LOCUS       sequence-358530-        6689 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 6689)
  AUTHORS   Scott KL, Kabbarah O, Liang MC, Ivanova E, Anagnostou V, Wu J, 
            Dhakal S, Wu M, Chen S, Feinberg T, Huang J, Saci A, Widlund HR, 
            Fisher DE, Xiao Y, Rimm DL, Protopopov A, Wong KK, Chin L
  TITLE     GOLPH3 modulates mTOR signalling and rapamycin sensitivity in 
            cancer.
  JOURNAL   Nature. 2009 Jun 25. 459(7250):1085-90.
  PUBMED    19553991
REFERENCE   2  (bases 1 to 6689)
  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: "Nature. 
            2009 Jun 25. 459(7250):1085-90."
COMMENT     Sequence Label: pEF-DEST51-GOLPH3
FEATURES             Location/Qualifiers
     source          1..6689
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..1179
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          232..1170
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1127..1147
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     primer_bind     1196..1215
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1196..1214
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1246..1270
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             1411..1425
                     /codon_start=1
                     /product="enterokinase recognition and cleavage site"
                     /label=enterokinase site
                     /translation="DDDDK"
     protein_bind    complement(2165..2189)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             2215..2256
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             2266..2283
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2306..2323)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2312..2536
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2582..3010
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2669..2688)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2879..2900
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3019..3039)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3024..3353
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3204..3339
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3266..3285
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        3401..3448
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             3467..3865
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     polyA_signal    4023..4144
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4060..4079)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4114..4133
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4193..4209)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4193..4209)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4206..4228)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4217..4233
                     /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(4241..4247,4248..4265,4266..4271))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4286..4307
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4424..4441)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4584..5172)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4664..4683)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5343..6134,6135..6203))
                     /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     5966..5985
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6204..6308)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6383..6402)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 ggctccggtg cccgtcagtg ggcagagcgc acatcgccca cagtccccga gaagttgggg
       61 ggaggggtcg gcaattgaac cggtgcctag agaaggtggc gcggggtaaa ctgggaaagt
      121 gatgtcgtgt actggctccg cctttttccc gagggtgggg gagaaccgta tataagtgca
      181 gtagtcgccg tgaacgttct ttttcgcaac gggtttgccg ccagaacaca ggtaagtgcc
      241 gtgtgtggtt cccgcgggcc tggcctcttt acgggttatg gcccttgcgt gccttgaatt
      301 acttccacct ggctgcagta cgtgattctt gatcccgagc ttcgggttgg aagtgggtgg
      361 gagagttcga ggccttgcgc ttaaggagcc ccttcgcctc gtgcttgagt tgaggcctgg
      421 cctgggcgct ggggccgccg cgtgcgaatc tggtggcacc ttcgcgcctg tctcgctgct
      481 ttcgataagt ctctagccat ttaaaatttt tgatgacctg ctgcgacgct ttttttctgg
      541 caagatagtc ttgtaaatgc gggccaagat ctgcacactg gtatttcggt ttttggggcc
      601 gcgggcggcg acggggcccg tgcgtcccag cgcacatgtt cggcgaggcg gggcctgcga
      661 gcgcggccac cgagaatcgg acgggggtag tctcaagctg gccggcctgc tctggtgcct
      721 ggcctcgcgc cgccgtgtat cgccccgccc tgggcggcaa ggctggcccg gtcggcacca
      781 gttgcgtgag cggaaagatg gccgcttccc ggccctgctg cagggagctc aaaatggagg
      841 acgcggcgct cgggagagcg ggcgggtgag tcacccacac aaaggaaaag ggcctttccg
      901 tcctcagccg tcgcttcatg tgactccacg gagtaccggg cgccgtccag gcacctcgat
      961 tagttctcga gcttttggag tacgtcgtct ttaggttggg gggaggggtt ttatgcgatg
     1021 gagtttcccc acactgagtg ggtggagact gaagttaggc cagcttggca cttgatgtaa
     1081 ttctccttgg aatttgccct ttttgagttt ggatcttggt tcattctcaa gcctcagaca
     1141 gtggttcaaa gtttttttct tccatttcag gtgtcgtgag gaattagctt ggtactaata
     1201 cgactcacta tagggagacc caagctggct aggtaagctt gatcaacaag tttgtacaaa
     1261 aaagttggca tgacctcgct gacccagcgc agctccggcc tggtgcagcg gcgcaccgag
     1321 gcctcccgca acgccgccga caaggagcgg gcggcgggcg gcggcgccgg cagcagcgag
     1381 gacgacgcgc agagccgccg cgacgagcag gacgacgacg acaagggcga ctccaaggaa
     1441 acgcggctga ccctgatgga ggaagtgctc ctgctgggcc tcaaggaccg cgagggttac
     1501 acatcatttt ggaatgactg tatatcatct ggattacgtg gctgtatgtt aattgaatta
     1561 gcattgagag gaaggttaca actagaggct tgtggaatga gacgtaaaag tctattaaca
     1621 agaaaggtaa tctgtaagtc agatgctcca acaggggatg ttcttcttga tgaagctctg
     1681 aagcatgtta aggaaactca gcctccagaa acggtccaga actggattga attacttagt
     1741 ggtgagacat ggaatccatt aaaattgcat tatcagttaa gaaatgtacg ggaacgatta
     1801 gctaaaaacc tggtggaaaa gggtgtattg acaacagaga aacagaactt cctacttttt
     1861 gacatgacaa cacatcccct caccaataac aacattaagc agcgcctcat caagaaagta
     1921 caggaagccg ttcttgacaa atgggtgaat gaccctcacc gcatggacag gcgcttgctg
     1981 gccctcattt acctggctca tgcctcggac gtcctggaga atgcttttgc tcctcttctg
     2041 gacgagcagt atgatttggc taccaagaga gtgcggcagc ttctcgactt agaccctgaa
     2101 gtggaatgtc tgaaggccaa caccaatgag gttctgtggg cggtggtggc ggcgttcacc
     2161 aagtacccaa ctttcttgta caaagtggtt gatctagagg gcccgcggtt cgaaggtaag
     2221 cctatcccta accctctcct cggtctcgat tctacgcgta ccggtcatca tcaccatcac
     2281 cattgagttt aaacccgctg atcagcctcg actgtgcctt ctagttgcca gccatctgtt
     2341 gtttgcccct cccccgtgcc ttccttgacc ctggaaggtg ccactcccac tgtcctttcc
     2401 taataaaatg aggaaattgc atcgcattgt ctgagtaggt gtcattctat tctggggggt
     2461 ggggtggggc aggacagcaa gggggaggat tgggaagaca atagcaggca tgctggggat
     2521 gcggtgggct ctatggcttc tgaggcggaa agaaccagct ggggctctag ggggtatccc
     2581 cacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc
     2641 gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc ctttctcgcc
     2701 acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt
     2761 agtgctttac ggcacctcga ccccaaaaaa cttgattagg gtgatggttc acgtagtggg
     2821 ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt ctttaatagt
     2881 ggactcttgt tccaaactgg aacaacactc aaccctatct cggtctattc ttttgattta
     2941 taagggattt tgccgatttc ggcctattgg ttaaaaaatg agctgattta acaaaaattt
     3001 aacgcgaatt aattctgtgg aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc
     3061 cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccagg tgtggaaagt
     3121 ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca
     3181 tagtcccgcc cctaactccg cccatcccgc ccctaactcc gcccagttcc gcccattctc
     3241 cgccccatgg ctgactaatt ttttttattt atgcagaggc cgaggccgcc tctgcctctg
     3301 agctattcca gaagtagtga ggaggctttt ttggaggcct aggcttttgc aaaaagctcc
     3361 cgggagcttg tatatccatt ttcggatctg atcagcacgt gttgacaatt aatcatcggc
     3421 atagtatatc ggcatagtat aatacgacaa ggtgaggaac taaaccatgg ccaagccttt
     3481 gtctcaagaa gaatccaccc tcattgaaag agcaacggct acaatcaaca gcatccccat
     3541 ctctgaagac tacagcgtcg ccagcgcagc tctctctagc gacggccgca tcttcactgg
     3601 tgtcaatgta tatcatttta ctgggggacc ttgtgcagaa ctcgtggtgc tgggcactgc
     3661 tgctgctgcg gcagctggca acctgacttg tatcgtcgcg atcggaaatg agaacagggg
     3721 catcttgagc ccctgcggac ggtgccgaca ggtgcttctc gatctgcatc ctgggatcaa
     3781 agccatagtg aaggacagtg atggacagcc gacggcagtt gggattcgtg aattgctgcc
     3841 ctctggttat gtgtgggagg gctaagcact tcgtggccga ggagcaggac tgacacgtgc
     3901 tacgagattt cgattccacc gccgccttct atgaaaggtt gggcttcgga atcgttttcc
     3961 gggacgccgg ctggatgatc ctccagcgcg gggatctcat gctggagttc ttcgcccacc
     4021 ccaacttgtt tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca
     4081 caaataaagc atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat
     4141 cttatcatgt ctgtataccg tcgacctcta gctagagctt ggcgtaatca tggtcatagc
     4201 tgtttcctgt gtgaaattgt tatccgctca caattccaca caacatacga gccggaagca
     4261 taaagtgtaa agcctggggt gcctaatgag tgagctaact cacattaatt gcgttgcgct
     4321 cactgcccgc tttccagtcg ggaaacctgt cgtgccagct gcattaatga atcggccaac
     4381 gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc actgactcgc
     4441 tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt
     4501 tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc caggaaccgt
     4561 aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa
     4621 aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt
     4681 ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg
     4741 tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc
     4801 agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc
     4861 gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta
     4921 tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct
     4981 acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc
     5041 tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa
     5101 caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa
     5161 aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa
     5221 aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt
     5281 ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac
     5341 agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc
     5401 atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt accatctggc
     5461 cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata
     5521 aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc
     5581 cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc
     5641 aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca
     5701 ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa
     5761 gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca
     5821 ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt
     5881 tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt
     5941 tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg
     6001 ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga
     6061 tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc
     6121 agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg
     6181 acacggaaat gttgaatact catactcttc ctttttcaat attattgaag catttatcag
     6241 ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg
     6301 gttccgcgca catttccccg aaaagtgcca cctgacgtcg acggatcggg agatctcccg
     6361 atcccctatg gtgcactctc agtacaatct gctctgatgc cgcatagtta agccagtatc
     6421 tgctccctgc ttgtgtgttg gaggtcgctg agtagtgcgc gagcaaaatt taagctacaa
     6481 caaggcaagg cttgaccgac aattgcatga agaatctgct tagggttagg cgttttgcgc
     6541 tgcttcgcga tgtacgggcc agatatacgc gttgacattg attattgact aggcttttgc
     6601 aaaaagcttt gcaaagatgg ataaagtttt aaacagagag gaatctttgc agctaatgga
     6661 ccttctaggt cttgaaagga gtgggaatt
//
