DECIPHER (version 2.0.2)

DesignArray: Design a set of DNA Microarray Probes for Detecting Sequences

Description

Chooses the set of microarray probes maximizing sensitivity and specificity to each target consensus sequence.

Usage

DesignArray(myDNAStringSet, maxProbeLength = 24, minProbeLength = 20, maxPermutations = 4, numRecordedMismatches = 500, numProbes = 10, start = 1, end = NULL, maxOverlap = 5, hybridizationFormamide = 10, minMeltingFormamide = 15, maxMeltingFormamide = 20, minScore = -1e+12, processors = 1, verbose = TRUE)

Arguments

myDNAStringSet

A DNAStringSet object of aligned consensus sequences.

maxProbeLength

The maximum length of probes, not including the poly-T spacer. Ideally less than 27 nucleotides.

minProbeLength

The minimum length of probes, not including the poly-T spacer. Ideally more than 18 nucleotides.

maxPermutations

The maximum number of probe permutations required to represent a target site. For example, if a target site has an 'N' then 4 probes are required because probes cannot be ambiguous. Typically fewer permutations are preferably because this requires less space on the microarray and simplifies interpretation of the results.

numRecordedMismatches

The maximum number of recorded potential cross-hybridizations for any target site.

numProbes

The target number of probes on the microarray per input consensus sequence.

start

Integer specifying the starting position in the alignment where potential forward primer target sites begin. Preferably a position that is included in most sequences in the alignment.

end

Integer specifying the ending position in the alignment where potential reverse primer target sites end. Preferably a position that is included in most sequences in the alignment.

maxOverlap

Maximum overlap in nucleotides between target sites on the sequence.

hybridizationFormamide

The formamide concentration (%, vol/vol) used in hybridization at 42 degrees Celsius. Note that this concentration is used to approximate hybridization efficiency of cross-amplifications.

minMeltingFormamide

The minimum melting point formamide concentration (%, vol/vol) of the designed probes. The melting point is defined as the concentration where half of the template is bound to probe.

maxMeltingFormamide

The maximum melting point formamide concentration (%, vol/vol) of the designed probes. Must be greater than the minMeltingFormamide.

minScore

The minimum score of designed probes before exclusion. A greater minScore will accelerate the code because more target sites will be excluded from consideration. However, if the minScore is too high it will prevent any target sites from being recorded.

processors

The number of processors to use, or NULL to automatically detect and use all available processors.

verbose

Logical indicating whether to display progress.

Value

A data.frame with the optimal set of probes matching the specified constraints. Each row lists the probe's target sequence (name), start position, length in nucleotides, start and end position in the sequence alignment, number of permutations, score, melt point in percent formamide at 42 degrees Celsius, hybridization efficiency (hyb_eff), target site, and probe(s). Probes are designed such that the stringency is determined by the equilibrium hybridization conditions and not subsequent washing steps.

Details

The algorithm begins by determining the optimal length of probes required to meet the input constraints while maximizing sensitivity to the target consensus sequence at the specified hybridization formamide concentration. This set of potential target sites is then scored based on the possibility of cross-hybridizing to the other non-target sequences. The set of probes is returned with the minimum possibility of cross-hybridizing.

References

ES Wright et al. (2013) Identification of Bacterial and Archaeal Communities From Source to Tap. Water Research Foundation, Denver, CO.

DR Noguera, et al. (2014). Mathematical tools to optimize the design of oligonucleotide probes and primers. Applied Microbiology and Biotechnology. doi:10.1007/s00253-014-6165-x.

See Also

Array2Matrix, NNLS

Examples

fas <- system.file("extdata", "Bacteria_175seqs.fas", package="DECIPHER")
dna <- readDNAStringSet(fas)
names(dna) <- 1:length(dna)
probes <- DesignArray(dna)
probes[1,]