LOCUS       sequence-12962-e        4651 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 4651)
  AUTHORS   Zhang H, Fire AZ
  TITLE     Cell autonomous specification of temporal identity by Caenorhabditis
            elegans microRNA lin-4.
  JOURNAL   Dev Biol. 2010 Aug 15. 344(2):603-10.
  PUBMED    20493184
REFERENCE   2  (bases 1 to 4651)
  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: "Dev Biol. 
            2010 Aug 15. 344(2):603-10."
COMMENT     Sequence Label: pHZ018
FEATURES             Location/Qualifiers
     source          1..4651
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    107..128
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(143..148,149..166,167..173)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    181..197
                     /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)."
     primer_bind     186..208
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     205..221
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     205..221
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        242..260
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(1023..1042)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1024..1042)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1049..1066)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1049..1065)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1058..1080)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     CDS             1203..1505
                     /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     1384..1403
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             1854..2648
                     /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             1854..2648
                     /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(1908..1927)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2518..2537
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     CDS             complement(join(2898..3689,3690..3758))
                     /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     3521..3540
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3882..4470
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4371..4390
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4624..4641
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 agcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc
       61 acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc
      121 tcactcatta ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa
      181 ttgtgagcgg ataacaattt cacacaggaa acagctatga ccatgattac gccaagctca
      241 gaattaaccc tcactaaagg gactagtcct gcaggtttaa acgaattcgc cctttagtcg
      301 acttctgaaa ataatcgttt gacccttttc cccgaatacc attttattgg aactagctcc
      361 cagtgtgaaa aaaaaccccg tgaaaacaga aaaaaagatc tgctcaaacc gtcctggtac
      421 ccggagagcc caggtgtgaa gcatcaatag tacactcaca cttgaggtct cagggaacag
      481 gccggaagca taaactcata aaccaaccaa aactctagaa attgtctaga atctaaaaac
      541 tataagattt attatattca ggaatagttt agatatttga aaggtaagtg tgatgagtcc
      601 gaccgattgt ggtagagtct attggaaaga gtgaccgaaa gggtctctgg gagtgatggg
      661 gcggaggatg gatggacatg ggaggagtag ctgaaggagg aggatcgaaa gtagagaagt
      721 gaagagactg ggtcattcgg tcgcttcgtc gttattcttt tttgccgcgg tccccccgcg
      781 cactgagaga gaggacagac gagaagcgga aggcacacgc agcagagcac ggagggacac
      841 gagcgcggcg ccccgcaagc ctttccgtga ggctggcacc accaccgctt ggcgacgcgt
      901 ccccgaaata gtgttaggtg ccacagagac gtagaccacc tcgctcccct ctcatttttg
      961 cagtaaggga gactcggcgt ctcgtcgaca agggcgaatt cgcggccgct aaattcaatt
     1021 cgccctatag tgagtcgtat tacaattcac tggccgtcgt tttacaacgt cgtgactggg
     1081 aaaaccctgg cgttacccaa cttaatcgcc ttgcagcaca tccccctttc gccagctggc
     1141 gtaatagcga agaggcccgc accgatcgcc cttcccaaca gttgcgcagc ctatacgtac
     1201 ggcagtttaa ggtttacacc tataaaagag agagccgtta tcgtctgttt gtggatgtac
     1261 agagtgatat tattgacacg ccggggcgac ggatggtgat ccccctggcc agtgcacgtc
     1321 tgctgtcaga taaagtctcc cgtgaacttt acccggtggt gcatatcggg gatgaaagct
     1381 ggcgcatgat gaccaccgat atggccagtg tgccggtctc cgttatcggg gaagaagtgg
     1441 ctgatctcag ccaccgcgaa aatgacatca aaaacgccat taacctgatg ttctggggaa
     1501 tataaatgtc aggcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttcacgtaga
     1561 aagccagtcc gcagaaacgg tgctgacccc ggatgaatgt cagctactgg gctatctgga
     1621 caagggaaaa cgcaagcgca aagagaaagc aggtagcttg cagtgggctt acatggcgat
     1681 agctagactg ggcggtttta tggacagcaa gcgaaccgga attgccagct ggggcgccct
     1741 ctggtaaggt tgggaagccc tgcaaagtaa actggatggc tttcttgccg ccaaggatct
     1801 gatggcgcag gggatcaagc tctgatcaag agacaggatg aggatcgttt cgcatgattg
     1861 aacaagatgg attgcacgca ggttctccgg ccgcttgggt ggagaggcta ttcggctatg
     1921 actgggcaca acagacaatc ggctgctctg atgccgccgt gttccggctg tcagcgcagg
     1981 ggcgcccggt tctttttgtc aagaccgacc tgtccggtgc cctgaatgaa ctgcaagacg
     2041 aggcagcgcg gctatcgtgg ctggccacga cgggcgttcc ttgcgcagct gtgctcgacg
     2101 ttgtcactga agcgggaagg gactggctgc tattgggcga agtgccgggg caggatctcc
     2161 tgtcatctca ccttgctcct gccgagaaag tatccatcat ggctgatgca atgcggcggc
     2221 tgcatacgct tgatccggct acctgcccat tcgaccacca agcgaaacat cgcatcgagc
     2281 gagcacgtac tcggatggaa gccggtcttg tcgatcagga tgatctggac gaagagcatc
     2341 aggggctcgc gccagccgaa ctgttcgcca ggctcaaggc gagcatgccc gacggcgagg
     2401 atctcgtcgt gacccatggc gatgcctgct tgccgaatat catggtggaa aatggccgct
     2461 tttctggatt catcgactgt ggccggctgg gtgtggcgga ccgctatcag gacatagcgt
     2521 tggctacccg tgatattgct gaagagcttg gcggcgaatg ggctgaccgc ttcctcgtgc
     2581 tttacggtat cgccgctccc gattcgcagc gcatcgcctt ctatcgcctt cttgacgagt
     2641 tcttctgaat tattaacgct tacaatttcc tgatgcggta ttttctcctt acgcatctgt
     2701 gcggtatttc acaccgcatc aggtggcact tttcggggaa atgtgcgcgg aacccctatt
     2761 tgtttatttt tctaaataca ttcaaatatg tatccgctca tgagattatc aaaaaggatc
     2821 ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag
     2881 taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt
     2941 ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag
     3001 ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca
     3061 gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact
     3121 ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca
     3181 gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg
     3241 tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc
     3301 atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg
     3361 gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca
     3421 tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt
     3481 atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc
     3541 agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc
     3601 ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca
     3661 tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa
     3721 aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat
     3781 tgaagcattt atcagggtta ttgtctcatg accaaaatcc cttaacgtga gttttcgttc
     3841 cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg
     3901 cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg
     3961 gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca
     4021 aatactgttc ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg
     4081 cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg
     4141 tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga
     4201 acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac
     4261 ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat
     4321 ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc
     4381 tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga
     4441 tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc
     4501 ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg
     4561 gataaccgta ttaccgcctt tgagtgagct gataccgctc gccgcagccg aacgaccgag
     4621 cgcagcgagt cagtgagcga ggaagcggaa g
//
