Table 1

LRE-based absolute quantification of eleven cDNA targets using five enzyme formulations.

Gain:

X8

X8

X2

X2

X1

OCF:

1092

1155

704

778

345

Enzyme:

QT 0XSG

QT 0.2XSG

DyNa 0.5XSG

DyNa 1.5XSG

FV 2.0XSG

LRE-No Average

CV (+/-)

Target


CHS K3K4

6,143

7,323

5,658

8,500

11,598

7,844

30.2%

SE K3K2

4,828

5,444

5,103

6,363

nd

5,435

12.3%

PHB K3K2

2,951

3,292

2,646

3,733

2,956

3,116

13.3%

EMF2 K2K3

2,301

3,256

3,094

2,752

nd

2,851

14.8%

NPR1 K1K4

1,872

1,973

1,565

1,657

2,334

1,880

16.0%

ANT K3K4

970

1,390

756

1,139

1,446

1,140

25.3%

FLF K3K4

661

553

466

596

nd

569

14.4%

HAP3C K3K4

145

198

254

255

nd

213

24.7%

ABI3 K3K4

66

105

129

122

48

94

37.8%

AP3 K3K4

38

52

61

34

22

41

37.3%

Lec2 K3K4

6

14

12

12

9

11

31.4%


Av. CV:

23.4%


Following LRE analysis (summarized in Figure 7), the FC readings encompassed by the LRE window were converted to F0 (see Figure 3) and the average F0 value converted to the number of target molecules (LRE-N0) using the OCF generated by each respective reaction formulation (see Figure 6 and additional file 3 for more details). The gain setting was adjusted to prevent saturation of the photomultiplier (see Figure 2). Gain: the photomultiplier gain setting, OCF: optical calibration factor (FU/ng dsDNA), QT: QuantiTect, DyNa: DyNAmo; FV: FullVelocity, SG: SYBR Green I, SD: standard deviation, nd: not determined.

Rutledge and Stewart BMC Biotechnology 2008 8:47   doi:10.1186/1472-6750-8-47

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