LOCUS       sequence-264452-        7749 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 7749)
  AUTHORS   Hageman J, Kampinga HH
  TITLE     Computational analysis of the human HSPH/HSPA/DNAJ family and 
            cloning of a human HSPH/HSPA/DNAJ expression library.
  JOURNAL   Cell Stress Chaperones. 2009 Jan;14(1):1-21. doi: 
            10.1007/s12192-008-0060-2. Epub 2008 Aug 7.
  PUBMED    18686016
REFERENCE   2  (bases 1 to 7749)
  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: 
            "10.1007/s12192-008-0060-2"; journalName: "Cell Stress Chaperones"; 
            date: "2009-01"; volume: "14"; issue: "1"; pages: "1-21"
COMMENT     Sequence Label: pcDNA5/FRT/TO GFP HSPA8
FEATURES             Location/Qualifiers
     source          1..7749
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        74..453
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        454..657
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     608..628
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    659..677
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    680..698
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     708..732
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     regulatory      823..832
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(829..831,832..834,835..1545)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(829..831,832..834,835..1545)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(874..895)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1135..1154)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1482..1503
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(3540..3557)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3546..3770
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(3903..3922)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    4054..4101
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     CDS             4109..5131
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             4109..5131
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     polyA_signal    5261..5382
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5298..5317)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5352..5371
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5431..5447)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5431..5447)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5444..5466)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5455..5471
                     /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(5479..5485,5486..5503,5504..5509))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5524..5545
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5662..5679)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5833..6421)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5913..5932)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6592..7383,7384..7452))
                     /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     7215..7234
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7453..7557)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(7632..7651)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 aattgcatga agaatctgct tagggttagg cgttttgcgc tgcttcgcga tgtacgggcc
       61 agatatacgc gttgacattg attattgact agttattaat agtaatcaat tacggggtca
      121 ttagttcata gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct
      181 ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta
      241 acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac
      301 ttggcagtac atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt
      361 aaatggcccg cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag
      421 tacatctacg tattagtcat cgctattacc atggtgatgc ggttttggca gtacatcaat
      481 gggcgtggat agcggtttga ctcacgggga tttccaagtc tccaccccat tgacgtcaat
      541 gggagtttgt tttggcacca aaatcaacgg gactttccaa aatgtcgtaa caactccgcc
      601 ccattgacgc aaatgggcgg taggcgtgta cggtgggagg tctatataag cagagctctc
      661 cctatcagtg atagagatct ccctatcagt gatagagatc gtcgacgagc tcgtttagtg
      721 aaccgtcaga tcgcctggag acgccatcca cgctgttttg acctccatag aagacaccgg
      781 gaccgatcca gcctccggac tctagcgttt aaacttaagc ttgccaccat ggtgagcaag
      841 ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg agctggacgg cgacgtaaac
      901 ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg ccacctacgg caagctgacc
      961 ctgaagttca tctgcaccac cggcaagctg cccgtgccct ggcccaccct cgtgaccacc
     1021 ctgacctacg gcgtgcagtg cttcagccgc taccccgacc acatgaagca gcacgacttc
     1081 ttcaagtccg ccatgcccga aggctacgtc caggagcgca ccatcttctt caaggacgac
     1141 ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg acaccctggt gaaccgcatc
     1201 gagctgaagg gcatcgactt caaggaggac ggcaacatcc tggggcacaa gctggagtac
     1261 aactacaaca gccacaacgt ctatatcatg gccgacaagc agaagaacgg catcaaggtg
     1321 aacttcaaga tccgccacaa catcgaggac ggcagcgtgc agctcgccga ccactaccag
     1381 cagaacaccc ccatcggcga cggccccgtg ctgctgcccg acaaccacta cctgagcacc
     1441 cagtccgccc tgagcaaaga ccccaacgag aagcgcgatc acatggtcct gctggagttc
     1501 gtgaccgccg ccgggatcac tctcggcatg gacgagctgt acaagggatc caccatgtcc
     1561 aagggacctg cagttggtat tgatcttggc accacctact cttgtgtggg tgttttccag
     1621 cacggaaaag tcgagataat tgccaatgat cagggaaacc gaaccactcc aagctatgtc
     1681 gcctttacgg acactgaacg gttgatcggt gatgccgcaa agaatcaagt tgcaatgaac
     1741 cccaccaaca cagtttttga tgccaaacgt ctgattggac gcagatttga tgatgctgtt
     1801 gtccagtctg atatgaaaca ttggcccttt atggtggtga atgatgctgg caggcccaag
     1861 gtccaagtag aatacaaggg agagaccaaa agcttctatc cagaggaggt gtcttctatg
     1921 gttctgacaa agatgaagga aattgcagaa gcctaccttg ggaagactgt taccaatgct
     1981 gtggtcacag tgccagctta ctttaatgac tctcagcgtc aggctaccaa agatgctgga
     2041 actattgctg gtctcaatgt acttagaatt attaatgagc caactgctgc tgctattgct
     2101 tacggcttag acaaaaaggt tggagcagaa agaaacgtgc tcatctttga cctgggaggt
     2161 ggcacttttg atgtgtcaat cctcactatt gaggatggaa tctttgaggt caagtctaca
     2221 gctggagaca cccacttggg tggagaagat tttgacaacc gaatggtcaa ccattttatt
     2281 gctgagttta agcgcaagca taagaaggac atcagtgaga acaagagagc tgtaagacgc
     2341 ctccgtactg cttgtgaacg tgctaagcgt accctctctt ccagcaccca ggccagtatt
     2401 gagatcgatt ctctctatga aggaatcgac ttctatacct ccattacccg tgcccgattt
     2461 gaagaactga atgctgacct gttccgtggc accctggacc cagtagagaa agcccttcga
     2521 gatgccaaac tagacaagtc acagattcat gatattgtcc tggttggtgg ttctactcgt
     2581 atccccaaga ttcagaagct tctccaagac ttcttcaatg gaaaagaact gaataagagc
     2641 atcaaccctg atgaagctgt tgcttatggt gcagctgtcc aggcagccat cttgtctgga
     2701 gacaagtctg agaatgttca agatttgctg ctcttggatg tcactcctct ttcccttggt
     2761 attgaaactg ctggtggagt catgactgtc ctcatcaagc gtaataccac cattcctacc
     2821 aagcagacac agaccttcac tacctattct gacaaccagc ctggtgtgct tattcaggtt
     2881 tatgaaggcg agcgtgccat gacaaaggat aacaacctgc ttggcaagtt tgaactcaca
     2941 ggcatacctc ctgcaccccg aggtgttcct cagattgaag tcacttttga cattgatgcc
     3001 aatggtatac tcaatgtctc tgctgtggac aagagtacgg gaaaagagaa caagattact
     3061 atcactaatg acaagggccg tttgagcaag gaagacattg aacgtatggt ccaggaagct
     3121 gagaagtaca aagctgaaga tgagaagcag agggacaagg tgtcatccaa gaattcactt
     3181 gagtcctatg ccttcaacat gaaagcaact gttgaagatg agaaacttca aggcaagatt
     3241 aacgatgagg acaaacagaa gattctggac aagtgtaatg aaattatcaa ctggcttgat
     3301 aagaatcaga ctgctgagaa ggaagaattt gaacatcaac agaaagagct ggagaaagtt
     3361 tgcaacccca tcatcaccaa gctgtaccag agtgcaggag gcatgccagg aggaatgcct
     3421 gggggatttc ctggtggtgg agctcctccc tctggtggtg cttcctcagg gcccaccatt
     3481 gaagaggttg attaagcggc cgctcgagtc tagagggccc gtttaaaccc gctgatcagc
     3541 ctcgactgtg ccttctagtt gccagccatc tgttgtttgc ccctcccccg tgccttcctt
     3601 gaccctggaa ggtgccactc ccactgtcct ttcctaataa aatgaggaaa ttgcatcgca
     3661 ttgtctgagt aggtgtcatt ctattctggg gggtggggtg gggcaggaca gcaaggggga
     3721 ggattgggaa gacaatagca ggcatgctgg ggatgcggtg ggctctatgg cttctgaggc
     3781 ggaaagaacc agctggggct ctagggggta tccccacgcg ccctgtagcg gcgcattaag
     3841 cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc
     3901 cgctcctttc gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc
     3961 tctaaatcgg gggctccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa
     4021 aaaacttgat tagggtgatg gttcacgtac ctagaagttc ctattccgaa gttcctattc
     4081 tctagaaagt ataggaactt ccttggccaa aaagcctgaa ctcaccgcga cgtctgtcga
     4141 gaagtttctg atcgaaaagt tcgacagcgt ctccgacctg atgcagctct cggagggcga
     4201 agaatctcgt gctttcagct tcgatgtagg agggcgtgga tatgtcctgc gggtaaatag
     4261 ctgcgccgat ggtttctaca aagatcgtta tgtttatcgg cactttgcat cggccgcgct
     4321 cccgattccg gaagtgcttg acattgggga attcagcgag agcctgacct attgcatctc
     4381 ccgccgtgca cagggtgtca cgttgcaaga cctgcctgaa accgaactgc ccgctgttct
     4441 gcagccggtc gcggaggcca tggatgcgat cgctgcggcc gatcttagcc agacgagcgg
     4501 gttcggccca ttcggaccgc aaggaatcgg tcaatacact acatggcgtg atttcatatg
     4561 cgcgattgct gatccccatg tgtatcactg gcaaactgtg atggacgaca ccgtcagtgc
     4621 gtccgtcgcg caggctctcg atgagctgat gctttgggcc gaggactgcc ccgaagtccg
     4681 gcacctcgtg cacgcggatt tcggctccaa caatgtcctg acggacaatg gccgcataac
     4741 agcggtcatt gactggagcg aggcgatgtt cggggattcc caatacgagg tcgccaacat
     4801 cttcttctgg aggccgtggt tggcttgtat ggagcagcag acgcgctact tcgagcggag
     4861 gcatccggag cttgcaggat cgccgcggct ccgggcgtat atgctccgca ttggtcttga
     4921 ccaactctat cagagcttgg ttgacggcaa tttcgatgat gcagcttggg cgcagggtcg
     4981 atgcgacgca atcgtccgat ccggagccgg gactgtcggg cgtacacaaa tcgcccgcag
     5041 aagcgcggcc gtctggaccg atggctgtgt agaagtactc gccgatagtg gaaaccgacg
     5101 ccccagcact cgtccgaggg caaaggaata gcacgtacta cgagatttcg attccaccgc
     5161 cgccttctat gaaaggttgg gcttcggaat cgttttccgg gacgccggct ggatgatcct
     5221 ccagcgcggg gatctcatgc tggagttctt cgcccacccc aacttgttta ttgcagctta
     5281 taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat ttttttcact
     5341 gcattctagt tgtggtttgt ccaaactcat caatgtatct tatcatgtct gtataccgtc
     5401 gacctctagc tagagcttgg cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta
     5461 tccgctcaca attccacaca acatacgagc cggaagcata aagtgtaaag cctggggtgc
     5521 ctaatgagtg agctaactca cattaattgc gttgcgctca ctgcccgctt tccagtcggg
     5581 aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg
     5641 tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg
     5701 gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa
     5761 cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc
     5821 gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc
     5881 aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag
     5941 ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct
     6001 cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta
     6061 ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc
     6121 cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc
     6181 agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt
     6241 gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct
     6301 gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc
     6361 tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca
     6421 agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta
     6481 agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa
     6541 atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg
     6601 cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg
     6661 actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc
     6721 aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc
     6781 cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa
     6841 ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc
     6901 cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg
     6961 ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc
     7021 cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat
     7081 ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg
     7141 tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc
     7201 ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg
     7261 aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat
     7321 gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg
     7381 gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg
     7441 ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct
     7501 catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac
     7561 atttccccga aaagtgccac ctgacgtcga cggatcggga gatctcccga tcccctatgg
     7621 tgcactctca gtacaatctg ctctgatgcc gcatagttaa gccagtatct gctccctgct
     7681 tgtgtgttgg aggtcgctga gtagtgcgcg agcaaaattt aagctacaac aaggcaaggc
     7741 ttgaccgac
//
