LOCUS       sequence-222560-        7085 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 7085)
  AUTHORS   Gupta PB, Kuperwasser C, Brunet JP, Ramaswamy S, Kuo WL, Gray JW, 
            Naber SP, Weinberg RA
  TITLE     The melanocyte differentiation program predisposes to metastasis 
            after neoplastic transformation.
  JOURNAL   Nat Genet. 2005 Oct . 37(10):1047-54.
  PUBMED    16142232
REFERENCE   2  (bases 1 to 7085)
  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: "Nat Genet. 
            2005 Oct . 37(10):1047-54."
COMMENT     Sequence Label: siSlug3
FEATURES             Location/Qualifiers
     source          1..7085
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     98..115
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    232..253
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(268..273,274..291,292..298)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    306..322
                     /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)."
     primer_bind     311..333
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     330..346
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     330..346
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     365..385
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        367..385
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        413..639
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             640..820
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    867..992
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1485..1718
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1903..1947
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
     promoter        2245..2493
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     2245..2265
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     2416..2435
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_feature    2591..2708
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2757..3267
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     3186..3204
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     CDS             3289..3888
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(3289..3308)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     3785..3805
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             4016..4249
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    4321..4442
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4358..4377)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4412..4431
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      4482..4617
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(4637..4656)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4638..4656)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4666..4683)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4666..4682)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4675..4697)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      4824..5279
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4911..4930)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5121..5142
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        5305..5409
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5410..5478,5479..6270)
                     /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     complement(5628..5647)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6441..7029
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6930..6949
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg cctttgagtg
       61 agctgatacc gctcgccgca gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc
      121 ggaagagcgc ccaatacgca aaccgcctct ccccgcgcgt tggccgattc attaatgcag
      181 ctggcacgac aggtttcccg actggaaagc gggcagtgag cgcaacgcaa ttaatgtgag
      241 ttagctcact cattaggcac cccaggcttt acactttatg cttccggctc gtatgttgtg
      301 tggaattgtg agcggataac aatttcacac aggaaacagc tatgaccatg attacgccaa
      361 gcgcgcaatt aaccctcact aaagggaaca aaagctggag ctgcaagctt aatgtagtct
      421 tatgcaatac tcttgtagtc ttgcaacatg gtaacgatga gttagcaaca tgccttacaa
      481 ggagagaaaa agcaccgtgc atgccgattg gtggaagtaa ggtggtacga tcgtgcctta
      541 ttaggaaggc aacagacggg tctgacatgg attggacgaa ccactgaatt gccgcattgc
      601 agagatattg tatttaagtg cctagctcga tacataaacg ggtctctctg gttagaccag
      661 atctgagcct gggagctctc tggctaacta gggaacccac tgcttaagcc tcaataaagc
      721 ttgccttgag tgcttcaagt agtgtgtgcc cgtctgttgt gtgactctgg taactagaga
      781 tccctcagac ccttttagtc agtgtggaaa atctctagca gtggcgcccg aacagggact
      841 tgaaagcgaa agggaaacca gaggagctct ctcgacgcag gactcggctt gctgaagcgc
      901 gcacggcaag aggcgagggg cggcgactgg tgagtacgcc aaaaattttg actagcggag
      961 gctagaagga gagagatggg tgcgagagcg tcagtattaa gcgggggaga attagatcgc
     1021 gatgggaaaa aattcggtta aggccagggg gaaagaaaaa atataaatta aaacatatag
     1081 tatgggcaag cagggagcta gaacgattcg cagttaatcc tggcctgtta gaaacatcag
     1141 aaggctgtag acaaatactg ggacagctac aaccatccct tcagacagga tcagaagaac
     1201 ttagatcatt atataataca gtagcaaccc tctattgtgt gcatcaaagg atagagataa
     1261 aagacaccaa ggaagcttta gacaagatag aggaagagca aaacaaaagt aagaccaccg
     1321 cacagcaagc ggccgctgat cttcagacct ggaggaggag atatgaggga caattggaga
     1381 agtgaattat ataaatataa agtagtaaaa attgaaccat taggagtagc acccaccaag
     1441 gcaaagagaa gagtggtgca gagagaaaaa agagcagtgg gaataggagc tttgttcctt
     1501 gggttcttgg gagcagcagg aagcactatg ggcgcagcgt caatgacgct gacggtacag
     1561 gccagacaat tattgtctgg tatagtgcag cagcagaaca atttgctgag ggctattgag
     1621 gcgcaacagc atctgttgca actcacagtc tggggcatca agcagctcca ggcaagaatc
     1681 ctggctgtgg aaagatacct aaaggatcaa cagctcctgg ggatttgggg ttgctctgga
     1741 aaactcattt gcaccactgc tgtgccttgg aatgctagtt ggagtaataa atctctggaa
     1801 cagatttgga atcacacgac ctggatggag tgggacagag aaattaacaa ttacacaagc
     1861 ttaatacact ccttaattga agaatcgcaa aaccagcaag aaaagaatga acaagaatta
     1921 ttggaattag ataaatgggc aagtttgtgg aattggttta acataacaaa ttggctgtgg
     1981 tatataaaat tattcataat gatagtagga ggcttggtag gtttaagaat agtttttgct
     2041 gtactttcta tagtgaatag agttaggcag ggatattcac cattatcgtt tcagacccac
     2101 ctcccaaccc cgaggggacc cgacaggccc gaaggaatag aagaagaagg tggagagaga
     2161 gacagagaca gatccattcg attagtgaac ggatctcgac ggtatcgatc acgagactag
     2221 cctcgagcgg ccgccccctt caccgagggc ctatttccca tgattccttc atatttgcat
     2281 atacgataca aggctgttag agagataatt ggaattaatt tgactgtaaa cacaaagata
     2341 ttagtacaaa atacgtgacg tagaaagtaa taatttcttg ggtagtttgc agttttaaaa
     2401 ttatgtttta aaatggacta tcatatgctt accgtaactt gaaagtattt cgatttcttg
     2461 gctttatata tcttgtggaa aggacgaaac accggtcaga cccattctga tgtaaagctc
     2521 gagctttaca tcagaatggg tctgtttttg aattctcgac ctcgagacaa atggcagtat
     2581 tcatccacaa ttttaaaaga aaagggggga ttggggggta cagtgcaggg gaaagaatag
     2641 tagacataat agcaacagac atacaaacta aagaattaca aaaacaaatt acaaaaattc
     2701 aaaattttcg ggtttattac agggacagca gagatccact ttggccgcgg ctcgaggggg
     2761 ttggggttgc gccttttcca aggcagccct gggtttgcgc agggacgcgg ctgctctggg
     2821 cgtggttccg ggaaacgcag cggcgccgac cctgggtctc gcacattctt cacgtccgtt
     2881 cgcagcgtca cccggatctt cgccgctacc cttgtgggcc ccccggcgac gcttcctgct
     2941 ccgcccctaa gtcgggaagg ttccttgcgg ttcgcggcgt gccggacgtg acaaacggaa
     3001 gccgcacgtc tcactagtac cctcgcagac ggacagcgcc agggagcaat ggcagcgcgc
     3061 cgaccgcgat gggctgtggc caatagcggc tgctcagcag ggcgcgccga gagcagcggc
     3121 cgggaagggg cggtgcggga ggcggggtgt ggggcggtag tgtgggccct gttcctgccc
     3181 gcgcggtgtt ccgcattctg caagcctccg gagcgcacgt cggcagtcgg ctccctcgtt
     3241 gaccgaatca ccgacctctc tccccagggg gatccaccgg agcttaccat gaccgagtac
     3301 aagcccacgg tgcgcctcgc cacccgcgac gacgtcccca gggccgtacg caccctcgcc
     3361 gccgcgttcg ccgactaccc cgccacgcgc cacaccgtcg atccggaccg ccacatcgag
     3421 cgggtcaccg agctgcaaga actcttcctc acgcgcgtcg ggctcgacat cggcaaggtg
     3481 tgggtcgcgg acgacggcgc cgcggtggcg gtctggacca cgccggagag cgtcgaagcg
     3541 ggggcggtgt tcgccgagat cggcccgcgc atggccgagt tgagcggttc ccggctggcc
     3601 gcgcagcaac agatggaagg cctcctggcg ccgcaccggc ccaaggagcc cgcgtggttc
     3661 ctggccaccg tcggcgtctc gcccgaccac cagggcaagg gtctgggcag cgccgtcgtg
     3721 ctccccggag tggaggcggc cgagcgcgcc ggggtgcccg ccttcctgga gacctccgcg
     3781 ccccgcaacc tccccttcta cgagcggctc ggcttcaccg tcaccgccga cgtcgaggtg
     3841 cccgaaggac cgcgcacctg gtgcatgacc cgcaagcccg gtgcctgacg cccgccccac
     3901 gacccgcagc gcccgaccga aaggagcgca cgaccccatg catcggtacc tttaagacca
     3961 atgacttaca aggcagctgt agatcttagc cactttttaa aagaaaaggg gggactggaa
     4021 gggctaattc actcccaacg aagacaagat ctgctttttg cttgtactgg gtctctctgg
     4081 ttagaccaga tctgagcctg ggagctctct ggctaactag ggaacccact gcttaagcct
     4141 caataaagct tgccttgagt gcttcaagta gtgtgtgccc gtctgttgtg tgactctggt
     4201 aactagagat ccctcagacc cttttagtca gtgtggaaaa tctctagcag tagtagttca
     4261 tgtcatctta ttattcagta tttataactt gcaaagaaat gaatatcaga gagtgagagg
     4321 aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca
     4381 aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct
     4441 tatcatgtct ggctctagct atcccgcccc taactccgcc catcccgccc ctaactccgc
     4501 ccagttccgc ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg
     4561 aggccgcctc ggcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag
     4621 ggacgtaccc aattcgccct atagtgagtc gtattacgcg cgctcactgg ccgtcgtttt
     4681 acaacgtcgt gactgggaaa accctggcgt tacccaactt aatcgccttg cagcacatcc
     4741 ccctttcgcc agctggcgta atagcgaaga ggcccgcacc gatcgccctt cccaacagtt
     4801 gcgcagcctg aatggcgaat gggacgcgcc ctgtagcggc gcattaagcg cggcgggtgt
     4861 ggtggttacg cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc
     4921 tttcttccct tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg
     4981 gctcccttta gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgatta
     5041 gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt
     5101 ggagtccacg ttctttaata gtggactctt gttccaaact ggaacaacac tcaaccctat
     5161 ctcggtctat tcttttgatt tataagggat tttgccgatt tcggcctatt ggttaaaaaa
     5221 tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa atattaacgc ttacaattta
     5281 ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat
     5341 tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa
     5401 aggaagagta tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt
     5461 tgccttcctg tttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag
     5521 ttgggtgcac gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt
     5581 tttcgccccg aagaacgttt tccaatgatg agcactttta aagttctgct atgtggcgcg
     5641 gtattatccc gtattgacgc cgggcaagag caactcggtc gccgcataca ctattctcag
     5701 aatgacttgg ttgagtactc accagtcaca gaaaagcatc ttacggatgg catgacagta
     5761 agagaattat gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg
     5821 acaacgatcg gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta
     5881 actcgccttg atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac
     5941 accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg cgaactactt
     6001 actctagctt cccggcaaca attaatagac tggatggagg cggataaagt tgcaggacca
     6061 cttctgcgct cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag
     6121 cgtgggtctc gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta
     6181 gttatctaca cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag
     6241 ataggtgcct cactgattaa gcattggtaa ctgtcagacc aagtttactc atatatactt
     6301 tagattgatt taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat
     6361 aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta
     6421 gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa
     6481 acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt
     6541 tttccgaagg taactggctt cagcagagcg cagataccaa atactgttct tctagtgtag
     6601 ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta
     6661 atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca
     6721 agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag
     6781 cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa
     6841 agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga
     6901 acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc
     6961 gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc
     7021 ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt
     7081 gctca
//
