LOCUS       sequence-134991-        5882 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 5882)
  AUTHORS   Yang Q, Manicone A, Coursen JD, Linke SP, Nagashima M, Forgues M, 
            Wang XW
  TITLE     Identification of a functional domain in a GADD45-mediated G2/M 
            checkpoint.
  JOURNAL   J Biol Chem. 2000 Nov 24. 275(47):36892-8.
  PUBMED    10973963
REFERENCE   2  (bases 1 to 5882)
  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: "J Biol 
            Chem. 2000 Nov 24. 275(47):36892-8."
COMMENT     Sequence Label: pcDNA3-HA+WT GADD45
FEATURES             Location/Qualifiers
     source          1..5882
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(11..28)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(182..770)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(262..281)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(941..1732,1733..1801))
                     /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     1564..1583
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1802..1906)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(1981..2000)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        2172..2551
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2552..2755
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2706..2726
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     2800..2819
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2800..2818
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             3333..3359
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     promoter        complement(3372..3390)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(3373..3390)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(3410..3427)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3416..3640
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3686..4114
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3773..3792)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3983..4004
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4123..4143)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4128..4457
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4308..4443
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4524..5318
                     /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(4578..4597)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5188..5207
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5492..5613
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5529..5548)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5583..5602
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5662..5678)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5662..5678)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5675..5697)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5686..5702
                     /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(5710..5716,5717..5734,5735..5740))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5755..5776
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
ORIGIN
        1 cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat
       61 cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga
      121 acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt
      181 ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt
      241 ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc
      301 gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa
      361 gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct
      421 ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta
      481 actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg
      541 gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc
      601 ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta
      661 ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg
      721 gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt
      781 tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg
      841 tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta
      901 aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg
      961 aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg
     1021 tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc
     1081 gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg
     1141 agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg
     1201 aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag
     1261 gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat
     1321 caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc
     1381 cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc
     1441 ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa
     1501 ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac
     1561 gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt
     1621 cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc
     1681 gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa
     1741 caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca
     1801 tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat
     1861 acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa
     1921 aagtgccacc tgacgtcgac ggatcgggag atctcccgat cccctatggt gcactctcag
     1981 tacaatctgc tctgatgccg catagttaag ccagtatctg ctccctgctt gtgtgttgga
     2041 ggtcgctgag tagtgcgcga gcaaaattta agctacaaca aggcaaggct tgaccgacaa
     2101 ttgcatgaag aatctgctta gggttaggcg ttttgcgctg cttcgcgatg tacgggccag
     2161 atatacgcgt tgacattgat tattgactag ttattaatag taatcaatta cggggtcatt
     2221 agttcatagc ccatatatgg agttccgcgt tacataactt acggtaaatg gcccgcctgg
     2281 ctgaccgccc aacgaccccc gcccattgac gtcaataatg acgtatgttc ccatagtaac
     2341 gccaataggg actttccatt gacgtcaatg ggtggagtat ttacggtaaa ctgcccactt
     2401 ggcagtacat caagtgtatc atatgccaag tacgccccct attgacgtca atgacggtaa
     2461 atggcccgcc tggcattatg cccagtacat gaccttatgg gactttccta cttggcagta
     2521 catctacgta ttagtcatcg ctattaccat ggtgatgcgg ttttggcagt acatcaatgg
     2581 gcgtggatag cggtttgact cacggggatt tccaagtctc caccccattg acgtcaatgg
     2641 gagtttgttt tggcaccaaa atcaacggga ctttccaaaa tgtcgtaaca actccgcccc
     2701 attgacgcaa atgggcggta ggcgtgtacg gtgggaggtc tatataagca gagctctctg
     2761 gctaactaga gaacccactg cttactggct tatcgaaatt aatacgactc actataggga
     2821 gacccaagct tatgactttg gaggaattct cggctggaga gcagaagacc gaaaggatgg
     2881 ataaggtggg ggatgccctg gaggaagtgc tcagcaaagc cctgagtcag cgcacgatca
     2941 ctgtcggggt gtacgaagcg gccaagctgc tcaacgtcga ccccgataac gtggtgttgt
     3001 gcctgctggc ggcggacgag gacgacgaca gagatgtggc tctgcagatc cacttcaccc
     3061 tgatccaggc gttttgctgc gagaacgaca tcaacatcct gcgcgtcagc aacccgggcc
     3121 ggctggcgga gctcctgctc ttggagaccg acgctggccc cgcggcgagc gagggcgccg
     3181 agcagccccc ggacctgcac tgcgtgctgg tgacgaatcc acattcatct caatggaagg
     3241 atcctgcctt aagtcaactt atttgttttt gccgggaaag tcgctacatg gatcaatggg
     3301 ttccagtgat taatctccct gaacggtcta gataccctta tgatgtgcca gattatgcct
     3361 gagggcccta ttctatagtg tcacctaaat gctagagctc gctgatcagc ctcgactgtg
     3421 ccttctagtt gccagccatc tgttgtttgc ccctcccccg tgccttcctt gaccctggaa
     3481 ggtgccactc ccactgtcct ttcctaataa aatgaggaaa ttgcatcgca ttgtctgagt
     3541 aggtgtcatt ctattctggg gggtggggtg gggcaggaca gcaaggggga ggattgggaa
     3601 gacaatagca ggcatgctgg ggatgcggtg ggctctatgg cttctgaggc ggaaagaacc
     3661 agctggggct ctagggggta tccccacgcg ccctgtagcg gcgcattaag cgcggcgggt
     3721 gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc cgctcctttc
     3781 gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc tctaaatcgg
     3841 gggctccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa aaaacttgat
     3901 tagggtgatg gttcacgtag tgggccatcg ccctgataga cggtttttcg ccctttgacg
     3961 ttggagtcca cgttctttaa tagtggactc ttgttccaaa ctggaacaac actcaaccct
     4021 atctcggtct attcttttga tttataaggg attttgccga tttcggccta ttggttaaaa
     4081 aatgagctga tttaacaaaa atttaacgcg aattaattct gtggaatgtg tgtcagttag
     4141 ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt
     4201 agtcagcaac caggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca
     4261 tgcatctcaa ttagtcagca accatagtcc cgcccctaac tccgcccatc ccgcccctaa
     4321 ctccgcccag ttccgcccat tctccgcccc atggctgact aatttttttt atttatgcag
     4381 aagccgaggc cgcctctgcc tctgagctat tccagaagta gtgaggaggc ttttttggag
     4441 gcctaggctt ttgcaaaaag ctcccgggag cttgtatatc cattttcgga tctgatcaag
     4501 agacaggatg aggatcgttt cgcatgattg aacaagatgg attgcacgca ggttctccgg
     4561 ccgcttgggt ggagaggcta ttcggctatg actgggcaca acagacaatc ggctgctctg
     4621 atgccgccgt gttccggctg tcagcgcagg ggcgcccggt tctttttgtc aagaccgacc
     4681 tgtccggtgc cctgaatgaa ctgcaggacg aggcagcgcg gctatcgtgg ctggccacga
     4741 cgggcgttcc ttgcgcagct gtgctcgacg ttgtcactga agcgggaagg gactggctgc
     4801 tattgggcga agtgccgggg caggatctcc tgtcatctca ccttgctcct gccgagaaag
     4861 tatccatcat ggctgatgca atgcggcggc tgcatacgct tgatccggct acctgcccat
     4921 tcgaccacca agcgaaacat cgcatcgagc gagcacgtac tcggatggaa gccggtcttg
     4981 tcgatcagga tgatctggac gaagagcatc aggggctcgc gccagccgaa ctgttcgcca
     5041 ggctcaaggc gcgcatgccc gacggcgagg atctcgtcgt gacccatggc gatgcctgct
     5101 tgccgaatat catggtggaa aatggccgct tttctggatt catcgactgt ggccggctgg
     5161 gtgtggcgga ccgctatcag gacatagcgt tggctacccg tgatattgct gaagagcttg
     5221 gcggcgaatg ggctgaccgc ttcctcgtgc tttacggtat cgccgctccc gattcgcagc
     5281 gcatcgcctt ctatcgcctt cttgacgagt tcttctgagc gggactctgg ggttcgaaat
     5341 gaccgaccaa gcgacgccca acctgccatc acgagatttc gattccaccg ccgccttcta
     5401 tgaaaggttg ggcttcggaa tcgttttccg ggacgccggc tggatgatcc tccagcgcgg
     5461 ggatctcatg ctggagttct tcgcccaccc caacttgttt attgcagctt ataatggtta
     5521 caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag
     5581 ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc tgtataccgt cgacctctag
     5641 ctagagcttg gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt atccgctcac
     5701 aattccacac aacatacgag ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt
     5761 gagctaactc acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc
     5821 gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg
     5881 ct
//
