Sequence Statistics

GC Content — The ratio of guanine + cytosine to total bases, expressed as a percentage. G-C base pairs form three hydrogen bonds (vs. two for A-T/A-U), so higher GC content increases thermal stability. GC content varies by genome region: coding sequences, CpG islands, and rRNA genes tend to be GC-rich.

Molecular Weight — Calculated from the sum of individual nucleotide monophosphate masses minus water released during polymerization. Essential for stoichiometric calculations, gel electrophoresis estimation, and mass spectrometry verification.

Melting Temperature (Tm) — The temperature at which half of the DNA duplexes denature. Critical for PCR primer design (annealing temperature $\approx$ Tm − 5°C), hybridization stringency, and understanding in vivo stability.

Codon Usage — The genetic code is degenerate: 61 sense codons encode 20 amino acids. Organisms exhibit codon usage bias, preferring certain synonymous codons. High-expression genes tend to use codons matching abundant tRNAs. The Codon Adaptation Index (CAI) quantifies how well a gene's codon usage matches the host organism.

Open Reading Frames (ORFs) — An ORF is a continuous stretch of codons beginning with ATG (start) and ending with a stop codon (TAA, TAG, or TGA). Three forward reading frames exist for each strand (six total for double-stranded DNA). The longest ORFs in each frame are candidates for protein-coding genes, though true gene identification requires promoter analysis, ribosome binding sites, and comparative evidence.

Reverse Complement — DNA is antiparallel: the two strands run 5'→3' and 3'→5'. The reverse complement converts between strands while maintaining the 5'→3' convention, essential for primer design and annotation.