captured by 5' and 3' rapid amplification of cDNA.
RNA was quantitated in duplicate using ultraviolet absorption spectrometry at 260 nm (Nanodrop Technologies, Wilmington, DE). For cDNA synthesis, 5 µg of total RNA was reverse transcribed with Superscript II reverse transcriptase (50 units, Invitrogen, Carlsbad, CA) using an oligo-(dT)12–18 primer as described . The single-stranded cDNA product was purified using column adsorption chromatography (Qiagen, Valencia, CA). A single cDNA preparation from each specimen was used for each assay. Alternatively, for 3'-rapid amplification of cDNA.