LOCUS       sequence-66680-e        7060 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express cGFP transcript ending in a poly(A) tail [using the bGH 
            poly(A) signal].
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7060)
  AUTHORS   Wilusz JE, JnBaptiste CK, Lu LY, Kuhn CD, Joshua-Tor L, Sharp PA
  TITLE     A triple helix stabilizes the 3' ends of long noncoding RNAs that 
            lack poly(A) tails.
  JOURNAL   Genes Dev. 2012 Nov 1;26(21):2392-407. doi: 10.1101/gad.204438.112. 
            Epub 2012 Oct 16.
  PUBMED    23073843
REFERENCE   2  (bases 1 to 7060)
  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"; doi: 
            "10.1101/gad.204438.112"; journalName: "Genes Dev"; date: 
            "2012-11-1- 1"; volume: "26"; issue: "21"; pages: "2392-407"
COMMENT     Sequence Label: pCRII-TOPO CMV-cGFP-bGH Poly(A) Sense
FEATURES             Location/Qualifiers
     source          1..7060
                     /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        239..257
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     239..256
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     enhancer        1075..1453
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1454..1657
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1608..1628
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     polyA_signal    2429..2653
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(3493..3512)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3494..3512)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3519..3536)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3519..3535)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3528..3550)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3676..4104
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3763..3782)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3973..3994
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             4448..5242
                     /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
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4502..4521)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5112..5131
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     CDS             join(5260..5328,5329..6120)
                     /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(5478..5497)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6291..6879
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6780..6799
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7033..7050
                     /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 gccaagctat
      241 ttaggtgaca ctatagaata ctcaagctat gcatcaagct tctttaaaag gccacttgaa
      301 ctcgctttcc atggcgattt gccttgtgag cactttcagg agagcctgga agctgaaaaa
      361 cggtagaaaa atttccgtgc gggccgtggg gggctggcgg caactggggg gccgcagatc
      421 agagtgggcc actggcagcc aacggccccc ggggctcagg cggggagcag ctctgtggtg
      481 tgggattgag gcgttttcca agagtgggtt ttcacgtttc taagatttcc caagcagaca
      541 gcccgtgctg ctccgatttc tcgaacaaaa aagcaaaacg tgtggctgtc ttgggagcaa
      601 gtcgcaggac tgcaagcagt tgggggagaa agtccgccat tttgccactt ctcaaccgtc
      661 cctgcaaggc tggggctcag ttgcgtaatg gaaagtaaag ccctgaacta tcacacttta
      721 atcttccttc aaaaggtggt aaactatacc tactgtccct caagagaaca caagaagtgc
      781 tttaagaggt attttaaaag ttccgggggt tttgtgaggt gtttgatgac ccgtttaaaa
      841 tatgatttcc atgtttcttt tgtctaaagt ttgcagctca aatctttcca cacgctagta
      901 atttaagtat ttctgcatgt gtagtttgca ttcaagttcc ataagctgtt aagaaaaatc
      961 tagaaaagta aaactagaac ctatttttaa ccgaagaact actttttgcc tccctcacaa
     1021 aggcggcgga aggtgatcga attccggtga tgcgagttgt tctccgtcgg atccacattg
     1081 attattgact agttattaat agtaatcaat tacggggtca ttagttcata gcccatatat
     1141 ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc ccaacgaccc
     1201 ccgcccattg acgtcaataa tgacgtatgt tcccatagta acgccaatag ggactttcca
     1261 ttgacgtcaa tgggtggact atttacggta aactgcccac ttggcagtac atcaagtgta
     1321 tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg cctggcatta
     1381 tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg tattagtcat
     1441 cgctattacc atggtgatgc ggttttggca gtacatcaat gggcgtggat agcggtttga
     1501 ctcacgggga tttccaagtc tccaccccat tgacgtcaat gggagtttgt tttggcacca
     1561 aaatcaacgg gactttccaa aatgtcgtaa caactccgcc ccattgacgc aaatgggcgg
     1621 taggcgtgta cggtgggagg tctatataag cagagctctc tggctaacta gagaacccac
     1681 tgcttactgg cgatatcatg gccagcaagg gcgaggagct gttcaccggc gtggtgccca
     1741 tcctggtgga gctggacggc gatgtgaatg gccacaagtt cagcgtgagc ggcgagggcg
     1801 agggcgatgc cacctacggc aagctgaccc tgaagttcat ctgcaccacc ggcaagctgc
     1861 ctgtgccctg gcccaccctg gtgaccaccc tgagctacgg cgtgcagtgc ttctcacgct
     1921 accccgatca catgaagcag cacgacttct tcaagagcgc catgcctgag ggctacatcc
     1981 aggagcgcac catcttcttc gaggatgacg gcaactacaa gtcgcgcgcc gaggtgaagt
     2041 tcgagggcga taccctggtg aatcgcatcg agctgaccgg caccgatttc aaggaggatg
     2101 gcaacatcct gggcaataag atggagtaca actacaacgc ccacaatgtg tacatcatga
     2161 ccgacaaggc caagaatggc atcaaggtga acttcaagat ccgccacaac atcgaggatg
     2221 gcagcgtgca gctggccgac cactaccagc agaatacccc catcggcgat ggccctgtgc
     2281 tgctgcccga taaccactac ctgtccaccc agagcgccct gtccaaggac cccaacgaga
     2341 agcgcgatca catgatctac ttcggcttcg tgaccgccgc cggcatcacc cacggcatgg
     2401 acgagctgta caagtgagcg gccgccgact gtgccttcta gttgccagcc atctgttgtt
     2461 tgcccctccc ccgtgccttc cttgaccctg gaaggtgcca ctcccactgt cctttcctaa
     2521 taaaatgagg aaattgcatc gcattgtctg agtaggtgtc attctattct ggggggtggg
     2581 gtggggcagg acagcaaggg ggaggattgg gaagacaata gcaggcatgc tggggatgcg
     2641 gtgggctcta tgggcggccg cgctgtgcgg taggcattga ggcagccagc gcaggggctt
     2701 ctgctgaggg ggcaggcgga gcttgaggaa accgcagata agtttttttc tctttgaaag
     2761 atagagatta atacaactac ttaaaaaata tagtcaatag gttactaaga tattgcttag
     2821 cgttaagttt ttaacgtaat tttaatagct taagatttta agagaaaata tgaagactta
     2881 gaagagtagc atgaggaagg aaaagataaa aggtttctaa aacatgacgg aggttgagat
     2941 gaagcttctt catggagtaa aaaatgtatt taaaagaaaa ttgagagaaa ggactacaga
     3001 gccccgaatt aataccaata gaagggcaat gcttttagat taaaatgaag gtgacttaaa
     3061 cagcttaaag tttagtttaa aagttgtagg tgattaaaat aatttgaagg cgatctttta
     3121 aaaagagatt aaaccgaagg tgattaaaag accttgaaat ccatgacgca gggagaattg
     3181 cgtcatttaa agcctagtta acgcatttac taaacgcaga cgaaaatgga aagattaatt
     3241 gggagtggta ggatgaaaca atttggagaa gatagaagtt tgaagtggaa aactggaaga
     3301 cagaagtacg ggaaggcgaa gaaaagaata gagaagatag ggaaattaga agataaaaac
     3361 atacttttag aagaaaaaag ataaatttaa acctgaaaag taggaagcag aagaaaaaag
     3421 acaagctagg aaacaaaaag ctaagggcaa aatgtacaaa ctctcgagca tgcatctaga
     3481 gggcccaatt cgccctatag tgagtcgtat tacaattcac tggccgtcgt tttacaacgt
     3541 cgtgactggg aaaaccctgg cgttacccaa cttaatcgcc ttgcagcaca tccccctttc
     3601 gccagctggc gtaatagcga agaggcccgc accgatcgcc cttcccaaca gttgcgcagc
     3661 ctgaatggcg aatggacgcg ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta
     3721 cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc cgctcctttc gctttcttcc
     3781 cttcctttct cgccacgttc gccggctttc cccgtcaagc tctaaatcgg gggctccctt
     3841 tagggttccg atttagtgct ttacggcacc tcgaccccaa aaaacttgat tagggtgatg
     3901 gttcacgtag tgggccatcg ccctgataga cggtttttcg ccctttgacg ttggagtcca
     3961 cgttctttaa tagtggactc ttgttccaaa ctggaacaac actcaaccct atctcggtct
     4021 attcttttga tttataaggg attttgccga tttcggccta ttggttaaaa aatgagctga
     4081 tttaacaaaa atttaacgcg aattttaaca aaattcaggg cgcaagggct gctaaaggaa
     4141 gcggaacacg tagaaagcca gtccgcagaa acggtgctga ccccggatga atgtcagcta
     4201 ctgggctatc tggacaaggg aaaacgcaag cgcaaagaga aagcaggtag cttgcagtgg
     4261 gcttacatgg cgatagctag actgggcggt tttatggaca gcaagcgaac cggaattgcc
     4321 agctggggcg ccctctggta aggttgggaa gccctgcaaa gtaaactgga tggctttctt
     4381 gccgccaagg atctgatggc gcaggggatc aagatctgat caagagacag gatgaggatc
     4441 gtttcgcatg attgaacaag atggattgca cgcaggttct ccggccgctt gggtggagag
     4501 gctattcggc tatgactggg cacaacagac aatcggctgc tctgatgccg ccgtgttccg
     4561 gctgtcagcg caggggcgcc cggttctttt tgtcaagacc gacctgtccg gtgccctgaa
     4621 tgaactgcag gacgaggcag cgcggctatc gtggctggcc acgacgggcg ttccttgcgc
     4681 agctgtgctc gacgttgtca ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc
     4741 ggggcaggat ctcctgtcat cccaccttgc tcctgccgag aaagtatcca tcatggctga
     4801 tgcaatgcgg cggctgcata cgcttgatcc ggctacctgc ccattcgacc accaagcgaa
     4861 acatcgcatc gagcgagcac gtactcggat ggaagccggt cttgtcgatc aggatgatct
     4921 ggacgaagag catcaggggc tcgcgccagc cgaactgttc gccaggctca aggcgcgcat
     4981 gcccgacggc gaggatctcg tcgtgaccca tggcgatgcc tgcttgccga atatcatggt
     5041 ggaaaatggc cgcttttctg gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta
     5101 tcaggacata gcgttggcta cccgtgatat tgctgaagag cttggcggcg aatgggctga
     5161 ccgcttcctc gtgctttacg gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg
     5221 ccttcttgac gagttcttct gaattgaaaa aggaagagta tgagtattca acatttccgt
     5281 gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca cccagaaacg
     5341 ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac gagtgggtta catcgaactg
     5401 gatctcaaca gcggtaagat ccttgagagt tttcgccccg aagaacgttt tccaatgatg
     5461 agcactttta aagttctgct atgtggcgcg gtattatccc gtattgacgc cgggcaagag
     5521 caactcggtc gccgcataca ctattctcag aatgacttgg ttgagtactc accagtcaca
     5581 gaaaagcatc ttacggatgg catgacagta agagaattat gcagtgctgc cataaccatg
     5641 agtgataaca ctgcggccaa cttacttctg acaacgatcg gaggaccgaa ggagctaacc
     5701 gcttttttgc acaacatggg ggatcatgta actcgccttg atcgttggga accggagctg
     5761 aatgaagcca taccaaacga cgagcgtgac accacgatgc ctgtagcaat ggcaacaacg
     5821 ttgcgcaaac tattaactgg cgaactactt actctagctt cccggcaaca attaatagac
     5881 tggatggagg cggataaagt tgcaggacca cttctgcgct cggcccttcc ggctggctgg
     5941 tttattgctg ataaatctgg agccggtgag cgtgggtctc gcggtatcat tgcagcactg
     6001 gggccagatg gtaagccctc ccgtatcgta gttatctaca cgacggggag tcaggcaact
     6061 atggatgaac gaaatagaca gatcgctgag ataggtgcct cactgattaa gcattggtaa
     6121 ctgtcagacc aagtttactc atatatactt tagattgatt taaaacttca tttttaattt
     6181 aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag
     6241 ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct
     6301 ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt
     6361 tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg
     6421 cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt caagaactct
     6481 gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc
     6541 gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg
     6601 tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa
     6661 ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg
     6721 gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg
     6781 ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga
     6841 tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt
     6901 ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct
     6961 gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg ccgcagccga
     7021 acgaccgagc gcagcgagtc agtgagcgag gaagcggaag
//
