LOCUS       sequence-190926-        8220 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Type II Hexokinase expressed in AS-30D rat hepatoma cell line.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8220)
  AUTHORS   Mathupala SP, Heese C, Pedersen PL
  TITLE     Glucose catabolism in cancer cells. The type II hexokinase promoter 
            contains functionally active response elements for the tumor 
            suppressor p53.
  JOURNAL   J Biol Chem. 1997 Sep 5;272(36):22776-80.
  PUBMED    9278438
REFERENCE   2  (bases 1 to 8220)
  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."; date: "1997-09-5"; volume: "272(36)"; pages: "22776-80"
COMMENT     Sequence Label: pCI-Neo-HKIImut
FEATURES             Location/Qualifiers
     source          1..8220
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        212..591
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        592..803
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     754..774
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     800..824
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     intron          964..1096
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     1079..1098
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     promoter        1141..1159
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      1176..1185
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     polyA_signal    3998..4119
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4009..4028)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     4063..4082
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     rep_origin      4305..4760
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4392..4411)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4602..4623
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        4877..5234
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        4877..5068
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(4900..4920)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      5085..5220
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             5285..6079
                     /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(5339..5358)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5949..5968
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    6143..6191
                     /note="synthetic polyadenylation signal"
     primer_bind     complement(6232..6251)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6351..6373
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(6411..6429)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        6497..6601
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6602..6670,6671..7462)
                     /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"
     rep_origin      join(7633..8220,1)
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8122..8141
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat
       61 ggctcgacag atcttcaata ttggccatta gccatattat tcattggtta tatagcataa
      121 atcaatattg gctattggcc attgcatacg ttgtatctat atcataatat gtacatttat
      181 attggctcat gtccaatatg accgccatgt tggcattgat tattgactag ttattaatag
      241 taatcaatta cggggtcatt agttcatagc ccatatatgg agttccgcgt tacataactt
      301 acggtaaatg gcccgcctgg ctgaccgccc aacgaccccc gcccattgac gtcaataatg
      361 acgtatgttc ccatagtaac gccaataggg actttccatt gacgtcaatg ggtggagtat
      421 ttacggtaaa ctgcccactt ggcagtacat caagtgtatc atatgccaag tccgccccct
      481 attgacgtca atgacggtaa atggcccgcc tggcattatg cccagtacat gaccttacgg
      541 gactttccta cttggcagta catctacgta ttagtcatcg ctattaccat ggtgatgcgg
      601 ttttggcagt acaccaatgg gcgtggatag cggtttgact cacggggatt tccaagtctc
      661 caccccattg acgtcaatgg gagtttgttt tggcaccaaa atcaacggga ctttccaaaa
      721 tgtcgtaaca actgcgatcg cccgccccgt tgacgcaaat gggcggtagg cgtgtacggt
      781 gggaggtcta tataagcaga gctcgtttag tgaaccgtca gatcactaga agctttattg
      841 cggtagttta tcacagttaa attgctaacg cagtcagtgc ttctgacaca acagtctcga
      901 acttaagctg cagtgactct cttaaggtag ccttgcagaa gttggtcgtg aggcactggg
      961 caggtaagta tcaaggttac aagacaggtt taaggagacc aatagaaact gggcttgtcg
     1021 agacagagaa gactcttgcg tttctgatag gcacctattg gtcttactga catccacttt
     1081 gcctttctct ccacaggtgt ccactcccag ttcaattaca gctcttaagg ctagagtact
     1141 taatacgact cactataggc tagcctcgag aattcgccac catggtcgcc tcgcatatga
     1201 tcgcctgctt attcacggag ctcaaccaaa accaagtgca gaaggttgac caatttctct
     1261 accacatgcg tctctcagat gagacccttc tggagatttc taggcggttc cggaaggaga
     1321 tggagaaagg gctaggagct accacgcacc ctacagcagc tgtgaaaatg ttgcctacct
     1381 ttgtgaggtc aactccggat gggacagaac atggggagtt cctggctctg gatcttggag
     1441 gaaccaactt ccgtgtgctc cgagtaaggg tgacggacaa tggcctccag agagtggaga
     1501 tggagaacca gatctacgcc atccttgagg acatcatgcg gggcagtgga acccagctgt
     1561 ttgaccacat cgccgaatgc ctggccaact tcatggacaa gctacaaatc aaagagaaga
     1621 agctccctct gggtttcacc ttctcgttcc cctgccacca gacaaaactg gatgagagtt
     1681 ttttggtctc gtggactaag gggttcaagt ccagtggcgt ggaaggcaga gatgtggtgg
     1741 acctgatccg gaaggctatc cagcgcagag gggactttga cattgacatt gtggccgtgg
     1801 tgaatgacac agttgggacc atgatgactt gtggctatga tgatcagaac tgcgagattg
     1861 gtctcattgt gggcactggc agcaacgcct gctacatgga ggaaatgcgt catattgaca
     1921 tggtggaggg agatgagggg cgcatgtgca tcaacatgga gtggggagcc tttggggacg
     1981 acggtacact caatgacatc cgaaccgagt ttgaccgaga gatcgacatg ggctcgctga
     2041 accctgggaa gcagctgttt gagaagatga ttagcgggat gtacatgggg gagctggtca
     2101 ggctcatcct ggtgaagatg gccaaggcag agctgttgtt ccaagggaaa ctcagcccag
     2161 aactccttac cactggctcc ttcgagacca aagatgtctc ggatattgaa gaggataagg
     2221 atggaatcga gaaggcctac caaatcctga tgcgcctggg tctgaatcca ttgcaggagg
     2281 attgtgtggc cacgcaccga atctgccaga ttgcgtccac gcgctcggcc agtctgtgcg
     2341 cagccaccct ggccgcggtg ctgtggcgaa tcaaagagaa caagggcgag gagcgacttc
     2401 gctccaccat cggtgtcgat ggctccgtct acaagaaaca tccccatttt gccaagcgtc
     2461 tccataaggc agtgaggagg ctggtgcccg actgtgatgt ccgcttcctc cgctctgagg
     2521 atggcagcgg caagggggct gctatggtga cggcggtggc ttaccgtctg gctgaccaac
     2581 accgggcccg ccagaagacc ctggagtctc tgaagctgag ccacgagcag cttctggagg
     2641 ttaagagaag aatgaaggtg gaaatggagc agggtctgag caaggagacg catgcggtcg
     2701 cccctgtgaa gatgctgccc acttacgtgt gtgccactcc agatggcaca gagaaaggag
     2761 acttcttggc cttggatctt ggaggaacaa acttccgggt cctgctggtg cgtgtgcgta
     2821 atggcaagcg gaggggcgtg gaggtgcata acaagatcta ctccatccca caggaggtta
     2881 tgcatggcac tggggaagag ctcttcgacc acattgtcca gtgcattgcg gacttcctgg
     2941 agtacatggg catgaagggc gtgtccctgc ctttgggttt cacattctcc ttcccttgcc
     3001 agcagaacag cctagaccag agcatcctcc tcaagtggac aaagggattc aaggcatctg
     3061 gctgcgaggg tgaggatgtg gtcaccttgc tgaaggaagc gattcaccgg cgagaggagt
     3121 ttgacctgga tgtggttgcc gtggtgaatg acacagttgg gactatgatg acttgtggct
     3181 acgaagaccc tcactgtgaa gttggcctta ttgttggcac cggaagcaac gcctgctaca
     3241 tggaagagat gcgtaatgtg gagctggtgg acggagggga gggacggatg tgtgtcaaca
     3301 tggagtgggg agcatttggg gacaatggct gcctggatga cttgcggacc gtgtttgatg
     3361 ttgctgtgga tgagctttct ctcaaccctg gcaaacagag gttcgagaag atgatcagcg
     3421 gcatgtactt gggagagatt gtgcgcaaca ttctcatcga tttcacgaag cgggggctgc
     3481 tcttccgagg ccgcatctca gagcgcctca agacaagggg aatctctgaa actaagttcc
     3541 tgtctcagat agagagcgac tgcctagccc tgctacaggt tcgtgccatc ctgcgccacc
     3601 tagggctgga gagcacgtgc gatgacagca tcatcgtgaa ggaggtgtgc actgtggttg
     3661 cccggcgcgc tgcacagctc tgtggcgcag gcatggccgc cgtagtggac aagataagag
     3721 agaaccgtgg gctggacaac cccaaagtga cagtgggcgt ggacgggact ctgtataagc
     3781 ttcatcctca ctttgccaag gtcatgcatg agacggtgag agatctggct ccgaaatgtg
     3841 acgtgtcctt cctggaatcc gaggacggca gtgggaaggg agcagctctc atcactgccg
     3901 tggcctgccg catccgggag gctgggcaga gatagtctag agtcgacccg ggcggccgct
     3961 tccctttagt gagggttaat gcttcgagca gacatgataa gatacattga tgagtttgga
     4021 caaaccacaa ctagaatgca gtgaaaaaaa tgctttattt gtgaaatttg tgatgctatt
     4081 gctttatttg taaccattat aagctgcaat aaacaagtta acaacaacaa ttgcattcat
     4141 tttatgtttc aggttcaggg ggagatgtgg gaggtttttt aaagcaagta aaacctctac
     4201 aaatgtggta aaatccgata aggatcgatc cgggctggcg taatagcgaa gaggcccgca
     4261 ccgatcgccc ttcccaacag ttgcgcagcc tgaatggcga atggacgcgc cctgtagcgg
     4321 cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg accgctacac ttgccagcgc
     4381 cctagcgccc gctcctttcg ctttcttccc ttcctttctc gccacgttcg ccggctttcc
     4441 ccgtcaagct ctaaatcggg ggctcccttt agggttccga tttagtgctt tacggcacct
     4501 cgaccccaaa aaacttgatt agggtgatgg ttcacgtagt gggccatcgc cctgatagac
     4561 ggtttttcgc cctttgacgt tggagtccac gttctttaat agtggactct tgttccaaac
     4621 tggaacaaca ctcaacccta tctcggtcta ttcttttgat ttataaggga ttttgccgat
     4681 ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga attttaacaa
     4741 aatattaacg cttacaattt cctgatgcgg tattttctcc ttacgcatct gtgcggtatt
     4801 tcacaccgca tacgcggatc tgcgcagcac catggcctga aataacctct gaaagaggaa
     4861 cttggttagg taccttctga ggcggaaaga accagctgtg gaatgtgtgt cagttagggt
     4921 gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt
     4981 cagcaaccag gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc
     5041 atctcaatta gtcagcaacc atagtcccgc ccctaactcc gcccatcccg cccctaactc
     5101 cgcccagttc cgcccattct ccgccccatg gctgactaat tttttttatt tatgcagagg
     5161 ccgaggccgc ctcggcctct gagctattcc agaagtagtg aggaggcttt tttggaggcc
     5221 taggcttttg caaaaagctt gattcttctg acacaacagt ctcgaactta aggctagagc
     5281 caccatgatt gaacaagatg gattgcacgc aggttctccg gccgcttggg tggagaggct
     5341 attcggctat gactgggcac aacagacaat cggctgctct gatgccgccg tgttccggct
     5401 gtcagcgcag gggcgcccgg ttctttttgt caagaccgac ctgtccggtg ccctgaatga
     5461 actgcaggac gaggcagcgc ggctatcgtg gctggccacg acgggcgttc cttgcgcagc
     5521 tgtgctcgac gttgtcactg aagcgggaag ggactggctg ctattgggcg aagtgccggg
     5581 gcaggatctc ctgtcatctc accttgctcc tgccgagaaa gtatccatca tggctgatgc
     5641 aatgcggcgg ctgcatacgc ttgatccggc tacctgccca ttcgaccacc aagcgaaaca
     5701 tcgcatcgag cgagcacgta ctcggatgga agccggtctt gtcgatcagg atgatctgga
     5761 cgaagagcat caggggctcg cgccagccga actgttcgcc aggctcaagg cgcgcatgcc
     5821 cgacggcgag gatctcgtcg tgacccatgg cgatgcctgc ttgccgaata tcatggtgga
     5881 aaatggccgc ttttctggat tcatcgactg tggccggctg ggtgtggcgg accgctatca
     5941 ggacatagcg ttggctaccc gtgatattgc tgaagagctt ggcggcgaat gggctgaccg
     6001 cttcctcgtg ctttacggta tcgccgctcc cgattcgcag cgcatcgcct tctatcgcct
     6061 tcttgacgag ttcttctgag cgggactctg gggttcgaaa tgaccgacca agcgacgccc
     6121 aacctgccat cacgatggcc gcaataaaat atctttattt tcattacatc tgtgtgttgg
     6181 ttttttgtgt gaatcgatag cgataaggat ccgcgtatgg tgcactctca gtacaatctg
     6241 ctctgatgcc gcatagttaa gccagccccg acacccgcca acacccgctg acgcgccctg
     6301 acgggcttgt ctgctcccgg catccgctta cagacaagct gtgaccgtct ccgggagctg
     6361 catgtgtcag aggttttcac cgtcatcacc gaaacgcgcg agacgaaagg gcctcgtgat
     6421 acgcctattt ttataggtta atgtcatgat aataatggtt tcttagacgt caggtggcac
     6481 ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac attcaaatat
     6541 gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa aaaggaagag
     6601 tatgagtatt caacatttcc gtgtcgccct tattcccttt tttgcggcat tttgccttcc
     6661 tgtttttgct cacccagaaa cgctggtgaa agtaaaagat gctgaagatc agttgggtgc
     6721 acgagtgggt tacatcgaac tggatctcaa cagcggtaag atccttgaga gttttcgccc
     6781 cgaagaacgt tttccaatga tgagcacttt taaagttctg ctatgtggcg cggtattatc
     6841 ccgtattgac gccgggcaag agcaactcgg tcgccgcata cactattctc agaatgactt
     6901 ggttgagtac tcaccagtca cagaaaagca tcttacggat ggcatgacag taagagaatt
     6961 atgcagtgct gccataacca tgagtgataa cactgcggcc aacttacttc tgacaacgat
     7021 cggaggaccg aaggagctaa ccgctttttt gcacaacatg ggggatcatg taactcgcct
     7081 tgatcgttgg gaaccggagc tgaatgaagc cataccaaac gacgagcgtg acaccacgat
     7141 gcctgtagca atggcaacaa cgttgcgcaa actattaact ggcgaactac ttactctagc
     7201 ttcccggcaa caattaatag actggatgga ggcggataaa gttgcaggac cacttctgcg
     7261 ctcggccctt ccggctggct ggtttattgc tgataaatct ggagccggtg agcgtgggtc
     7321 tcgcggtatc attgcagcac tggggccaga tggtaagccc tcccgtatcg tagttatcta
     7381 cacgacgggg agtcaggcaa ctatggatga acgaaataga cagatcgctg agataggtgc
     7441 ctcactgatt aagcattggt aactgtcaga ccaagtttac tcatatatac tttagattga
     7501 tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg ataatctcat
     7561 gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg tagaaaagat
     7621 caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa
     7681 accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa
     7741 ggtaactggc ttcagcagag cgcagatacc aaatactgtt cttctagtgt agccgtagtt
     7801 aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt
     7861 accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata
     7921 gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt
     7981 ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac
     8041 gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga
     8101 gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg
     8161 ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa
//
