LOCUS       Exported                6257 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    pcDNA3.1 Flag hHMGB1
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6257)
  AUTHORS   Taniguchi N, Carames B, Kawakami Y, Amendt BA, Komiya S, Lotz M
  TITLE     Chromatin protein HMGB2 regulates articular cartilage surface 
            maintenance via beta-catenin pathway.
  JOURNAL   Proc Natl Acad Sci U S A. 2009 Sep 29. 106(39):16817-22.
  PUBMED    19805379
REFERENCE   2  (bases 1 to 6257)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6257
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     enhancer        97..476
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        477..680
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     631..651
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     725..744
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        725..743
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             952..975
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     promoter        complement(1672..1690)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(1673..1690)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(1710..1727)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1716..1940
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1986..2414
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2073..2092)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2283..2304
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2423..2443)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2428..2757
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2608..2743
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2670..2689
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2824..3618
                     /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(2878..2897)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3488..3507
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3792..3913
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3829..3848)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3883..3902
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3962..3978)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3962..3978)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3975..3997)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3986..4002
                     /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(4010..4040)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4055..4076
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4193..4210)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4364..4952)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4444..4463)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(5123..5983)
                     /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     5746..5765
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5984..6088)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6163..6182)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 caacaaggca aggcttgacc gacaattgca tgaagaatct gcttagggtt aggcgttttg
       61 cgctgcttcg cgatgtacgg gccagatata cgcgttgaca ttgattattg actagttatt
      121 aatagtaatc aattacgggg tcattagttc atagcccata tatggagttc cgcgttacat
      181 aacttacggt aaatggcccg cctggctgac cgcccaacga cccccgccca ttgacgtcaa
      241 taatgacgta tgttcccata gtaacgccaa tagggacttt ccattgacgt caatgggtgg
      301 agtatttacg gtaaactgcc cacttggcag tacatcaagt gtatcatatg ccaagtacgc
      361 cccctattga cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag tacatgacct
      421 tatgggactt tcctacttgg cagtacatct acgtattagt catcgctatt accatggtga
      481 tgcggttttg gcagtacatc aatgggcgtg gatagcggtt tgactcacgg ggatttccaa
      541 gtctccaccc cattgacgtc aatgggagtt tgttttggca ccaaaatcaa cgggactttc
      601 caaaatgtcg taacaactcc gccccattga cgcaaatggg cggtaggcgt gtacggtggg
      661 aggtctatat aagcagagct ctctggctaa ctagagaacc cactgcttac tggcttatcg
      721 aaattaatac gactcactat agggagaccc aagctccaca attcgggcgg cggctgctgc
      781 tgcctgtggc ccgggcggct gggagaagcc gagtgttggt gagtgacgcg gcggaggtgt
      841 agtttgacgc ggtgtgttac gtgggggaga gaataaaact ccagcgagat ccgggccgcg
      901 aacgaaagca gtgacggagg agcttgtacc accggtgact agatgaccat ggactacaag
      961 gacgacgatg acaagctgca ggaattcatg ggcaaaggag atcctaagaa gccgagaggc
     1021 aaaatgtcat catatgcatt ttttgtgcaa acttgtcggg aggagcataa gaagaagcac
     1081 ccagatgctt cagtcaactt ctcagagttt tctaagaagt gctcagagag gtggaagacc
     1141 atgtctgcta aagagaaagg aaaatttgaa gatatggcaa aggcggacaa ggcccgttat
     1201 gaaagagaaa tgaaaaccta tatccctccc aaaggggaga caaaaaagaa gttcaaggat
     1261 cccaatgcac ccaagaggcc tccttcggcc ttcttcctct tctgctctga gtatcgccca
     1321 aaaatcaaag gagaacatcc tggcctgtcc attggtgatg ttgcgaagaa actgggagag
     1381 atgtggaata acactgctgc agatgacaag cagccttatg aaaagaaggc tgcgaagctg
     1441 aaggaaaaat acgaaaagga tattgctgca tatcgagcta aaggaaagcc tgatgcagca
     1501 aaaaagggag ttgtcaaggc tgaaaaaagc aagaaaaaga aggaagagga ggaagatgag
     1561 gaagatgaag aggatgagga ggaggaggaa gatgaagaag atgaagatga agaagaagat
     1621 gatgatgatg aataagtcga cgcgtctgca gaagcttcta gagggcccta ttctatagtg
     1681 tcacctaaat gctagagctc gctgatcagc ctcgactgtg ccttctagtt gccagccatc
     1741 tgttgtttgc ccctcccccg tgccttcctt gaccctggaa ggtgccactc ccactgtcct
     1801 ttcctaataa aatgaggaaa ttgcatcgca ttgtctgagt aggtgtcatt ctattctggg
     1861 gggtggggtg gggcaggaca gcaaggggga ggattgggaa gacaatagca ggcatgctgg
     1921 ggatgcggtg ggctctatgg cttctgaggc ggaaagaacc agctggggct ctagggggta
     1981 tccccacgcg ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt
     2041 gaccgctaca cttgccagcg ccctagcgcc cgctcctttc gctttcttcc cttcctttct
     2101 cgccacgttc gccggctttc cccgtcaagc tctaaatcgg gggctccctt tagggttccg
     2161 atttagtgct ttacggcacc tcgaccccaa aaaacttgat tagggtgatg gttcacgtag
     2221 tgggccatcg ccctgataga cggtttttcg ccctttgacg ttggagtcca cgttctttaa
     2281 tagtggactc ttgttccaaa ctggaacaac actcaaccct atctcggtct attcttttga
     2341 tttataaggg attttgccga tttcggccta ttggttaaaa aatgagctga tttaacaaaa
     2401 atttaacgcg aattaattct gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc
     2461 tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac caggtgtgga
     2521 aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca
     2581 accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag ttccgcccat
     2641 tctccgcccc atggctgact aatttttttt atttatgcag aggccgaggc cgcctctgcc
     2701 tctgagctat tccagaagta gtgaggaggc ttttttggag gcctaggctt ttgcaaaaag
     2761 ctcccgggag cttgtatatc cattttcgga tctgatcaag agacaggatg aggatcgttt
     2821 cgcatgattg aacaagatgg attgcacgca ggttctccgg ccgcttgggt ggagaggcta
     2881 ttcggctatg actgggcaca acagacaatc ggctgctctg atgccgccgt gttccggctg
     2941 tcagcgcagg ggcgcccggt tctttttgtc aagaccgacc tgtccggtgc cctgaatgaa
     3001 ctgcaggacg aggcagcgcg gctatcgtgg ctggccacga cgggcgttcc ttgcgcagct
     3061 gtgctcgacg ttgtcactga agcgggaagg gactggctgc tattgggcga agtgccgggg
     3121 caggatctcc tgtcatctca ccttgctcct gccgagaaag tatccatcat ggctgatgca
     3181 atgcggcggc tgcatacgct tgatccggct acctgcccat tcgaccacca agcgaaacat
     3241 cgcatcgagc gagcacgtac tcggatggaa gccggtcttg tcgatcagga tgatctggac
     3301 gaagagcatc aggggctcgc gccagccgaa ctgttcgcca ggctcaaggc gcgcatgccc
     3361 gacggcgagg atctcgtcgt gacccatggc gatgcctgct tgccgaatat catggtggaa
     3421 aatggccgct tttctggatt catcgactgt ggccggctgg gtgtggcgga ccgctatcag
     3481 gacatagcgt tggctacccg tgatattgct gaagagcttg gcggcgaatg ggctgaccgc
     3541 ttcctcgtgc tttacggtat cgccgctccc gattcgcagc gcatcgcctt ctatcgcctt
     3601 cttgacgagt tcttctgagc gggactctgg ggttcgaaat gaccgaccaa gcgacgccca
     3661 acctgccatc acgagatttc gattccaccg ccgccttcta tgaaaggttg ggcttcggaa
     3721 tcgttttccg ggacgccggc tggatgatcc tccagcgcgg ggatctcatg ctggagttct
     3781 tcgcccaccc caacttgttt attgcagctt ataatggtta caaataaagc aatagcatca
     3841 caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca
     3901 tcaatgtatc ttatcatgtc tgtataccgt cgacctctag ctagagcttg gcgtaatcat
     3961 ggtcatagct gtttcctgtg tgaaattgtt atccgctcac aattccacac aacatacgag
     4021 ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt gagctaactc acattaattg
     4081 cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa
     4141 tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca
     4201 ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg
     4261 taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc
     4321 agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc
     4381 cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac
     4441 tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc
     4501 tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata
     4561 gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc
     4621 acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca
     4681 acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag
     4741 cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta
     4801 gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg
     4861 gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc
     4921 agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt
     4981 ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa
     5041 ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat
     5101 atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga
     5161 tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac
     5221 gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg
     5281 ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg
     5341 caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt
     5401 cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct
     5461 cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat
     5521 cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta
     5581 agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca
     5641 tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat
     5701 agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac
     5761 atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa
     5821 ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt
     5881 cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg
     5941 caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat
     6001 attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt
     6061 agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtcg
     6121 acggatcggg agatctcccg atcccctatg gtgcactctc agtacaatct gctctgatgc
     6181 cgcatagtta agccagtatc tgctccctgc ttgtgtgttg gaggtcgctg agtagtgcgc
     6241 gagcaaaatt taagcta
//
