Table 4.

Most successful primer combinations and central cycles of the PCR programmes.

DNA region	Primers	Central cycles
18S	Whole region: NS1/NS8 or NS1/SR6
	Two parts:
	1st: NS1/NS4	(i) 5 cycles of 45 s at 94 °C, 45 s at 53 °C (decreasing 1° C/cycle), 1.5 min at 72 °C
	2nd: NS21/SR6	(ii) 35 cycles of 45 s at 94 °C, 45 s at 48 °C, 1.5 min at 72 °C
ITS+28S	Both regions: ITS1F/LR5
	Two parts:
	ITS: ITS1F/ITS4	(i) 5 cycles of 45 s at 94 °C, 45 s at 55 °C (decreasing 1 °C/cycle), 1.5 min at 72 °C
	28S: LR0R/LR5	(ii) 35 cycles of 45 s at 94 °C, 45 s at 50 °C, 1.5 min at 72 °C
	Other smaller parts: ITS1F/ITS2, ITS3/ITS4, LR0R/LR3, and LR3R/LR5	Same as above, but extension reduced to 45 s at 72 °C
	Specific for Dacryonaema/Unilacryma:	40 cycles of 45 s at 94 °C, 45 s at 60 °C, 30 s at 72 °C
	ITS1: ITS1DacryoUnil-F/ITS1Dacryo-r or ITS1Unil-r
	ITS2: ITS2Dacryo-F or ITS2Unil-F/ITS2DacryoUnil-r
RPB1	gRPB1-A/fRPB1-C, or DacryRPB1-1F/DacryRPB1-2r, or DacryRPB1-A/DacryRPB1-C	(i) 5 cycles of 45 s at 94 °C, 45 s at 58 °C (decreasing 1 °C/cycle), 1.5 min at 72 °C
		(ii) 35 cycles of 45 s at 94 °C, 45 s at 53 °C, 1.5 min at 72 °C
RPB2	RPB2-6F or bRPB2-6F or fRPB2-7cF/fRPB2-11aR or bRPB2-10.9R (all six combinations), or DacryRPB2-6F/ DacryRPB2-11aR, or DacryRPB2-6.2F/DacryRPB2-11bR	(i) 5 cycles of 45 s at 94 °C, 45 s at 58 °C (decreasing 1 °C/cycle), 1.5 min at 72 °C
		(ii) 35 cycles of 45 s at 94 °C, 45 s at 53 °C, 1.5 min at 72 °C
TEF-1α	EF1-1018F or Efdf/EF1-1953R or EF1-2218R (all four combinations)	(i) 5 cycles of 45 s at 94 °C, 45 s at 60 °C (decreasing 1 °C/cycle), 1.5 min at 72 °C
		(ii) 35 cycles of 45 s at 94 °C, 45 s at 55 °C, 1.5 min at 72 °C
12S	MS1/MS2, or DacryMS1/Dacry12S-2r, or DacryMS1/Dacry12S-4r	(i) 5 cycles of 45 s at 94 °C, 45 s at 53 °C (decreasing 1 °C/cycle), 1 min at 72 °C
		(ii) 35 cycles of 45 s at 94 °C, 45 s at 48 °C, 1 min at 72 °C
ATP6	ATP6-3/ATP6-2 or ATP6-3/ATP6-4	Programme according to Kretzer & Bruns (1999)