The internal deletion mutants of the NtMyb2 promoter were made as follows: upstream regions of the internal deletions were made by PCR with or without point mutations to make an AseI and pSK-LIP as a template. PCR products were digested with AseI, filled in with Klenow enzyme, and digested with XbaI. The downstream regions were made by PCR, which has a mutation to generate a BamHI site in the position corresponding to +80 of the NtMyb2 promoter, and a specific primer with a mutation to generate a DraI, followed by digestion with BamHI and DraI. An appropriate set of fragments was inserted between the XbaI and BamHI sites of pLTR-CAT4 (Hirochika et al., 1996), resulting in pLIP-CAT-2–pLIP-CAT-6. The full-length NtMyb2 promoter fragment (−694 to +80) and truncated fragments (−617 to +80 and −71 to +80) were amplified by PCR, digested with XbaI and BamHI, and inserted into pLTR-CAT4 XbaI and BamHI sites, resulting in pLIP-CAT-0, pLIP-CAT-1, and pLIP-CAT-7, respectively. The deletion start and end points of the resulting plasmids pLIP-CAT-1–pLIP-CAT-7 were −694 to −620, −617 to −500, −505 to −408, −414 to −315, −320 to −222, −218 to −72, and −694 to −72, respectively, from the transcription start site. The 169-bp region from −231 to −62 was amplified by PCR and cloned into the EcoRV site of pBlueScriptSK(+), resulting in pSK-LIP6. The HindIII–PstI fragment from pSK-LIP6 was inserted between the HindIII and PstI sites of p35S(40)-CAT (Takeda et al., 1999), resulting in pLIP(40)-CAT-6. The internal small (<24 bp) deletion mutants of the 169-bp region were made as follows: upstream fragments were made by PCR using universal M13 primer F (M13-F: 5′-CGTTGTAAAACGACGGCCAG-3′) and a specific primer whose 5′-terminal 20 nt matched the upstream side of the deletion start point and 3′-terminal 10 nt matched the downstream side of the deletion end point. The downstream fragments were also made by PCR using universal M13-RV primer and a specific primer whose 5′-terminal 20 nt matched the downstream side of the deletion end point and whose 3′-terminal 10 nt matched the upstream side of deletion start point. Next, appropriate PCR products were mixed (1/10 volume each) and amplified with the two universal primers (M13-F and M13-RV) to make fused fragments, which were then digested with HindIII and PstI and ligated with a HindIII and PstI digest of p35S(40)-CAT, resulting in pLIP(40)-CAT-62, -63, -64, -65, -66, -653, -654, -655, -656, -661, -662, and -663. The PCR products amplified using primers 61F: 5′-CGATAAGCTTGTGTAATACACACTTGTCGT-3′ and M13-RV, and M13-F and 67R: 5′-CGGGCTGCAGTGTTTGCGATTTCCACGAAG-3′ were digested with HindIII and PstI and inserted into p35S(40)-CAT, resulting in pLIP(40)-CAT-61 and -67, respectively. The deletion start and end points of the resulting plasmids pLIP-CAT-61–pLIP-CAT-67 were −221 to −206, −205 to −182, −181 to −158, −157 to −134, −133 to −110, −109 to −86, and −85 to −62, respectively, from the transcription start site. The point mutations were introduced by PCR as previously described by Sugimoto et al. (2000), resulting in pLIP(40)-CAT-651 and -652. The oligonucleotides 5′-agctaa(AAGATCCAA)8aactgca-3′ and 5′-gtt(TTGGATCCTT)8tta-3′, and 5′-agctaa(AAGCGCCAA)8aactgca-3′ and 5′-gtt(TTGGCGCCTT)8tta-3′ were annealed and ligated between the HindIII and PstI sites of p35S(40)-CAT, resulting in p35S(40)-AG8W-CAT and p35S(40)-AG8M-CAT. The sequences of all of the PCR-amplified fragments or inserted oligonucleotides were re-confirmed by sequencing.