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Mutant cDNA sequences were obtained by reverse transcription followed by PCR amplification with total RNAs and NPH1-specific primers. PCR products were purified with a QIAGEN PCR purification kit and sequenced directly. Both strands of the entire cDNA sequence for each mutant were examined for mutations, and each lesion was confirmed by two independent PCR experiments.
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Because nph1-2 is in the Estland ecotype, we also sequenced an Estland wild-type cDNA for comparison.
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An additional allele, nph1-3, is identical in sequence to nph1-1.
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Single-letter abbreviations for the amino acid residues are as follows: A, Ala; C, Cys; D, Asp; E, Glu; F, Phe; G, Gly; H, His; I, Ile; K, Lys; L, Leu; M, Met; N, Asn; P, Pro; Q, Gln; R, Arg; S, Ser; T, Thr; V, Val; W, Trp; and Y, Tyr.
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We thank K. Meyer and W. Lukowitz for providing the genomic library, J. R. Ecker for providing the cDNA and YAC libraries, C. Somerville for assistance with the AFLP technique, and P. Reymond for many helpful comments on the manuscript. This work was funded by NSF grants MCB-9219256 and IBN-9601164. This paper is Carnegie Institution of Washington. Department of Plant Biology publication 1367.
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