Table 3

Absolute quantification via limiting dilution assay.

Dilution

Total

Nil

N per

Target

Factor

Rxns

Rxns

Aliquot

LDA-No


CHS

15,000

48

30

0.47

7,050

SE

6,222

48

22

0.78

4,854

PHB

6,222

48

28

0.54

3,354

EMF2

8,000

48

33

0.37

2,998

NPR1

3,000

48

26

0.61

1,839

ANT

1,250

48

20

0.88

1,094

FLF

500

24

9

0.98

490

HAP3C

466

48

29

0.50

235

ABI3

250

48

28

0.54

135

AP3

67

48

19

0.93

62

Lec2

20

48

27

0.58

12


Based upon the N0 values produced by LRE quantification, a sample of each reverse transcriptase reaction was diluted to a predicted 0.5–1.0 target molecules per aliquot for each of the eleven cDNA targets, and replicate amplification reactions were conducted to determine the frequency of aliquots lacking a target molecule (Nil Rxns). The number of target molecules (N) per aliquot was then calculated using equation 14 and the predicted number of target molecules in the reverse transcriptase reaction (LDA-No) was calculated by multiplying by the dilution factor (see text for additional details).

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

Open Data