Package {TmCalculator}

Type:	Package
Title:	Genome-Wide Nucleic Acid Melting Temperature Profiling and Multi-Omics Integration
Version:	1.0.6
Date:	2026-06-12
Description:	Accurate calculation of nucleic acid melting temperature (Tm) is fundamental to many molecular biology applications, and this software scales Tm analysis from individual sequences to genome‑wide thermodynamic profiling. This package extends Tm analysis from simple sequence level computation to comprehensive genome-wide thermodynamic profiling. It takes multiple input formats including sequence strings, FASTA files, genomic coordinates. The implementation provides three Tm calculation methods: the Wallace rule (Thein & Wallace, 1986), empirical GC‑content formulas (Marmur, 1962; Schildkraut, 2010; Wetmur, 1991; Untergasser, 2012; von Ahsen, 2001), and nearest‑neighbor thermodynamics (Breslauer, 1986; Sugimoto, 1996; Allawi, 1998; SantaLucia, 2004; Freier, 1986; Xia, 1998; Chen, 2012; Bommarito, 2000; Turner, 2010; Sugimoto, 1995; Allawi, 1997; SantaLucia, 2005). Corrections are supported for salt ions (SantaLucia, 1996, 1998; Owczarzy, 2004, 2008) and for chemical conditions such as dimethyl sulfoxide and formamide. This package returns result as a GRanges object for interoperability with Bioconductor workflows and downstream multi-omics analyses. Data-level integration reconciles Tm windows with external multi-omics GRanges objects through overlap, nearest-feature, windowed-count, and binned-average strategies, returning a single unified GRanges object ready for downstream analysis. Visualization-level integration renders multiple feature layers as independent concentric tracks on a shared genomic axis, each retaining its native coordinate resolution. Group comparison supports Wilcoxon rank-sum and Student's t-tests with multiple available correction methods for contrasting Tm and other features across region classes.
BugReports:	https://github.com/JunhuiLi1017/TmCalculator/issues
License:	MIT + file LICENSE
Depends:	R (≥ 3.5)
VignetteBuilder:	knitr
Imports:	BSgenome, BiocGenerics, Biostrings, GenomeInfoDb, GenomicRanges, IRanges, S4Vectors, graphics, grDevices, methods
Suggests:	testthat (≥ 3.0.0), knitr, utils, rmarkdown, remotes, BiocManager, BSgenomeForge, BSgenome.Hsapiens.UCSC.hg38, karyoploteR, ggridges, ggforce, ggplot2, rlang, seqinr
NeedsCompilation:	no
Repository:	CRAN
RoxygenNote:	7.3.2
LazyData:	true
LazyDataCompression:	xz
Encoding:	UTF-8
Packaged:	2026-06-13 05:24:04 UTC; lij11
Author:	Junhui Li [cre, aut], Lihua Julie Zhu [aut]
Maintainer:	Junhui Li <ljh.biostat@gmail.com>
Date/Publication:	2026-06-13 16:30:08 UTC

TmCalculator: Genome-Wide Nucleic Acid Melting Temperature Profiling and Multi-Omics Integration

Description

Accurate calculation of nucleic acid melting temperature (Tm) is fundamental to many molecular biology applications, and this software scales Tm analysis from individual sequences to genome‑wide thermodynamic profiling. This package extends Tm analysis from simple sequence level computation to comprehensive genome-wide thermodynamic profiling. It takes multiple input formats including sequence strings, FASTA files, genomic coordinates. The implementation provides three Tm calculation methods: the Wallace rule (Thein & Wallace, 1986), empirical GC‑content formulas (Marmur, 1962; Schildkraut, 2010; Wetmur, 1991; Untergasser, 2012; von Ahsen, 2001), and nearest‑neighbor thermodynamics (Breslauer, 1986; Sugimoto, 1996; Allawi, 1998; SantaLucia, 2004; Freier, 1986; Xia, 1998; Chen, 2012; Bommarito, 2000; Turner, 2010; Sugimoto, 1995; Allawi, 1997; SantaLucia, 2005). Corrections are supported for salt ions (SantaLucia, 1996, 1998; Owczarzy, 2004, 2008) and for chemical conditions such as dimethyl sulfoxide and formamide. This package returns result as a GRanges object for interoperability with Bioconductor workflows and downstream multi-omics analyses. Data-level integration reconciles Tm windows with external multi-omics GRanges objects through overlap, nearest-feature, windowed-count, and binned-average strategies, returning a single unified GRanges object ready for downstream analysis. Visualization-level integration renders multiple feature layers as independent concentric tracks on a shared genomic axis, each retaining its native coordinate resolution. Group comparison supports Wilcoxon rank-sum and Student's t-tests with multiple available correction methods for contrasting Tm and other features across region classes.

Author(s)

Maintainer: Junhui Li ljh.biostat@gmail.com (ORCID)

Authors:

• Lihua Julie Zhu julie.zhu@umassmed.edu (ORCID)

See Also

Useful links:

• Report bugs at https://github.com/JunhuiLi1017/TmCalculator/issues

Filter windows with too many N bases

Description

Reads each window sequence and drops those where the N fraction exceeds max_N_frac. Uses batched getSeq for efficiency.

Usage

.filter_N_windows(bsgenome, chr, win_starts, win_ends, max_N_frac, window)

Detect the first and last non-N positions on a chromosome

Description

Uses a coarse block scan to find the positions of the first and last non-N base on a chromosome. Efficient because it only reads N_scan_block bp at a time from each end.

Usage

.find_N_bounds(bsgenome, chr, chr_start, chr_end, block_size)

Load the BSgenome object from an installed BSgenome.* data package

Description

The genome object name is given by the BSgenomeObjname field in DESCRIPTION (e.g. Hsapiens), not necessarily the package name.

Usage

.get_bsgenome_from_pkg(pkg)

Arguments

Value

A BSgenome object.

Sequence extraction by preloading whole chromosomes

Description

Sequence extraction by preloading whole chromosomes

Usage

.getseq_preload_chr(gr_query, genome_objs, parsed)

Arguments

Value

DNAStringSet of extracted sequences.

Vectorized sequence extraction with getSeq

Description

Vectorized sequence extraction with getSeq

Usage

.getseq_vectorized(gr_query, genome_objs, parsed)

Arguments

Value

DNAStringSet of extracted sequences.

Load installed BSgenome packages

Description

Load installed BSgenome packages

Usage

.load_genome_packages(pkg_names)

Arguments

Value

Named list of genome objects.

Normalize Tm/GC metadata column names on a GRanges object

Description

Renames legacy lowercase tm/gc columns to Tm/GC.

Usage

.normalize_tm_gc_metadata(gr)

Parse coordinate strings into a data frame

Description

Parse coordinate strings into a data frame

Usage

.parse_coord_strings(coord_vec, pkg_name_override = NULL)

Arguments

Value

Data frame with columns chr, win_start, win_end, strand, pkg_name, and region_id.

convert a vector of characters into a string

Description

Simply convert a vector of characters such as c("H","e","l","l","o","W","o","r","l","d") into a single string "HelloWorld".

Usage

c2s(characters)

Arguments

Value

Retrun a strings

Author(s)

Junhui Li

References

citation("TmCalculator")

Filter invalid bases in nucleotide sequences

Description

This function processes nucleotide sequences by converting characters to uppercase and replacing invalid bases with "". based on the specified method. The function preserves the sequence length and attributes (name and Tm) of each sequence.

Usage

check_filter_seq(seq_list, method)

Arguments

Value

Returns a list of sequences with the same structure as input, where invalid bases are replaced with ""

Author(s)

Junhui Li

References

citation("TmCalculator")

Corrections of melting temperature with chemical substances

Description

Apply corrections to melting temperature calculations based on the presence of DMSO and formamide. These corrections are rough approximations and should be used with caution.

Usage

chem_correct(
  DMSO = 0,
  formamide_unit = list(value = 0, unit = "percent"),
  dmso_factor = 0.75,
  formamide_factor = 0.65,
  pt_gc = NULL
)

Arguments

Details

When formamide_unit$unit = "percent": Correction = - factor * percentage_of_formamide

When formamide_unit$unit = "molar": Correction = (0.453 * GC/100 - 2.88) * formamide

Author(s)

Junhui Li

References

von Ahsen N, Wittwer CT, Schutz E, et al. Oligonucleotide melting temperatures under PCR conditions: deoxynucleotide Triphosphate and Dimethyl sulfoxide concentrations with comparison to alternative empirical formulas. Clin Chem 2001, 47:1956-1961.

Examples

# DMSO correction
chem_correct(DMSO = 3)

# Formamide correction (percent)
chem_correct(formamide_unit = list(value = 1.25, unit = "percent"), pt_gc = 50)

# Formamide correction (molar)
chem_correct(formamide_unit = list(value = 1.25, unit = "molar"), pt_gc = 50)

Compare numeric GRanges metadata across groups

Description

Test whether one or more numeric metadata columns (e.g. Tm, GC) differ between levels of a categorical grouping column in a GRanges object.

Usage

compare_groups(
  gr,
  target = "Tm",
  method = c("wilcoxon", "t.test"),
  group,
  min_n_per_group = 2L,
  paired = FALSE,
  alternative = c("two.sided", "less", "greater"),
  posthoc = TRUE,
  p.adjust.method = c("holm", "hochberg", "hommel", "bonferroni", "BH", "BY", "fdr",
    "none")
)

Arguments

Details

The test used depends on method and the number of group levels:

When there are three or more groups and posthoc = TRUE, pairwise follow-up tests are run (Wilcoxon or Welch t-test) with p.adjust multiple-testing correction.

Value

A list with:

results

Data frame with one row per target: omnibus test name, statistic, p.value, and group information.

summary

Per-group descriptive statistics (n, mean, sd, median).

pairwise

Pairwise comparisons when posthoc = TRUE and there are \ge 3 groups; otherwise NULL.

Author(s)

Junhui Li

Examples

## Not run: 
library(GenomicRanges)
set.seed(1)
gr <- GRanges(
  seqnames = rep("chr1", 90),
  ranges = IRanges(start = sort(sample(1e6, 90)), width = 50),
  Tm = c(rnorm(30, 74, 2), rnorm(30, 70, 2), rnorm(30, 66, 2)),
  GC = runif(90, 40, 60)
)
gr$group <- rep(c("high", "mid", "low"), each = 30)

# Two groups
gr2 <- gr[gr$group %in% c("high", "low")]
compare_groups(gr2, target = "Tm", group = "group", posthoc = FALSE)

# Three or more groups (Kruskal-Wallis + pairwise Wilcoxon)
compare_groups(gr, target = c("Tm", "GC"), group = "group",
                     method = "wilcoxon")

# Three or more groups (one-way ANOVA + pairwise t-tests)
compare_groups(gr, target = "Tm", group = "group", method = "t.test")

# Post-hoc with Benjamini-Hochberg FDR control
compare_groups(gr, target = "Tm", group = "group",
                     p.adjust.method = "BH")

## End(Not run)

Fast complement and reverse complement

Description

Uses chartr() instead of character-by-character translation. ~10x faster than the seqinr-style implementation for long sequences.

Usage

complement_fast(seq_str, rev = FALSE)

Arguments

Value

Character string(s) with complement.

Convert genomic coordinate strings to a GRanges object

Description

Fast conversion of genomic coordinate strings to a GRanges object with reference sequences fetched from installed BSgenome.* packages. Designed for large sliding-window inputs: genome packages are loaded once, coordinate strings are parsed in one pass, and sequences are extracted with vectorized getSeq (or optional chromosome preloading).

Usage

coor_to_genomic_ranges(
  input,
  complement_seq = NULL,
  method = c("vectorized", "preload_chr")
)

Arguments

Details

For genome-wide tiling with thousands of windows, pass coordinates as list(pkg_name = "BSgenome.Hsapiens.UCSC.hg38", seq = ...) so the genome package is loaded once instead of per interval.

Value

A GRanges object with metadata columns:

sequence

Reference sequence for each interval.

complement

Complementary sequence.

GC

GC fraction (0-1) per interval.

region_id

Region identifier from the coordinate string.

genome_pkg

BSgenome package name used.

Author(s)

Junhui Li

See Also

to_genomic_ranges, to_genomic_ranges_fast

Examples

## Not run: 
coords <- c(
  "chr1:1000-1199:+:win1",
  "chr1:1200-1399:+:win2"
)
gr <- coor_to_genomic_ranges(
  list(pkg_name = "BSgenome.Hsapiens.UCSC.hg38", seq = coords)
)
gr

## End(Not run)

E. coli K-12 MG1655 replication-associated hotspot annotations

Description

A named list of genomic feature tables for *Escherichia coli* K-12 MG1655 (NCBI assembly GCF_000005845.2 / ASM584v2, chromosome U00096.3). The data support the genome-wide Tm vignette and plot_genome_track examples by providing independent multi-omics layers that can be overlaid on the same genomic axis alongside Tm profiles computed with tm_calculate.

Usage

ecoli_rep_hotspots

Format

A named list with five elements:

all_peaks_IP_mutH

A data frame (38 rows) of MutL protein ChIP-seq peaks marking mismatch-repair-associated regions (MutL-AR). Columns: chr, start, end, Sample, name, Size.

bins_rep

A data frame (4,642 rows) of tandem-repeat (microsatellite) counts in non-overlapping 1 kb bins. Columns: chr, start, end, count.

bins_cru

A data frame (4,642 rows) of cruciform-forming sequence counts in non-overlapping 1 kb bins. Columns: chr, start, end, count.

ssdna

A data frame (2,636 rows) of single-stranded DNA regions. Columns: chr, start, end, Cells..strand., Region.

bins_gatc

A data frame (4,642 rows) of GATC methylation-site counts in non-overlapping 1 kb bins. Columns: chr, start, end, count.

All coordinate-based tables use chr = "U00096.3" and are compatible with plot_genome_track and compare_groups.

Source

Curated from published *E. coli* K-12 MG1655 multi-omics datasets used in the package vignette vignette("genome_wide_tm_ecoli", package = "TmCalculator").

Examples

data(ecoli_rep_hotspots)
names(ecoli_rep_hotspots)

# MutL-AR peak coordinates
head(ecoli_rep_hotspots$all_peaks_IP_mutH)

# Microsatellite density in 1 kb bins
summary(ecoli_rep_hotspots$bins_rep$count)

Convert FASTA file to GenomicRanges object

Description

This function reads sequences from a FASTA file and converts them to a GenomicRanges object. If named with format ">chr2:1-10:[+|-]:[seq_name]", the name will be parsed into GRanges components.

Usage

fa_to_genomic_ranges(input_seq)

Arguments

Value

A GenomicRanges object containing: - GRanges information (seqnames, ranges, strand) - sequence data from FASTA file - Complementary sequences (if provided) - Names from FASTA headers

Examples

# Example with single FASTA file
input_seq <- system.file("extdata", "example1.fasta", package = "TmCalculator")
gr <- fa_to_genomic_ranges(input_seq)

Calculate G and C content of nucleotide sequences

Description

Calculate G and C content of nucleotide sequences. The function calculates the percentage of G and C bases relative to the total number of A, T, G, and C bases in the sequence.

Usage

gc(input_seq, ambiguous = FALSE)

Arguments

Value

Content of G and C as a percentage (range from 0 to 100

Author(s)

Junhui Li

Examples

# Calculate GC content of a DNA sequence
gc(c("a","t","c","t","g","g","g","c","c","a","g","t","a"))  # 53.85%

# Calculate GC content including ambiguous bases
gc("GCATSWSYK", ambiguous = TRUE)  # 55.56%

Generate complementary sequence

Description

Generate the complementary sequence of a nucleic acid sequence, with an option to reverse it.

Usage

generate_complement(input_seq, reverse = FALSE)

Arguments

Value

Returns the complementary sequence(s) in the specified direction.

Author(s)

Junhui Li

References

citation("TmCalculator")

Examples

# Generate complementary sequence in same direction (5' to 3')
generate_complement("ATGCG", reverse = FALSE)

# Generate complementary sequence in reverse direction (3' to 5')
generate_complement("ATGCG", reverse = TRUE)

Integrate a Tm GRanges with multi-omic feature ranges

Description

Combines the output of tm_calculate (a GRanges object with Tm and GC columns) with a second GRanges carrying arbitrary multi-omic metadata (ChIP-seq peaks, ATAC-seq signal, methylation sites, gene annotations, etc.) using one of four positional strategies:

"overlap"

Each tm range is annotated with the aggregated metadata of all feature ranges it directly overlaps.

"nearest"

Each tm range is annotated with the metadata of its single closest feature range, plus an added distance column.

"window"

Each tm range is expanded symmetrically by window_size bp and annotated with aggregated metadata from all features that fall within the expanded window.

"bin"

The genomic space covered by the data is tiled into equal-width bins. Each bin is annotated with the mean tm / GC of overlapping tm ranges and the aggregated feature values - suitable for joint heatmaps and genome-wide correlation analyses.

For strategies "overlap" and "window", when a single Tm range matches multiple features the default behaviour is to summarise: numeric columns are aggregated via agg_fun (default mean), and categorical columns are collapsed to a comma-separated string of unique values.

Usage

integrate_granges(
  gr_tm,
  gr_features,
  strategy = c("overlap", "nearest", "window", "bin"),
  feature_cols = NULL,
  prefix = "",
  window_size = 1000L,
  bin_size = 1e+06,
  agg_fun = mean,
  min_overlap = 1L,
  ignore_strand = TRUE,
  keep_unmatched = TRUE,
  distance_col = "distance_to_feature"
)

Arguments

Value

• "overlap", "nearest", "window": A GRanges object with the same ranges as gr_tm (minus unmatched ranges if keep_unmatched = FALSE), with additional metadata columns from gr_features.

• "bin": A new GRanges of genomic bins. Each bin carries Tm_mean, GC_mean (if available), n_tm_ranges, n_features, and one aggregated column per requested feature column.

Author(s)

Junhui Li

Examples

## Not run: 
library(GenomicRanges)

# -- Sample data ----------------------------------------------------------
set.seed(42)
gr_tm <- GRanges(
  seqnames = c(rep("chr1", 60), rep("chr2", 30)),
  ranges   = IRanges(
    start = c(sort(sample(1:249e6, 60)),
              sort(sample(1:243e6, 30))),
    width = sample(50:200, 90, replace = TRUE)
  ),
  Tm = runif(90, 55, 85),
  GC = runif(90, 30, 70)
)

gr_features <- GRanges(
  seqnames = c(rep("chr1", 40), rep("chr2", 20)),
  ranges   = IRanges(
    start = c(sort(sample(1:249e6, 40)),
              sort(sample(1:243e6, 20))),
    width = sample(500:5000, 60, replace = TRUE)
  ),
  score      = runif(60, 0, 100),
  peak_type  = sample(c("narrow", "broad"), 60, replace = TRUE),
  signal     = rnorm(60, 5, 2)
)

# Strategy 1: overlap - annotate Tm ranges with overlapping peak features
res_overlap <- integrate_granges(gr_tm, gr_features,
                                  strategy = "overlap")

# Strategy 2: nearest - every Tm range gets its closest peak + distance
res_nearest <- integrate_granges(gr_tm, gr_features,
                                  strategy = "nearest")
head(res_nearest$distance_to_feature)

# Strategy 3: window - 5 kb window around each probe
res_window <- integrate_granges(gr_tm, gr_features,
                                 strategy = "window", window_size = 5000)

# Strategy 4: bin - 500 kb genome bins with mean Tm and aggregated signal
res_bin <- integrate_granges(gr_tm, gr_features,
                              strategy = "bin", bin_size = 5e5)
as.data.frame(res_bin) |> head()

# Use a subset of feature columns and add a prefix
integrate_granges(gr_tm, gr_features,
                  strategy    = "overlap",
                  feature_cols = c("score", "peak_type"),
                  prefix       = "chip_")

## End(Not run)

Generate sliding-window genomic coordinate strings for Tm calculation

Description

Tiles one or more chromosomes from a BSgenome object into overlapping windows and returns a named character vector of coordinate strings in the format "chr:start-end:strand:genome_pkg:region_id". This vector is the primary input for downstream Tm calculation functions (tm_nn, tm_gc, tm_wallace) that accept genomic coordinate strings.

Usage

make_genomiccoord(
  bsgenome,
  chromosomes = NULL,
  window = 200L,
  slide = 50L,
  start = NULL,
  end = NULL,
  strand = "+",
  trim_N = c("ends", "filter", "none"),
  max_N_frac = 0.1,
  N_scan_block = window,
  region_prefix = "region",
  genome_pkg_name = NULL,
  as_vector = TRUE,
  verbose = TRUE
)

Arguments

Value

When as_vector = TRUE (default): a named character vector of coordinate strings, one element per window. Names are the region IDs (region1, region2, ...).

When as_vector = FALSE: a data.frame with columns coord, chr, win_start, win_end, strand, genome, region_id, chr_start_used, chr_end_used.

Coordinate string format

Each element of the returned vector follows the pattern:

  chr1:10001-10200:+:BSgenome.Hsapiens.UCSC.hg38:region1
  chr1:10051-10250:+:BSgenome.Hsapiens.UCSC.hg38:region2
  ...

Fields (colon-separated):

1. chromosome – e.g. chr1

2. start-end – 1-based, inclusive coordinates

3. strand – + or -

4. genome – BSgenome package name (character)

5. region_id – unique label regionN

N-base trimming

Human (and most mammalian) chromosomes begin and end with long stretches of N bases that represent assembly gaps. Windows that consist entirely or predominantly of Ns produce meaningless Tm values. The function offers two trimming strategies controlled by trim_N:

"none"

No trimming. Windows start at start and end at end (or chromosome length).

"ends"

Detect the first and last positions of non-N bases on each chromosome using Biostrings::letterFrequency() on a coarse block scan, then trim start and end to those positions. Efficient: reads the chromosome sequence only once in blocks.

"filter"

Generate windows across the full start-end range but then remove any window whose N-base fraction exceeds max_N_frac (default 0.1, i.e. 10%). More granular than "ends" but slower because it reads each window sequence.

Author(s)

Junhui Li

See Also

tm_nn, tm_gc, tm_wallace, BSgenome, letterFrequency

Examples

## Not run: 
library(BSgenome.Hsapiens.UCSC.hg38)

## -- Basic usage: tile chr1 with 200 bp windows, 50 bp slide ----------
coords <- make_genomiccoord(
  bsgenome    = BSgenome.Hsapiens.UCSC.hg38,
  chromosomes = "chr1",
  window      = 200L,
  slide       = 50L
)
length(coords)      # number of windows on chr1
head(coords, 3)
# region1 "chr1:10001-10200:+:BSgenome.Hsapiens.UCSC.hg38:region1"
# region2 "chr1:10051-10250:+:BSgenome.Hsapiens.UCSC.hg38:region2"
# region3 "chr1:10101-10300:+:BSgenome.Hsapiens.UCSC.hg38:region3"

## -- Non-overlapping tiling (slide == window) -------------------------
coords_nonoverlap <- make_genomiccoord(
  bsgenome    = BSgenome.Hsapiens.UCSC.hg38,
  chromosomes = paste0("chr", 1:22),
  window      = 200L,
  slide       = 200L    # no overlap
)
length(coords_nonoverlap)   # ~15 million windows across autosomes

## -- Custom start/end (e.g. a specific sub-region) --------------------
coords_sub <- make_genomiccoord(
  bsgenome    = BSgenome.Hsapiens.UCSC.hg38,
  chromosomes = "chr1",
  window      = 200L,
  slide       = 50L,
  start       = 1000000L,
  end         = 2000000L
)
length(coords_sub)   # 19,981 windows in 1 Mb region

## -- No N-trimming (use full chromosome length) ------------------------
coords_noN <- make_genomiccoord(
  bsgenome    = BSgenome.Hsapiens.UCSC.hg38,
  chromosomes = "chr1",
  window      = 200L,
  slide       = 50L,
  trim_N      = "none"
)

## -- Per-window N-filtering (removes windows with >10% N) -------------
coords_filt <- make_genomiccoord(
  bsgenome    = BSgenome.Hsapiens.UCSC.hg38,
  chromosomes = "chr1",
  window      = 200L,
  slide       = 50L,
  trim_N      = "filter",
  max_N_frac  = 0.10
)

## -- Get data.frame output for GRanges construction -------------------
df <- make_genomiccoord(
  bsgenome    = BSgenome.Hsapiens.UCSC.hg38,
  chromosomes = "chr1",
  window      = 200L,
  slide       = 50L,
  as_vector   = FALSE
)
gr <- GenomicRanges::GRanges(
  seqnames = df$chr,
  ranges   = IRanges::IRanges(start = df$win_start, end = df$win_end),
  strand   = df$strand
)

## -- Pass directly to tm_nn -------------------------------------------
coords <- make_genomiccoord(
  bsgenome    = BSgenome.Hsapiens.UCSC.hg38,
  chromosomes = "chr1",
  window      = 200L,
  slide       = 200L,
  start       = 1000000L,
  end         = 1010000L
)
tm_results <- tm_nn(coords, Na = 50)

## End(Not run)

Plot genome tracks in linear or circular layout

Description

Visualise genomic tracks as either a linear karyoploteR plot or a circular plot (base R graphics). Set circular = TRUE to switch to the circular layout; the same track_list works for both views without modification.

Supported track types in each list element:

• type = "rect" : data as GRanges/data.frame, drawn as rectangles

• type = "line" : data as GRanges/data.frame with value_col, drawn as lines

• type = "area" : same as line but filled (linear only)

• type = "region": piled-up regions (linear only)

• type = "coverage": coverage area plot (linear only)

A track with ideogram = TRUE is drawn inside the chromosome bar (linear) or as the outermost ring with a grey background (circular). Only "rect" type is supported for ideogram tracks.

A per-track height field (numeric, default 1) controls relative sizing. In linear mode heights are proportional fractions of the data panel; in circular mode they scale the radial thickness of each ring.

A per-track highlight field (list or list-of-lists with data/col/alpha/border) draws highlight bands within that track only.

Entries with type = "highlight" draw translucent bands spanning all real tracks. These accept: data, col (default "#F1C40F"), alpha (default 0.18), border (default NA), and min.degree (circular only, default 0.4).

Each list element may also contain: name, col, bg.col, border, ylim, lwd, alpha (0–1 transparency), legend_font_col, ideogram, height, highlight.

Usage

plot_genome_track(
  genome_name,
  genome_size = NULL,
  genome = NULL,
  track_list,
  circular = FALSE,
  label = NULL,
  chromosomes = NULL,
  zoom = NULL,
  plot.type = 1,
  track.gap = 0.01,
  legend.show = TRUE,
  legend.position = NULL,
  legend.cex = 0.6,
  legend.bty = "n",
  legend.border = NA,
  legend.font.col = "black",
  legend.lwd = 1,
  legend.seg.len = -0.5,
  legend.box.lwd = 0.25,
  legend.x.intersp = 1,
  legend.lty = 1,
  axis.cex = NULL,
  title.cex = NULL,
  base.tick.dist = NULL,
  start.degree = 34.47,
  track.height = 0.05,
  gap.after = 0,
  cell.padding = c(0, 0, 0, 0),
  track.margin = c(0.005, 0.005),
  circle.margin = c(0.001, 0.001),
  canvas.xlim = c(-1, 1),
  canvas.ylim = c(-1, 1),
  axis.unit = "Mb",
  axis.step = NULL,
  axis.show.unit = FALSE,
  label.column = 4,
  label.niceFacing = TRUE,
  label.cex = 0.6,
  label.side = "inside",
  label.labels_height = 0.01,
  label.connection_height = 0.03,
  label.line_lwd = 0.25
)

Arguments

Value

Invisibly returns the KaryoPlot object (linear) or NULL (circular).

Examples

## Not run: 
data(ecoli_rep_hotspots)

library("BSgenome.Ecoli.NCBI.ASM584v2")
genome_name <- "BSgenome.Ecoli.NCBI.ASM584v2"
chr_name    <- "U00096.3"
genome <- get(genome_name, envir = asNamespace(genome_name))
chr_length  <- length(genome[[chr_name]])
genome_name="BSgenome.Ecoli.NCBI.ASM584v2"
bins_gc <- make_genomiccoord(
  bsgenome    = genome_name,
  chromosomes = chr_name,
  window      = 200L,
  slide       = 200L,
  start       = 1,
  end         = chr_length,
  strand      = "+"
)
input_new <- list(pkg_name = genome_name, seq = bins_gc)
gr_batch <- to_genomic_ranges_fast(input_new)
tm_ASM584v2 <- tm_calculate(
  gr_batch,
  method   = "tm_nn"
)
Tm <- as.data.frame(tm_ASM584v2$gr[, c("Tm", "GC")])

tracks <- list(
  list(type = "rect", data = ecoli_rep_hotspots$all_peaks_IP_mutH,
       col = "#2C3E50", bg.col = "grey", name = "MutL-AR",
       legend_font_col = "#2C3E50", ideogram = TRUE, height = 0.5),
  list(type = "line", data = Tm, value_col = "GC",
       name = "GC content", col = "#4A90E2",
       legend_font_col = "#4A90E2"),
  list(type = "line", data = Tm, value_col = "Tm",
       name = "Melting temp", col = "#E06666",
       legend_font_col = "#E06666", height = 2),
  list(type = "line", data = ecoli_rep_hotspots$bins_rep,
       value_col = "count", name = "Microsatellites", col = "#2ECC71",
       legend_font_col = "#2ECC71"),
  list(type = "line", data = ecoli_rep_hotspots$bins_cru,
       value_col = "count", name = "Cruciform", col = "#3B3E6B",
       legend_font_col = "#3B3E6B"),
  list(data = ecoli_rep_hotspots$ssdna, name = "ssDNA",
       col = "#8E44AD", legend_font_col = "#8E44AD"),
  list(type = "line", data = ecoli_rep_hotspots$bins_gatc,
       value_col = "count", name = "GATC sites", col = "#D35400",
       legend_font_col = "#D35400"),
  list(type = "highlight", data = ecoli_rep_hotspots$all_peaks_IP_mutH,
       col = "#F1C40F", alpha = 0.18)
)

# Circular
plot_genome_track("E. coli", genome_size = 4641652,
                  track_list = tracks, circular = TRUE)

# Linear
plot_genome_track("E. coli", genome_size = 4641652,
                  track_list = tracks)

# Linear zoom (single region)
plot_genome_track("E. coli", genome_size = 4641652,
                  track_list = tracks,
                  zoom = "U00096.3:1000000-2000000")

# Linear zoom (multiple regions — stacked panels)
plot_genome_track("E. coli", genome_size = 4641652,
                  track_list = tracks,
                  zoom = c("U00096.3:1000000-1200000",
                           "U00096.3:3000000-3200000"))

# Circular zoom (multiple regions — concatenated arcs)
plot_genome_track("E. coli", genome_size = 4641652,
                  track_list = tracks, circular = TRUE,
                  zoom = c("U00096.3:1000000-1200000",
                           "U00096.3:3000000-3200000"))

# Circular with canvas panning
plot_genome_track("E. coli", genome_size = 4641652,
                  track_list = tracks, circular = TRUE,
                  canvas.xlim = c(0.5, 1), canvas.ylim = c(0, 1),
                  circle.margin = c(0.05, 0.05))

## End(Not run)

Compare Tm distributions across groups

Description

Visualise how melting temperature (and optionally other numeric metadata) differs between region classes such as peak vs. non-peak, mutant vs. wild-type, or any categorical annotation stored in a GRanges object. Designed as the visual companion to compare_groups().

Usage

plot_tm(
  gr,
  group,
  value = "Tm",
  plot_type = c("box", "violin", "rank", "density", "ridgeline", "ecdf", "sina"),
  color_palette = c("viridis", "magma", "plasma", "inferno", "cividis"),
  show_points = FALSE,
  point_size = 1,
  point_alpha = 0.3,
  notch = FALSE,
  add_mean = FALSE,
  facet_by = NULL,
  show_pvalue = FALSE,
  p_adjust_method = "BH",
  title = NULL,
  ylab = "Tm (°C)",
  xlab = NULL,
  show_legend = TRUE
)

Arguments

Details

The function pairs naturally with integrate_granges() and compare_groups():

1. Use integrate_granges() to annotate Tm windows with feature overlaps (e.g. ChIP-seq peaks, repeat classes).

2. Use compare_groups() for the statistical test.

3. Use plot_tm() to visualise the distributional difference.

Value

A ggplot object.

Examples

## Not run: 
library(GenomicRanges)
data(ecoli_rep_hotspots)

library("BSgenome.Ecoli.NCBI.ASM584v2")
genome_name <- "BSgenome.Ecoli.NCBI.ASM584v2"
chr_name    <- "U00096.3"
genome <- get(genome_name, envir = asNamespace(genome_name))
chr_length  <- length(genome[[chr_name]])
genome_name="BSgenome.Ecoli.NCBI.ASM584v2"
bins_gc <- make_genomiccoord(
  bsgenome    = genome_name,
  chromosomes = chr_name,
  window      = 200L,
  slide       = 200L,
  start       = 1,
  end         = chr_length,
  strand      = "+"
)
input_new <- list(pkg_name = genome_name, seq = bins_gc)
gr_batch <- to_genomic_ranges_fast(input_new)
tm_ASM584v2 <- tm_calculate(
  gr_batch,
  method   = "tm_nn"
)
gr_tm <- tm_ASM584v2$gr

# Annotate with MutL-AR peak membership
mutH_peaks <- GRanges(
  seqnames = ecoli_rep_hotspots$all_peaks_IP_mutH$chr,
  ranges   = IRanges(start = ecoli_rep_hotspots$all_peaks_IP_mutH$start,
                     end   = ecoli_rep_hotspots$all_peaks_IP_mutH$end)
)
mutH_peaks$peak_id <- paste0("mutH_", seq_along(mutH_peaks))
gr_annot <- integrate_granges(
  gr_tm = gr_tm, gr_features = mutH_peaks,
  strategy = "overlap", feature_cols = "peak_id",
  keep_unmatched = TRUE
)
gr_annot$in_mutH <- ifelse(is.na(gr_annot$peak_id), "non_peak", "peak")

# Box plot — peak vs non-peak Tm
plot_tm(gr_annot, group = "in_mutH")

# Box plot with Wilcoxon p-value bracket
plot_tm(gr_annot, group = "in_mutH", show_pvalue = TRUE)

# Violin plot with p-value and jittered points
plot_tm(gr_annot, group = "in_mutH", plot_type = "violin",
        show_points = TRUE, show_pvalue = TRUE)

# Rank plot — visual Wilcoxon test with p-value subtitle
plot_tm(gr_annot, group = "in_mutH", plot_type = "rank",
        show_pvalue = TRUE)

# Density overlay with p-value
plot_tm(gr_annot, group = "in_mutH", plot_type = "density",
        show_pvalue = TRUE)

# ECDF — cumulative distribution comparison
plot_tm(gr_annot, group = "in_mutH", plot_type = "ecdf",
        show_pvalue = TRUE)

# Compare GC content instead of Tm
plot_tm(gr_annot, group = "in_mutH", value = "GC", ylab = "GC content",
        show_pvalue = TRUE)

# Notched box plot with mean markers and p-value
plot_tm(gr_annot, group = "in_mutH", notch = TRUE, add_mean = TRUE,
        show_pvalue = TRUE)

## End(Not run)

Prints melting temperature from a TmCalculator object

Description

print.TmCalculator prints to console the melting temperature value from an object of class TmCalculator.

Usage

## S3 method for class 'TmCalculator'
print(x, ...)

Arguments

Value

The melting temperature value.

convert a string into a vector of characters

Description

Simply convert a single string such as "HelloWorld" into a vector of characters such as c("H","e","l","l","o","W","o","r","l","d")

Usage

s2c(strings)

Arguments

Value

Retrun a vector of characters

Author(s)

Junhui Li

References

citation("TmCalculator")

Corrections of melting temperature with salt concentration

Description

Apply corrections to melting temperature calculations based on salt concentrations. Different correction methods are available for various experimental conditions.

Usage

salt_correction(
  Na = 0,
  K = 0,
  Tris = 0,
  Mg = 0,
  dNTPs = 0,
  method = c("Schildkraut2010", "Wetmur1991", "SantaLucia1996", "SantaLucia1998-1",
    "SantaLucia1998-2", "Owczarzy2004", "Owczarzy2008"),
  input_seq,
  ambiguous = FALSE
)

Arguments

Details

Different correction methods are available for various experimental conditions:

- Schildkraut2010: Updated salt correction method that accounts for monovalent and divalent cations - Wetmur1991: Classic salt correction method for monovalent cations - SantaLucia1996: DNA-specific salt correction - SantaLucia1998-1: Improved DNA salt correction - SantaLucia1998-2: Alternative DNA salt correction (requires sequence information) - Owczarzy2004: Comprehensive salt correction including effects of divalent cations (requires sequence information) - Owczarzy2008: Updated comprehensive salt correction (requires sequence information)

Author(s)

Junhui Li

References

Schildkraut C, Lifson S. Dependence of the melting temperature of DNA on salt concentration. Biopolymers. 1965;3(2):195-208.

Wetmur JG. DNA Probes: Applications of the Principles of Nucleic Acid Hybridization. Critical Reviews in Biochemistry and Molecular Biology. 1991;26(3-4):227-259.

SantaLucia J. A unified view of polymer, dumbbell, and oligonucleotide DNA nearest-neighbor thermodynamics. Proceedings of the National Academy of Sciences. 1998;95(4):1460-1465.

Owczarzy R, Moreira BG, Manthey JA, et al. Predicting stability of DNA duplexes in solutions containing magnesium and monovalent cations. Biochemistry. 2008;47(19):5336-5353.

Examples

salt_correction(Na = 50, Mg = 1.5, method = "Owczarzy2008", 
               input_seq = "ATGCGATGCG")

Thermodynamic parameters for GC-based Tm calculation methods

Description

A data frame containing coefficients and parameters for different GC-based Tm calculation methods. Each row represents a different method with its specific coefficients (A, B, C, D) and salt correction method.

Usage

thermodynamic_gc_params

Format

A data frame with 8 rows and 5 columns:

A

Intercept coefficient

B

GC content coefficient

C

Length correction coefficient

D

Mismatch coefficient

salt_correction

Associated salt correction method

Details

The methods included are: - Chester1993: Tm = 69.3 + 0.41(Percentage_GC) - 650/N - QuikChange: Tm = 81.5 + 0.41(Percentage_GC) - 675/N - Percentage_mismatch - Schildkraut1965: Tm = 81.5 + 0.41(Percentage_GC) - 675/N + 16.6 x log10[Na+] - Wetmur1991_MELTING: Tm = 81.5 + 0.41(Percentage_GC) - 500/N + Wetmur salt - - Wetmur1991_RNA: Tm = 78 + 0.7(Percentage_GC) - 500/N + Wetmur salt - - Wetmur1991_RNA/DNA: Tm = 67 + 0.8(Percentage_GC) - 500/N + Wetmur salt - - Primer3Plus: Tm = 81.5 + 0.41(Percentage_GC) - 600/N + 16.6 x log10[Na+] - vonAhsen2001: Tm = 77.1 + 0.41(Percentage_GC) - 528/N + 11.7 x log10[Na+]

Thermodynamic Tables for Nucleic Acid Hybridization

Description

A comprehensive collection of thermodynamic parameters used for calculating melting temperatures of nucleic acid duplexes. The dataset includes parameters for DNA/DNA, RNA/RNA, RNA/DNA and 2'-O-methylRNA/RNA hybridizations, plus parameters for mismatches, dangling ends, internal / bulge loops, GU wobble, CNG repeats, inosine, hydroxyadenine, azobenzene and locked nucleic acids (LNA).

Usage

thermodynamic_nn_params

Format

A named list of two-column matrices with colnames = c("left", "right") (left = \Delta H, right = \Delta S). Rownames are dinucleotide step keys such as "AT/TA", mismatch / dangling-end keys, or feature-specific keys (e.g. "CAG_4" for the (CAG)4 CNG entry).

Populated tables

DNA_NN_Breslauer_1986

DNA/DNA NN, Breslauer et al. (1986)

DNA_NN_Sugimoto_1996

DNA/DNA NN, Sugimoto et al. (1996)

DNA_NN_Allawi_1998

DNA/DNA NN, Allawi (1998)

DNA_NN_SantaLucia_2004

DNA/DNA NN, SantaLucia & Hicks (2004)

RNA_NN_Freier_1986

RNA/RNA NN, Freier (1986)

RNA_NN_Xia_1998

RNA/RNA NN, Xia (1998)

RNA_NN_Chen_2012

RNA/RNA NN with GU, Chen / Serra (2012)

RNA_DNA_NN_Sugimoto_1995

RNA/DNA hybrid NN, Sugimoto (1995)

DNA_IMM_Peyret_1999

DNA single internal mismatch, Peyret (1999)

DNA_TMM_Bommarito_2000

DNA terminal mismatch, Bommarito (2000)

DNA_DE_Bommarito_2000

DNA single dangling end, Bommarito (2000)

RNA_DE_Turner_2010

RNA single dangling end, Turner (2010)

Registered placeholders (values TBD - populate from primary literature)

DNA_NN_AllSan_1997

Allawi & SantaLucia (1997) Biochemistry 36:10581

DNA_NN_SantaLucia_1996

SantaLucia et al. (1996) Biochemistry 35:3555

DNA_NN_Tanaka_2004

Tanaka et al. (2004) Biochemistry 43:7143

MeRNA_RNA_NN_Kierzek_2006

Kierzek et al. (2006) Biochemistry 45:581

DNA_RNA_IMM_Watkins_2011

Watkins et al. (2011) Nucleic Acids Res 39:1894

RNA_IMM_Lu_2006

Lu et al. (2006) Nucleic Acids Res 34:4912

RNA_IMM_Davis_Znosko_2007

Davis & Znosko (2007) Biochemistry 46:13425

RNA_IMM_Davis_Znosko_2008

Davis & Znosko (2008) Biochemistry 47:10178

DNA_TandemMM_AllSanPey

Allawi & SantaLucia (1997/1998); Peyret (1999)

RNA_TandemMM_Mathews_1999

Mathews et al. (1999) JMB 288:911

DNA_DE_Ohmichi_2002

Ohmichi et al. (2002) JACS 124:10367

RNA_DE_Ohmichi_2002

Ohmichi et al. (2002) JACS 124:10367

RNA_DE_Serra_2008

O'Toole / Serra (2006, 2008)

DNA_DDE_Ohmichi_2002

Ohmichi et al. (2002) JACS 124:10367

RNA_DDE_Ohmichi_2002

Ohmichi et al. (2002) JACS 124:10367

RNA_DDE_OToole_2005

O'Toole et al. (2005) Biochemistry 44:14914

RNA_DDE_OToole_2006

O'Toole et al. (2006) NAR 34:3338

DNA_LDE_Ohmichi_2002

Ohmichi et al. (2002) JACS 124:10367

RNA_LDE_Ohmichi_2002

Ohmichi et al. (2002) JACS 124:10367

DNA_IL_SantaLucia_2004

SantaLucia & Hicks (2004)

RNA_IL_Lu_2006

Lu et al. (2006) NAR 34:4912

RNA_IL_Badhwar_2007

Badhwar et al. (2007) Biochemistry 46:14715

DNA_SBL_Tanaka_2007

Tan & Chen (2007) Biophys J 92:3615

DNA_SBL_SantaLucia_2004

SantaLucia & Hicks (2004)

RNA_SBL_Lu_2006

Lu et al. (2006) NAR 34:4912

RNA_SBL_Blose_2007

Blose et al. (2007) Biochemistry 46:15123

DNA_LBL_SantaLucia_2004

SantaLucia & Hicks (2004)

RNA_LBL_Lu_2006

Mathews (1999); Lu et al. (2006)

RNA_GU_Mathews_1999

Mathews & Turner (1999) JMB 288:911

RNA_GU_Chen_2012

Chen / Serra (2012)

DNA_CNG_Broda_2005

Broda et al. (2005) Biochemistry 44:13851 - rownames use paste0("C", N, "G_", n), e.g. "CAG_4".

DNA_INO_SantaLucia_2005

Watkins & SantaLucia (2005) NAR 33:6258

RNA_INO_Wright_2007

Wright et al. (2007) Biochemistry 46:4625

DNA_HA_Kawakami_2001

Kawakami / Sugimoto (2001) Biochemistry 40:14040

DNA_AZB_Asanuma_2005

Asanuma et al. (2005) Angew Chem Int Ed 44:2747

DNA_LNA_Owczarzy_2011

Owczarzy et al. (2011) Biochemistry 50:9352

DNA_LNA_McTigue_2004

McTigue et al. (2004) Biochemistry 43:5388

DNA_cLNA_Owczarzy_2011

Owczarzy et al. (2011) Biochemistry 50:9352

DNA_cLNA_MM_Owczarzy_2011

Owczarzy et al. (2011) Biochemistry 50:9352

To add the numeric values for any placeholder table, construct a matrix with two columns (\Delta H kcal/mol, \Delta S cal/(mol*K)) and the dinucleotide-step rownames described above, then assign it:

thermodynamic_nn_params$DNA_CNG_Broda_2005 <- new_table

Details

Coverage mirrors the rmelting (Bioconductor) vignette sections 4.2.1 - 4.4. Tables whose numeric values are not yet ported from primary literature are included in the list as NULL placeholders so that tm_nn can still dispatch on the table name; the corresponding contribution is silently skipped after a one-time package-startup notice.

Source

Various publications as cited above.

References

Breslauer K J (1986) <doi:10.1073/pnas.83.11.3746> Sugimoto N (1996) <doi:10.1093/nar/24.22.4501> Allawi H (1998) <doi:10.1093/nar/26.11.2694> SantaLucia J (2004) <doi:10.1146/annurev.biophys.32.110601.141800> Freier S (1986) <doi:10.1073/pnas.83.24.9373> Xia T (1998) <doi:10.1021/bi9809425> Chen JL (2012) <doi:10.1021/bi3002709> Sugimoto N (1995) <doi:10.1016/S0048-9697(98)00088-6> Bommarito S (2000) <doi:10.1093/nar/28.9.1929> Peyret N (1999) <doi:10.1021/bi9825091> Allawi H T & SantaLucia J (1997) <doi:10.1021/bi962590c> SantaLucia J (2005) <doi:10.1093/nar/gki918> Turner D H (2010) <doi:10.1093/nar/gkp892> Tanaka F (2004) Biochemistry 43:7143 Kierzek E (2006) Biochemistry 45:581 Watkins N E (2011) Nucleic Acids Res 39:1894 Lu Z J (2006) Nucleic Acids Res 34:4912 Davis A R & Znosko B M (2007, 2008) Biochemistry Mathews D H (1999) <doi:10.1006/jmbi.1999.2700> Ohmichi T (2002) J Am Chem Soc 124:10367 O'Toole A S (2005, 2006) Biochemistry / Nucleic Acids Res Badhwar J (2007) Biochemistry 46:14715 Tan Z J & Chen S J (2007) Biophys J 92:3615 Blose J M (2007) Biochemistry 46:15123 Broda M (2005) <doi:10.1021/bi0501447> Wright D J (2007) Biochemistry 46:4625 Kawakami J / Sugimoto N (2001) <doi:10.1021/bi010918b> Asanuma H (2005) Angew Chem Int Ed 44:2747 McTigue P M (2004) Biochemistry 43:5388 Owczarzy R (2011) Biochemistry 50:9352

Examples

# Access DNA/DNA nearest neighbor parameters
thermodynamic_nn_params$DNA_NN_SantaLucia_2004

# Access DNA internal mismatch parameters
thermodynamic_nn_params$DNA_IMM_Peyret_1999

# See which tables are still placeholders (NULL)
names(Filter(is.null, thermodynamic_nn_params))

Calculate melting temperature using multiple methods

Description

Calculates melting temperature using multiple methods: - Nearest Neighbor thermodynamics (tm_nn) - GC content-based method (tm_gc) - Wallace rule (tm_wallace)

Usage

tm_calculate(
  input_seq,
  method = c("tm_nn", "tm_gc", "tm_wallace"),
  complement_seq = NULL,
  ambiguous = FALSE,
  shift = 0,
  nn_table = c("DNA_NN_SantaLucia_2004", "DNA_NN_Breslauer_1986", "DNA_NN_Sugimoto_1996",
    "DNA_NN_Allawi_1998", "RNA_NN_Freier_1986", "RNA_NN_Xia_1998", "RNA_NN_Chen_2012",
    "RNA_DNA_NN_Sugimoto_1995"),
  tmm_table = "DNA_TMM_Bommarito_2000",
  imm_table = "DNA_IMM_Peyret_1999",
  de_table = c("DNA_DE_Bommarito_2000", "RNA_DE_Turner_2010"),
  dnac_high = 25,
  dnac_low = 25,
  self_comp = FALSE,
  variant = c("Primer3Plus", "Chester1993", "QuikChange", "Schildkraut1965",
    "Wetmur1991_MELTING", "Wetmur1991_RNA", "Wetmur1991_RNA/DNA", "vonAhsen2001"),
  userset = NULL,
  Na = 50,
  K = 0,
  Tris = 0,
  Mg = 0,
  dNTPs = 0,
  salt_method = c("Schildkraut2010", "Wetmur1991", "SantaLucia1996", "SantaLucia1998-1",
    "Owczarzy2004", "Owczarzy2008"),
  DMSO = 0,
  formamide_unit = list(value = 0, unit = "percent"),
  dmso_factor = 0.75,
  formamide_factor = 0.65,
  mismatch = TRUE
)

Arguments

Details

The function calculates melting temperature using the specified method(s). For each method: - NN: Uses nearest neighbor thermodynamics with detailed sequence analysis - GC: Uses GC content-based calculation with various empirical formulas - Wallace: Uses the simple Wallace rule (2deg C per A/T, 4deg C per G/C)

The function processes the input sequence once and applies it to all selected methods, making it more efficient than calling each method separately.

Value

A TmCalculator list with:

Available Options

Method Selection:

• method: c("tm_nn", "tm_gc", "tm_wallace")

Nearest Neighbor (NN) Method Options:

• nn_table:

  • "DNA_NN_Breslauer_1986"

  • "DNA_NN_Sugimoto_1996"

  • "DNA_NN_Allawi_1998"

  • "DNA_NN_SantaLucia_2004" (default)

  • "RNA_NN_Freier_1986"

  • "RNA_NN_Xia_1998"

  • "RNA_NN_Chen_2012"

  • "RNA_DNA_NN_Sugimoto_1995"

• tmm_table (Terminal Mismatches):

  • "DNA_TMM_Bommarito_2000" (default)

• imm_table (Internal Mismatches):

  • "DNA_IMM_Peyret_1999" (default)

• de_table (Dangling Ends):

  • "DNA_DE_Bommarito_2000" (default)

  • "RNA_DE_Turner_2010"

GC Method Options:

• variant:

  • "Primer3Plus" (default)

  • "Chester1993"

  • "QuikChange"

  • "Schildkraut1965"

  • "Wetmur1991_MELTING"

  • "Wetmur1991_RNA"

  • "Wetmur1991_RNA/DNA"

  • "vonAhsen2001"

Salt Correction Options:

• salt_method:

  • "Schildkraut2010" (default)

  • "Wetmur1991"

  • "SantaLucia1996"

  • "SantaLucia1998-1"

  • "SantaLucia1998-2"

  • "Owczarzy2004"

  • "Owczarzy2008"

Formamide Unit Options:

• formamide_unit$unit:

  • "percent" (default)

  • "molar"

Other Parameters:

• ambiguous: TRUE/FALSE (default: FALSE)

• shift: Integer value (default: 0)

• dnac_high: Numeric value in nM (default: 25)

• dnac_low: Numeric value in nM (default: 25)

• self_comp: TRUE/FALSE (default: FALSE)

• Na: Millimolar concentration (default: 50)

• K: Millimolar concentration (default: 0)

• Tris: Millimolar concentration (default: 0)

• Mg: Millimolar concentration (default: 0)

• dNTPs: Millimolar concentration (default: 0)

• DMSO: Percent concentration (default: 0)

• dmso_factor: Numeric value (default: 0.75)

• formamide_factor: Numeric value (default: 0.65)

• mismatch: TRUE/FALSE (default: TRUE)

Author(s)

Junhui Li

Examples

## Not run: 
input_seq <- c("chr1:1000100-1000150:+:BSgenome.Hsapiens.UCSC.hg38")
result <- tm_calculate(
  input_seq,
  method = "tm_nn",
  nn_table = "DNA_NN_SantaLucia_2004",
  salt_method = "Owczarzy2008"
)

## End(Not run)

Calculate the melting temperature using empirical formulas based on GC content

Description

Calculate the melting temperature using empirical formulas based on GC content with different options. The function returns a list of sequences with updated Tm attributes and calculation options.

Usage

tm_gc(
  gr_seq,
  ambiguous = FALSE,
  userset = NULL,
  variant = c("Primer3Plus", "Chester1993", "QuikChange", "Schildkraut1965",
    "Wetmur1991_MELTING", "Wetmur1991_RNA", "Wetmur1991_RNA/DNA", "vonAhsen2001"),
  Na = 50,
  K = 0,
  Tris = 0,
  Mg = 0,
  dNTPs = 0,
  salt_method = c("Schildkraut2010", "Wetmur1991", "SantaLucia1996", "SantaLucia1998-1",
    "Owczarzy2004", "Owczarzy2008"),
  mismatch = TRUE,
  DMSO = 0,
  formamide_unit = list(value = 0, unit = "percent"),
  dmso_factor = 0.75,
  formamide_factor = 0.65
)

Arguments

Value

Returns a list with two components: - Tm: A list of sequences with updated Tm attributes - Options: A list containing calculation parameters and method information

Author(s)

Junhui Li

References

Marmur J, Doty P. Determination of the base composition of deoxyribonucleic acid from its thermal denaturation temperature. Journal of Molecular Biology, 1962, 5(1):109-118.

Schildkraut C. Dependence of the melting temperature of DNA on salt concentration. Biopolymers, 2010, 3(2):195-208.

Wetmur JG. DNA Probes: Applications of the Principles of Nucleic Acid Hybridization. CRC Critical Reviews in Biochemistry, 1991, 26(3-4):33.

Untergasser A, Cutcutache I, Koressaar T, et al. Primer3–new capabilities and interfaces. Nucleic Acids Research, 2012, 40(15):e115-e115.

von Ahsen N, Wittwer CT, Schutz E, et al. Oligonucleotide melting temperatures under PCR conditions: deoxynucleotide Triphosphate and Dimethyl sulfoxide concentrations with comparison to alternative empirical formulas. Clin Chem 2001, 47:1956-1961.

Examples

# Example with multiple sequences
input_seq <- c("ATCGTGCGTAGCAGTACGATCAGTAG", "ATCGTGCGTAGCAGTACGATCAGTAG")
gr_seq <- to_genomic_ranges(input_seq)
out <- tm_gc(gr_seq, ambiguous = TRUE, variant = "Primer3Plus", Na = 50, mismatch = TRUE)
out
out$Options

Calculate melting temperature using nearest neighbor thermodynamics

Description

Calculate melting temperature using nearest neighbor thermodynamics. The function checks if all sequence combinations in the input sequence are present in the thermodynamic parameter tables before performing calculations.

Usage

tm_nn(
  gr_seq,
  ambiguous = FALSE,
  shift = 0,
  nn_table = c("DNA_NN_SantaLucia_2004", "DNA_NN_Breslauer_1986", "DNA_NN_Sugimoto_1996",
    "DNA_NN_Allawi_1998", "RNA_NN_Freier_1986", "RNA_NN_Xia_1998", "RNA_NN_Chen_2012",
    "RNA_DNA_NN_Sugimoto_1995"),
  tmm_table = "DNA_TMM_Bommarito_2000",
  imm_table = "DNA_IMM_Peyret_1999",
  de_table = c("DNA_DE_Bommarito_2000", "RNA_DE_Turner_2010"),
  dnac_high = 25,
  dnac_low = 25,
  self_comp = FALSE,
  Na = 50,
  K = 0,
  Tris = 0,
  Mg = 0,
  dNTPs = 0,
  salt_method = c("Schildkraut2010", "Wetmur1991", "SantaLucia1996", "SantaLucia1998-1",
    "Owczarzy2004", "Owczarzy2008"),
  DMSO = 0,
  formamide_unit = list(value = 0, unit = "percent"),
  dmso_factor = 0.75,
  formamide_factor = 0.65
)

Arguments

Details

DNA_NN_Breslauer_1986: Breslauer K J (1986) <doi:10.1073/pnas.83.11.3746>

DNA_NN_Sugimoto_1996: Sugimoto N (1996) <doi:10.1093/nar/24.22.4501>

DNA_NN_Allawi_1998: Allawi H (1998) <doi:10.1093/nar/26.11.2694>

DNA_NN_SantaLucia_2004: SantaLucia J (2004) <doi:10.1146/annurev.biophys.32.110601.141800>

RNA_NN_Freier_1986: Freier S (1986) <doi:10.1073/pnas.83.24.9373>

RNA_NN_Xia_1998: Xia T (1998) <doi:10.1021/bi9809425>

RNA_NN_Chen_2012: Chen JL (2012) <doi:10.1021/bi3002709>

RNA_DNA_NN_Sugimoto_1995: Sugimoto N (1995)<doi:10.1016/S0048-9697(98)00088-6>

DNA_TMM_Bommarito_2000: Bommarito S (2000) <doi:10.1093/nar/28.9.1929>

DNA_IMM_Peyret_1999: Peyret N (1999) <doi:10.1021/bi9825091> & Allawi H T (1997) <doi:10.1021/bi962590c> & Santalucia N (2005) <doi:10.1093/nar/gki918>

DNA_DE_Bommarito_2000: Bommarito S (2000) <doi:10.1093/nar/28.9.1929>

RNA_DE_Turner_2010: Turner D H (2010) <doi:10.1093/nar/gkp892>

Author(s)

Junhui Li

References

Breslauer K J , Frank R , Blocker H , et al. Predicting DNA duplex stability from the base sequence.[J]. Proceedings of the National Academy of Sciences, 1986, 83(11):3746-3750.

Sugimoto N , Nakano S , Yoneyama M , et al. Improved Thermodynamic Parameters and Helix Initiation Factor to Predict Stability of DNA Duplexes[J]. Nucleic Acids Research, 1996, 24(22):4501-5.

Allawi, H. Thermodynamics of internal C.T mismatches in DNA[J]. Nucleic Acids Research, 1998, 26(11):2694-2701.

Hicks L D , Santalucia J . The thermodynamics of DNA structural motifs.[J]. Annual Review of Biophysics & Biomolecular Structure, 2004, 33(1):415-440.

Freier S M , Kierzek R , Jaeger J A , et al. Improved free-energy parameters for predictions of RNA duplex stability.[J]. Proceedings of the National Academy of Sciences, 1986, 83(24):9373-9377.

Xia T , Santalucia , J , Burkard M E , et al. Thermodynamic Parameters for an Expanded Nearest-Neighbor Model for Formation of RNA Duplexes with Watson-Crick Base Pairs,[J]. Biochemistry, 1998, 37(42):14719-14735.

Chen J L , Dishler A L , Kennedy S D , et al. Testing the Nearest Neighbor Model for Canonical RNA Base Pairs: Revision of GU Parameters[J]. Biochemistry, 2012, 51(16):3508-3522.

Bommarito S, Peyret N, Jr S L. Thermodynamic parameters for DNA sequences with dangling ends[J]. Nucleic Acids Research, 2000, 28(9):1929-1934.

Turner D H , Mathews D H . NNDB: the nearest neighbor parameter database for predicting stability of nucleic acid secondary structure[J]. Nucleic Acids Research, 2010, 38(Database issue):D280-D282.

Sugimoto N , Nakano S I , Katoh M , et al. Thermodynamic Parameters To Predict Stability of RNA/DNA Hybrid Duplexes[J]. Biochemistry, 1995, 34(35):11211-11216.

Allawi H, SantaLucia J: Thermodynamics and NMR of internal G-T mismatches in DNA. Biochemistry 1997, 36:10581-10594.

Santalucia N E W J . Nearest-neighbor thermodynamics of deoxyinosine pairs in DNA duplexes[J]. Nucleic Acids Research, 2005, 33(19):6258-67.

Peyret N , Seneviratne P A , Allawi H T , et al. Nearest-Neighbor Thermodynamics and NMR of DNA Sequences with Internal A-A, C-C, G-G, and T-T Mismatches, [J]. Biochemistry, 1999, 38(12):3468-3477.

Examples

input_seq <- c("AAAATTTTTTTCCCCCCCCCCCCCCGGGGGGGGGGGGTGTGCGCTGC",
"AAAATTTTTTTCCCCCCCCCCCCCCGGGGGGGGGGGGTGTGCGCTGC")
seqs <- to_genomic_ranges(input_seq)
out <- tm_nn(seqs, Na=50)
out

Calculate the melting temperature using the 'Wallace rule'

Description

The Wallace rule is often used as rule of thumb for approximate melting temperature calculations for primers with 14 to 20 nt length.

Usage

tm_wallace(gr_seq, ambiguous = FALSE)

Arguments

Value

Returns a list of sequences with updated Tm attributes

Author(s)

Junhui Li

References

Thein S L , Lynch J R , Weatherall D J , et al. DIRECT DETECTION OF HAEMOGLOBIN E WITH SYNTHETIC OLIGONUCLEOTIDES[J]. The Lancet, 1986, 327(8472):93.

Examples

input_seq = c('acgtTGCAATGCCGTAWSDBSY','acgtTGCCCCGGCCGCGCCGTAWSDBSY') #for wallace rule
gr_seq <- to_genomic_ranges(input_seq)
out <- tm_wallace(gr_seq, ambiguous = TRUE)
out
out$Options

Convert input sequences to a GRanges object (fast backend)

Description

Drop-in replacement for to_genomic_ranges that uses the fast coordinate backend in coor_to_genomic_ranges for genomic coordinate input. Accepts the same input_seq and complement_seq arguments as to_genomic_ranges, plus a list input list(pkg_name = ..., seq = ...) for large tiling jobs.

This function processes a vector of sequences string, a FASTA file, or a character vector with genomic coordinates into a GenomicRanges object, optionally including complementary sequences. sequence names are parsed based on their format: - If names have this pattern "chr:start-end:strand:species[:name]" (e.g., "chr1:1-5:+:seq_1"), parse components into seqnames, ranges, strand, and name. - If names have this pattern "chr:start-end:strand" (e.g., "chr1:1-5:+"), parse components into seqnames, ranges, and strand. - If names have this pattern "chr:start-end" (e.g., "chr1:1-5"), parse components into seqnames and ranges. - If no names are provided, use default values: seqnames = "chr1", start = 1, width = sequence length, strand = "*", name = "1", etc. Complementary sequences are either provided or automatically generated.

Usage

to_genomic_ranges_fast(
  input_seq,
  complement_seq = NULL,
  method = c("vectorized", "preload_chr")
)

to_genomic_ranges(input_seq, complement_seq = NULL)

Arguments

Value

A GRanges object. See coor_to_genomic_ranges for metadata columns when coordinate input is used.

A GenomicRanges object with seqnames, ranges, strand, name, sequence, Complement, and Tm as metadata.

Author(s)

Junhui Li

Examples

## Not run: 
gr <- to_genomic_ranges_fast(
  list(
    pkg_name = "BSgenome.Hsapiens.UCSC.hg38",
    seq = c("chr1:1000-1199:+:win1", "chr1:1200-1399:+:win2")
  )
)

## End(Not run)

# Using a character vector with auto-generated complementary sequences
seqs <- c("ATGCG", "GCTAG")
names(seqs) <- c("chr1:1-5:+:seq_1", "chr2:1-5:+")
gr <- to_genomic_ranges(seqs)
gr

# Using a character vector with provided complementary sequences
seqs <- c("ATGCG", "GCTAG")
comp_seqs <- c("TACGC", "CGTA")
gr <- to_genomic_ranges(seqs, comp_seqs)
gr

# Using a FASTA file
gr <- to_genomic_ranges(system.file("extdata", "example1.fasta", package = "TmCalculator"))
## Not run: 
# Using a character vector with genomic coordinates
seqs <- c(
  "chr1:1898000-1898050:+:BSgenome.Hsapiens.UCSC.hg38",
  "chr2:2563000-2563050:-:BSgenome.Hsapiens.UCSC.hg38"
)
gr <- to_genomic_ranges(seqs)
gr

## End(Not run)

Convert sequence strings to GenomicRanges object

Description

This function converts sequence strings to a GenomicRanges object, handling both named and unnamed sequences. It can also process complementary sequences if provided. sequence names can be in the format ">chr2:1-10:+:seq2" which will be parsed into chromosome, position, strand, and name components.

Usage

vec_to_genomic_ranges(input_seq)

Arguments

Value

A GenomicRanges object containing: - GRanges information (seqnames, ranges, strand) - sequence data - Complementary sequences - Names from input or auto-generated

Examples

# Example with named sequences in GRanges format
seqs <- c("ATGCG", "GCTAG")
names(seqs) <- c("chr1:1111-1115:+:seq1", "chr2:1221-1225:+")
gr <- vec_to_genomic_ranges(seqs)

# Example with unnamed sequences
seqs <- c("ATGCG", "GCTAG")
gr <- vec_to_genomic_ranges(seqs)