Table 2.

Primers and PCR Conditions Used for the SMADIP1-Coding-Sequence Screening^[Note]

	Primer
Exon	Forward	Reverse	AnnealingTemperature (°C)	PCRProduct(bp)
2	GCGATGCCCACATTGTCG	TCTCGAGCCGCGTAGTG	53	229
3	ATCTCAACAGAGTGTTTAGAG	GAAGATGTGAAGATGGTAC	50	354
4	TGTGCTTGGCATGCTTAG	CATGACACAGGACAAATG	50	191
5	GAGATGGACTTGGGATCC	CAGAGGTCAGTGCAGTG	57a	297
6	GCTGCCATTGATTTACTG	GCCAATCAAAGCAATATCG	50	348
7	TTCCTCCTGCACAAAACTG	AAATGCAAATGCGAGCTCC	50	191
8:
A	ATCAAGGAAATGTAACGG	GTGTGTAGCCATAAGAAC	50	346
B	CTGAACAGACAGGCTTAC	TGACTCACTACCGGAAG	50	411
C	GAGGAACAAGGAGTTAC	CTCAGAAAGTACAGATGAC	50	434
D	CCAACAAAGCCGGAGTT	TGTTAGCCTGAGAGGAG	56b	421
E	CATCACCATCTATAGCAG	CTGAACACTGGGTTAGTG	50	438
F	TGAGCCTCTGAACTTGAC	GCCACCTCTTTTCTTCATAG	53	361
9	GTGAGCACAAAGAGAAGC	GAAATGTACAGCAGGACG	50	284
10:
A	CTAATCACCCTGTCATC	TAATGCTCTGCAAGTAAGC	53	317
B	GAAAGGGCACTTGGAAC	CAGCAGTGTTTTCAAGC	50	427

Note.— For PCR amplification, the reaction mixture (25 μl) contained 100 ng leukocyte DNA, 20 pmol each primer, 0.1 μM dNTP, 0.75 μl MgCl₂ 0.1 M, and 1 U Taq DNA polymerase (Gibco). Thirty cycles of amplification were performed, each cycle consisting of 30 s denaturation at 96°C, 30 s annealing, and 30 s elongation. PCR products were heated for 10 min at 96°C, loaded onto a GeneGel Excel 12.5/24 kit (Amersham Pharmacia Biotech), and electrophoresed for 2.5 h at 5°C and at 15°C.

^a+ 5% Dimethyl sulfoxide.

^b+ 2.5% Dimethyl sulfoxide.