LOCUS       sequence-380752-        5266 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5266)
  AUTHORS   Bertrand E, Chartrand P, Schaefer M, Shenoy SM, Singer RH, Long RM
  TITLE     Localization of ASH1 mRNA particles in living yeast.
  JOURNAL   Mol Cell. 1998 Oct . 2(4):437-45.
  PUBMED    9809065
REFERENCE   2  (bases 1 to 5266)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Mol Cell. 
            1998 Oct . 2(4):437-45."
COMMENT     Sequence Label: pCR4-24XMS2SL-stable
FEATURES             Location/Qualifiers
     source          1..5266
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(231..533)
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="QFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDKV
                     SRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     complement(333..352)
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     primer_bind     656..678
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     670..687
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     671..687
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     694..713
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        694..712
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        complement(2091..2109)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2130..2146)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2130..2146)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2143..2165)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2154..2170
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(2178..2184,2185..2202,2203..2208))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2223..2244
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2361..2378)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2532..3120)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2612..2631)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             join(3244..3312,3313..4104)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(3462..3481)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     CDS             complement(4354..5148)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             complement(4354..5148)
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4465..4484)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     5075..5094
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
ORIGIN
        1 gccccagctg gcaattccgg ttcgcttgct gtccataaaa ccgcccagtc tagctatcgc
       61 catgtaagcc cactgcaagc tacctgcttt ctctttgcgc ttgcgttttc ccttgtccag
      121 atagcccagt agctgacatt catccggggt cagcaccgtt tctgcggact ggctttctac
      181 gtgaaaagga tctaggtgaa gatccttttt gataatctca tgcctgacat ttatattccc
      241 cagaacatca ggttaatggc gtttttgatg tcattttcgc ggtggctgag atcagccact
      301 tcttccccga taacggagac cggcacactg gccatatcgg tggtcatcat gcgccagctt
      361 tcatccccga tatgcaccac cgggtaaagt tcacgggaga ctttatctga cagcagacgt
      421 gcactggcca gggggatcac catccgtcgc cccggcgtgt caataatatc actctgtaca
      481 tccacaaaca gacgataacg gctctctctt ttataggtgt aaaccttaaa ctgccgtacg
      541 tataggctgc gcaactgttg ggaagggcga tcggtgcggg cctcttcgct attacgccag
      601 ctggcgaaag ggggatgtgc tgcaaggcga ttaagttggg taacgccagg gttttcccag
      661 tcacgacgtt gtaaaacgac ggccagtgaa ttgtaatacg actcactata gggcgaattg
      721 aatttagcgg ccgcgaattc gcccttcgcg ggatcctacg gtacttattg ccaagaaagc
      781 acgagcatca gccgtgcctc caggtcgaat cttcaaacga cgacgatcac gcgtcgctcc
      841 agtattccag ggttcatcag atcctacggt acttattgcc aagaaagcac gagcatcagc
      901 cgtgcctcca ggtcgaatct tcaaacgacg acgatcacgc gtcgctccag tattccaggg
      961 ttcatcagat cctacggtac ttattgccaa gaaagcacga gcatcagccg tgcctccagg
     1021 tcgaatcttc aaacgacgac gatcacgcgt cgctccagta ttccagggtt catcagatcc
     1081 tacggtactt attgccaaga aagcacgagc atcagccgtg cctccaggtc gaatcttcaa
     1141 acgacgacga tcacgcgtcg ctccagtatt ccagggttca tcagatccta cggtacttat
     1201 tgccaagaaa gcacgagcat cagccgtgcc tccaggtcga atcttcaaac gacgacgatc
     1261 acgcgtcgct ccagtattcc agggttcatc agatcctacg gtacttattg ccaagaaagc
     1321 acgagcatca gccgtgcctc caggtcgaat cttcaaacga cgacgatcac gcgtcgctcc
     1381 agtattccag ggttcatcag atcctacggt acttattgcc aagaaagcac gagcatcagc
     1441 cgtgcctcca ggtcgaatct tcaaacgacg acgatcacgc gtcgctccag tattccaggg
     1501 ttcatcagat cctacggtac ttattgccaa gaaagcacga gcatcagccg tgcctccagg
     1561 tcgaatcttc aaacgacgac gatcacgcgt cgctccagta ttccagggtt catcagatcc
     1621 tacggtactt attgccaaga aagcacgagc atcagccgtg cctccaggtc gaatcttcaa
     1681 acgacgacga tcacgcgtcg ctccagtatt ccagggttca tcagatccta cggtacttat
     1741 tgccaagaaa gcacgagcat cagccgtgcc tccaggtcga atcttcaaac gacgacgatc
     1801 acgcgtcgct ccagtattcc agggttcatc agatcctacg gtacttattg ccaagaaagc
     1861 acgagcatca gccgtgcctc caggtcgaat cttcaaacga cgacgatcac gcgtcgctcc
     1921 agtattccag ggttcatcag atcctacggt acttattgcc aagaaagcac gagcatcagc
     1981 cgtgcctcca ggtcgaatct tcaaacgacg acgatcacgc gtcgctccag tattccaggg
     2041 ttcatcagat ctcgcgaagg gcgaattcgt ttaaacctgc aggactagtc cctttagtga
     2101 gggttaattc tgagcttggc gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat
     2161 ccgctcacaa ttccacacaa catacgagcc ggaagcataa agtgtaaagc ctggggtgcc
     2221 taatgagtga gctaactcac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga
     2281 aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt
     2341 attgggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg
     2401 cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac
     2461 gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg
     2521 ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca
     2581 agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc
     2641 tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc
     2701 ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag
     2761 gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc
     2821 ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca
     2881 gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg
     2941 aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg
     3001 aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct
     3061 ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa
     3121 gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa
     3181 gggattttgg tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag
     3241 agtatgagta ttcaacattt ccgtgtcgcc cttattccct tttttgcggc attttgcctt
     3301 cctgtttttg ctcacccaga aacgctggtg aaagtaaaag atgctgaaga tcagttgggt
     3361 gcacgagtgg gttacatcga actggatctc aacagcggta agatccttga gagttttcgc
     3421 cccgaagaac gttttccaat gatgagcact tttaaagttc tgctatgtgg cgcggtatta
     3481 tcccgtattg acgccgggca agagcaactc ggtcgccgca tacactattc tcagaatgac
     3541 ttggttgagt actcaccagt cacagaaaag catcttacgg atggcatgac agtaagagaa
     3601 ttatgcagtg ctgccataac catgagtgat aacactgcgg ccaacttact tctgacaacg
     3661 atcggaggac cgaaggagct aaccgctttt ttgcacaaca tgggggatca tgtaactcgc
     3721 cttgatcgtt gggaaccgga gctgaatgaa gccataccaa acgacgagcg tgacaccacg
     3781 atgcctgtag caatggcaac aacgttgcgc aaactattaa ctggcgaact acttactcta
     3841 gcttcccggc aacaattaat agactggatg gaggcggata aagttgcagg accacttctg
     3901 cgctcggccc ttccggctgg ctggtttatt gctgataaat ctggagccgg tgagcgtggg
     3961 tctcgcggta tcattgcagc actggggcca gatggtaagc cctcccgtat cgtagttatc
     4021 tacacgacgg ggagtcaggc aactatggat gaacgaaata gacagatcgc tgagataggt
     4081 gcctcactga ttaagcattg gtaactgtca gaccaagttt actcatatat actttagatt
     4141 gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc
     4201 atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca
     4261 tttccccgaa aagtgccacc tgatgcggtg tgaaataccg cacagatgcg taaggagaaa
     4321 ataccgcatc aggaaattgt aagcgttaat aattcagaag aactcgtcaa gaaggcgata
     4381 gaaggcgatg cgctgcgaat cgggagcggc gataccgtaa agcacgagga agcggtcagc
     4441 ccattcgccg ccaagctctt cagcaatatc acgggtagcc aacgctatgt cctgatagcg
     4501 gtccgccaca cccagccggc cacagtcgat gaatccagaa aagcggccat tttccaccat
     4561 gatattcggc aagcaggcat cgccatgggt cacgacgaga tcctcgccgt cgggcatgct
     4621 cgccttgagc ctggcgaaca gttcggctgg cgcgagcccc tgatgctctt cgtccagatc
     4681 atcctgatcg acaagaccgg cttccatccg agtacgtgct cgctcgatgc gatgtttcgc
     4741 ttggtggtcg aatgggcagg tagccggatc aagcgtatgc agccgccgca ttgcatcagc
     4801 catgatggat actttctcgg caggagcaag gtgagatgac aggagatcct gccccggcac
     4861 ttcgcccaat agcagccagt cccttcccgc ttcagtgaca acgtcgagca cagctgcgca
     4921 aggaacgccc gtcgtggcca gccacgatag ccgcgctgcc tcgtcttgca gttcattcag
     4981 ggcaccggac aggtcggtct tgacaaaaag aaccgggcgc ccctgcgctg acagccggaa
     5041 cacggcggca tcagagcagc cgattgtctg ttgtgcccag tcatagccga atagcctctc
     5101 cacccaagcg gccggagaac ctgcgtgcaa tccatcttgt tcaatcatgc gaaacgatcc
     5161 tcatcctgtc tcttgatcag agcttgatcc cctgcgccat cagatccttg gcggcaagaa
     5221 agccatccag tttactttgc agggcttccc aaccttacca gagggc
//
