Probability-directed isolation of nucleotide sequences (PINS)

An in vitro method for enriching for a pre-defined target DNA molecule containing a known unique sequence of consecutive nucleotides from a mixed polynucleotide sample comprising the steps of:

• a) providing a mixed polynucleotide sample containing said target DNA molecule and non-target DNA molecules, wherein said target DNA molecule comprises a known unique sequence of at least 10 consecutive nucleotides which is distinguishable from sequences in the non-target DNA molecules,
• b) serially diluting said mixed polynucleotide sample until the probability that the target DNA molecule is present in a diluted sample of mixed polynucleotides is less than 0.75,
• c) producing a sufficient number of replicates of the diluted sample of mixed polynucleotides until the probability that the target DNA molecule is present in at least one of the replicate dilution samples is at least 0.75;
• d) amplifying all DNA in said replicate diluted samples to increase the abundance of the DNA in each sample;
• wherein the frequency of said target DNA molecule in said at least one replicate dilution sample is increased compared to the mixed polynucleotide sample in step (a);
• and
• wherein the probability in (b) and (c) is calculated using a hypergeometric distribution according to the following equation:
• P
• ⁡
• (
• X
• =
• x
• )
• =
• h
• ⁡
• (
• x
• ;
• n
• ,
• M
• ,
• N
• )
• =
• (
• M
• x
• )
• ⁢
• (
• N
• -
• M
• n
• -
• x
• )
• (
• N
• n
• )
• where x is the number of target DNA molecules in the diluted sample, n is the volume of the diluted sample multiplied by 1,000, M is the number of target DNA molecules in the mixed polynucleotide sample, and N is the volume of the mixed polynucleotide sample multiplied by 1,000.