Drybench Plasmid.
pUC19 plasmid map: restriction and primer sites
Restriction sites and unique cutters on pUC19, and the universal primers that flank its cloning site, from NCBI's record with sources shown.
NCBI L09137.2, “Cloning vector pUC19c, complete sequence”, 2,686 bp, circular. Shipped with the app: opening it sends nothing to NCBI.
NCBI titles this record 'pUC19c': its comment explains that the sequence is written with the beta-galactosidase (lacZ) gene on the complementary strand. The vector is pUC19.
The map
Library features, the 10 enzymes of the cloning site that cut once, and the sequencing-primer sites. Pick a feature for its evidence.
Arrow keys move between features in map order; Enter says what the one in focus is; [ and ] step back and forward; Home and End go to the first and last.
lacZα, which carries the cloning site and is the basis of the blue-white screen, is not drawn: we have found no NCBI record whose authors annotate pUC19's own lacZα, so the annotation library has no entry for it.
Unique cutters
42 enzymes of the table cut this plasmid once (Host: dam⁺ dcm⁺).
AatII, Acc65I, AccI, AflIII, AhdI, AlwNI, ApoI, AvaI, BamHI, BanII, BpmI, BsaI, BseYI, BspMI, BsrFI, BstAPI, Eco53kI, EcoO109I, EcoRI, HincII, HindIII, KasI, KpnI, NarI, NdeI, NmeAIII, PciI, PluTI, PstI, SacI, SalI, SapI, SbfI, ScaI, SfoI, SmaI, SphI, SspI, XbaI, XmaI, XmnI, ZraI
Depends on methylation (not recorded)
Enzymes with a site whose methylation sensitivity is not recorded: they may or may not cut, so they are not counted.
BsaJI, BsiEI, CviKI-1, Hpy188I, Hpy188III, MwoI, NlaIV, PspGI, StyD4I
Features
| Feature | Location | Strand | Evidence |
|---|---|---|---|
| M13mp19 polylinker | complement(396..452) | reverse | Exact match |
| lac promoter | complement(470..591) | reverse | Exact match |
| lac operator | complement(487..507) | reverse | Exact match |
| CAP binding site | complement(554..591) | reverse | Exact match |
| RNA I (ColE1-type ori) | 1309..1415 | forward | Match: 99.0% identity, 100.0% coverage, 1 mismatch, 0 gaps |
| AmpR (bla) | complement(1626..2486) | reverse | Match: 99.7% identity, 100.0% coverage, 2 mismatches, 0 gaps |
Sequencing-primer sites
| Primer | Location | Strand | Binding |
|---|---|---|---|
| pUC/M13 Forward Primer | 352 to 375 | forward | Exact |
| M13 Forward (−20) | 379 to 394 | forward | Exact |
| M13 Reverse | 465 to 481 | reverse | Exact |
| pUC/M13 Reverse Primer | 465 to 486 | reverse | Exact |
Whether the primer primes at this site is not assessed.
Numbers
- Double-stranded weight: 1.65969 × 10⁶ g/mol (circular, no end groups).
- 1 µg is 0.6025 pmol.
- 1 ng is 3.628 × 10⁸ copies.
Next: pick a diagnostic digest
The digest picker finds the digests whose gels tell a correct construct from an empty or reversed one.
Computed when this page was built, from data release f1.e1.p1.
How it is computed
- Feature annotation from the library. Local alignment with affine gaps (Smith TF, Waterman MS. J Mol Biol 1981;147:195-197; Gotoh O. J Mol Biol 1982;162:705-708), seeded by shared 12-base words and banded; match +2, mismatch -3, gap open -5, gap extend -2, this app's parameters, checked against the Python reference; a gap of k bases scores -5 - 2(k - 1), so one long gap costs less than several short ones. Identity is identical columns over alignment columns, gaps included; coverage is the feature's aligned bases over its length. The exported local_alignment_score is this score, for re-deriving a call, not a quality measure; for a feature under 30 bases no alignment runs and the score is set to twice its length, the score a whole exact alignment would have.Checked against Biopython 1.88 PairwiseAligner with the same scoring on 26 alignments of library parts to other NCBI records: identity and coverage within 0.1 percentage points, place, strand, call and score exact; 12 synthetic inserts on each side of every threshold; no match and 0 partial calls on 10 random sequences of 10,000 bp and both golden plasmids reversed (2026-10-06). Library 2 (2026-10-17): all 4,005 pairs of its parts aligned against each other with the same scoring, 66 of them reaching the thresholds and each published with how it is shown; no call on 12 random controls; what is shown at a locus, the parts named beside it and the inside labels exact on 14 sequences.
- Restriction sites and cut positions. Recognition sequences and cut positions as published facts, typed by hand (pipeline/plasmid-map/enzymes.csv); every row checked against Biopython 1.88 Bio.Restriction, whose data are REBASE release 404 (Roberts RJ et al. REBASE: a database for DNA restriction and modification. Nucleic Acids Res 2023;51:D629-D630).Every row checked against Biopython 1.88 Bio.Restriction (enzyme data release 404) in its own notation, overhang and cut positions: 223 enzymes on pUC19 and pBR322, 446 cases, cut positions exact, with sites across the origin and 6 double digests (2026-10-06).
- dam and dcm methylation of sites. McClelland M, Nelson M, Raschke E. Effect of site-specific modification on restriction endonucleases and DNA modification methyltransferases. Nucleic Acids Res 1994;22:3640-3659, Table I.Checked against 19 cases worked by hand from McClelland et al. 1994, Table I (one per outcome, flank and origin overlaps, fully methylated DpnI and host settings, plus hemi-site sensitivity entries for other enzymes; not a hemimethylated-DpnI model), and the cited blocked examples on pUC19 (MboI) and pBR322 (EcoRII), outcomes exact (2026-10-06). Which enzymes need a Table I row (82 whose site can carry a dam or dcm methylated base, 57 of them with a row) checked by a second implementation sharing no code with the engine (2026-10-17); whether Table I lists the other 25 has not been checked.
- Sequencing-primer sites. Exact and one-mismatch matches of the shipped primers on both strands; a mismatch is reported with its distance from the primer's 3′ end, and nothing is claimed about whether such a site primes.Each of the 6 shipped primers found exactly at its NCBI binding site at build and every week; one-mismatch places unit-tested (2026-10-06).
- ng, pmol, copies and ligation amounts. pmol = ng × 1000 / MW; copies = ng × 10⁻⁹ × N_A / MW, with N_A = 6.02214076 × 10²³ per mole (exact, SI 2019); insert ng = vector ng × insert bp / vector bp × ratio. MW of the double-stranded plasmid from its own sequence (both strands, no end groups when circular) with the CIAAW 2021 atomic weights.Checked against exact rational arithmetic on 11 cases (pmol, copies per µL, ng for copies, ligation, the 650 Da per bp rule and the weights of pUC19 and pBR322), to full double precision (2026-10-06).
Limits
- For research and education only, and not medical advice: plasmid maps and sequence assembly designs. A designed product is not evidence that the reaction occurred; nothing here predicts expression or how well a reaction will work.
- Enzyme sites and cut positions are from a hand-curated table of 223 enzymes (version 1, built 2026-10-06); it may differ from a vendor's list, so check NEBcutter before a critical digest. Star activity, Type IIB and nicking enzymes are not modelled.
- Methylation blocking is called only where McClelland et al. (1994) record it; elsewhere a site reads 'may be blocked: sensitivity not recorded', which means unknown, never 'cuts'. 25 of the 82 enzymes whose site can carry a dam or dcm methylated base have no row in this app's copy of Table I, and whether Table I lists them has not been checked.
- Feature badges are alignment matches to public records: evidence of sequence, not of function.
- Primer sites are exact and one-mismatch sites of the shipped primers; whether a primer primes at a mismatched site is not assessed, and a Tm is given only against a perfect match.