LOCUS       sequence-7041-ex        6912 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 6912)
  AUTHORS   Colwill K, Wells CD, Elder K, Goudreault M, Hersi K, Kulkarni S, 
            Hardy WR, Pawson T, Morin GB
  TITLE     Modification of the Creator recombination system for proteomics 
            applications--improved expression by addition of splice sites.
  JOURNAL   BMC Biotechnol. 2006 Mar 6;6:13. doi: 10.1186/1472-6750-6-13.
  PUBMED    16519801
REFERENCE   2  (bases 1 to 6912)
  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: "BMC 
            Biotechnol."; date: "2006-03-6"; volume: "6:13. doi"; pages: " 
            10.1186/1472-6750-6-13"
COMMENT     Sequence Label: V207 pRETRO-Triple-Flag SD - Acceptor
FEATURES             Location/Qualifiers
     source          1..6912
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    579..920
                     /label=MESV Psi
                     /note="packaging signal of murine embryonic stem cell 
                     virus"
     CDS             987..1403
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     1333..1355
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     1371..1387
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     CDS             1429..1494
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     protein_bind    complement(1878..1911)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        2055..2554
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(2090..2113)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     2487..2506
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     CDS             2575..3174
                     /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(2575..2594)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     3071..3091
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             3274..3788
                     /label=3' LTR
                     /note="3' long terminal repeat from murine embryonic stem 
                     cell virus"
     primer_bind     complement(3957..3973)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3957..3973)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3970..3992)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3981..3997
                     /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(4005..4011,4012..4029,4030..4035))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4050..4071
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4188..4205)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4359..4947)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4439..4458)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5118..5909,5910..5978))
                     /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     5741..5760
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5979..6083)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6151..6169
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(6207..6229)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     6329..6348
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6542..6564
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     6692..6711
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     LTR             join(6911..6912,1..515)
                     /label=5' LTR
                     /note="5' long terminal repeat from murine embryonic stem 
                     cell virus"
ORIGIN
        1 tgaaagaccc cacctgtagg tttggcaagc tagcttaagt aacgccattt tgcaaggcat
       61 ggaaaataca taactgagaa tagagaagtt cagatcaagg ttaggaacag agagacagca
      121 gaatatgggc caaacaggat atctgtggta agcagttcct gccccggctc agggccaaga
      181 acagatggtc cccagatgcg gtcccgccct cagcagtttc tagagaacca tcagatgttt
      241 ccagggtgcc ccaaggacct gaaatgaccc tgtgccttat ttgaactaac caatcagttc
      301 gcttctcgct tctgttcgcg cgcttctgct ccccgagctc aataaaagag cccacaaccc
      361 ctcactcggc gcgccagtcc tccgatagac tgcgtcgccc gggtacccgt attcccaata
      421 aagcctcttg ctgtttgcat ccgaatcgtg gactcgctga tccttgggag ggtctcctca
      481 gattgattga ctgcccacct cgggggtctt tcatttggag gttccaccga gatttggaga
      541 cccctgccca gggaccaccg acccccccgc cgggaggtaa gctggccagc ggtcgtttcg
      601 tgtctgtctc tgtctttgtg cgtgtttgtg ccggcatcta atgtttgcgc ctgcgtctgt
      661 actagttagc taactagctc tgtatctggc ggacccgtgg tggaactgac gagttctgaa
      721 cacccggccg caaccctggg agacgtccca gggactttgg gggccgtttt tgtggcccga
      781 cctgaggaag ggagtcgatg tggaatccga ccccgtcagg atatgtggtt ctggtaggag
      841 acgagaacct aaaacagttc ccgcctccgt ctgaattttt gctttcggtt tggaaccgaa
      901 gccgcgcgtc ttgtctgctg cagcgctgca gcatcgttct gtgttgtctc tgtctgactg
      961 tgtttctgta tttgtctgaa aattagggcc agactgttac cactccctta agtttgacct
     1021 taggtcactg gaaagatgtc gagcggatcg ctcacaacca gtcggtagat gtcaagaaga
     1081 gacgttgggt taccttctgc tctgcagaat ggccaacctt taacgtcgga tggccgcgag
     1141 acggcacctt taaccgagac ctcatcaccc aggttaagat caaggtcttt tcacctggcc
     1201 cgcatggaca cccagaccag gtcccctaca tcgtgacctg ggaagccttg gcttttgacc
     1261 cccctccctg ggtcaagccc tttgtacacc ctaagcctcc gcctcctctt cctccatccg
     1321 ccccgtctct cccccttgaa cctcctcgtt cgaccccgcc tcgatcctcc ctttatccag
     1381 ccctcactcc ttctctaggc gccggaatta gatctctcga gccacatgga ctacaaggac
     1441 cacgacggcg actacaagga ccacgacatc gactacaagg acgacgacga caaggtaagt
     1501 agatcttgac ctctcaaaag cgggcatgac ttctgcgcta agattgtcag tttccaaaaa
     1561 cgaggaggat ttgatattca cctggcccgc ggtgatgcct ttgagggtgg ccgcatccat
     1621 ctggtcagaa aagacaatct ttttgttgtc aagcttggtg gcaaacgacc cgtagagggc
     1681 gttggacagc aacttggcga tggagcgcag ggtttggttt ttgtcgcgat cggcgcgctc
     1741 cttggccgcg atgtttagct gcacgtattc gcgcgcaacg caccgccatt cgggaaagac
     1801 ggtggtgcgc tcgtcgggca ccaggtgcac gcgccaacag cggttgtgca gggtgacaag
     1861 gtcaacgcta agcttcgata acttcgtata gcatacatta tacgaagtta tagatccaat
     1921 attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt
     1981 agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtgt
     2041 taacgaattc taccgggtag gggaggcgct tttcccaagg cagtctggag catgcgcttt
     2101 agcagccccg ctgggcactt ggcgctacac aagtggcctc tggcctcgca cacattccac
     2161 atccaccggt aggcgccaac cggctccgtt ctttggtggc cccttcgcgc caccttctac
     2221 tcctccccta gtcaggaagt tcccccccgc cccgcagctc gcgtcgtgca ggacgtgaca
     2281 aatggaagta gcacgtctca ctagtctcgt gcagatggac agcaccgctg agcaatggaa
     2341 gcgggtaggc ctttggggca gcggccaata gcagctttgc tccttcgctt tctgggctca
     2401 gaggctggga aggggtgggt ccgggggcgg gctcaggggc gggctcaggg gcggggcggg
     2461 cgcccgaagg tcctccggag gcccggcatt ctgcacgctt caaaagcgca cgtctgccgc
     2521 gctgttctcc tcttcctcat ctccgggcct ttcgacctgc agcccaagct taccatgacc
     2581 gagtacaagc ccacggtgcg cctcgccacc cgcgacgacg tccccagggc cgtacgcacc
     2641 ctcgccgccg cgttcgccga ctaccccgcc acgcgccaca ccgtcgatcc ggaccgccac
     2701 atcgagcggg tcaccgagct gcaagaactc ttcctcacgc gcgtcgggct cgacatcggc
     2761 aaggtgtggg tcgcggacga cggcgccgcg gtggcggtct ggaccacgcc ggagagcgtc
     2821 gaagcggggg cggtgttcgc cgagatcggc ccgcgcatgg ccgagttgag cggttcccgg
     2881 ctggccgcgc agcaacagat ggaaggcctc ctggcgccgc accggcccaa ggagcccgcg
     2941 tggttcctgg ccaccgtcgg cgtctcgccc gaccaccagg gcaagggtct gggcagcgcc
     3001 gtcgtgctcc ccggagtgga ggcggccgag cgcgccgggg tgcccgcctt cctggagacc
     3061 tccgcgcccc gcaacctccc cttctacgag cggctcggct tcaccgtcac cgccgacgtc
     3121 gaggtgcccg aaggaccgcg cacctggtgc atgacccgca agcccggtgc ctgacgcccg
     3181 ccccacgacc cgcagcgccc gaccgaaagg agcgcacgac cccatgcatc gataaaataa
     3241 aagattttat ttagtctcca gaaaaagggg ggaatgaaag accccacctg taggtttggc
     3301 aagctagctt aagtaacgcc attttgcaag gcatggaaaa tacataactg agaatagaga
     3361 agttcagatc aaggttagga acagagagac agcagaatat gggccaaaca ggatatctgt
     3421 ggtaagcagt tcctgccccg gctcagggcc aagaacagat ggtccccaga tgcggtcccg
     3481 ccctcagcag tttctagaga accatcagat gtttccaggg tgccccaagg acctgaaatg
     3541 accctgtgcc ttatttgaac taaccaatca gttcgcttct cgcttctgtt cgcgcgcttc
     3601 tgctccccga gctcaataaa agagcccaca acccctcact cggcgcgcca gtcctccgat
     3661 agactgcgtc gcccgggtac ccgtgtatcc aataaaccct cttgcagttg catccgactt
     3721 gtggtctcgc tgttccttgg gagggtctcc tctgagtgat tgactacccg tcagcggggg
     3781 tctttcatgg gtaacagttt cttgaagttg gagaacaaca ttctgagggt aggagtcgaa
     3841 tattaagtaa tcctgactca attagccact gttttgaatc cacatactcc aatactcctg
     3901 aaatagttca ttatggacag cgcagaagag ctggggagaa ttaattcgta atcatggtca
     3961 tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat acgagccgga
     4021 agcataaagt gtaaagcctg gggtgcctaa tgagtgagct aactcacatt aattgcgttg
     4081 cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc
     4141 caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc gctcactgac
     4201 tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata
     4261 cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa
     4321 aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct
     4381 gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa
     4441 agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg
     4501 cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca
     4561 cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa
     4621 ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg
     4681 gtaagacacg acttatcgcc actggcagca gccactggta acaggattag cagagcgagg
     4741 tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta cactagaagg
     4801 acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc
     4861 tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag
     4921 attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac
     4981 gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc
     5041 ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag
     5101 taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt
     5161 ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag
     5221 ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca
     5281 gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact
     5341 ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca
     5401 gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg
     5461 tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc
     5521 atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg
     5581 gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca
     5641 tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt
     5701 atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc
     5761 agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc
     5821 ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca
     5881 tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa
     5941 aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat
     6001 tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa
     6061 aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctga cgtctaagaa
     6121 accattatta tcatgacatt aacctataaa aataggcgta tcacgaggcc ctttcgtctc
     6181 gcgcgtttcg gtgatgacgg tgaaaacctc tgacacatgc agctcccgga gacggtcaca
     6241 gcttgtctgt aagcggatgc cgggagcaga caagcccgtc agggcgcgtc agcgggtgtt
     6301 ggcgggtgtc ggggctggct taactatgcg gcatcagagc agattgtact gagagtgcac
     6361 catatgcggt gtgaaatacc gcacagatgc gtaaggagaa aataccgcat caggcgccat
     6421 tcgccattca ggctgcgcaa ctgttgggaa gggcgatcgg tgcgggcctc ttcgctatta
     6481 cgccagctgg cgaaaggggg atgtgctgca aggcgattaa gttgggtaac gccagggttt
     6541 tcccagtcac gacgttgtaa aacgacggcg caaggaatgg tgcatgcaag gagatggcgc
     6601 ccaacagtcc cccggccacg gggcctgcca ccatacccac gccgaaacaa gcgctcatga
     6661 gcccgaagtg gcgagcccga tcttccccat cggtgatgtc ggcgatatag gcgccagcaa
     6721 ccgcacctgt ggcgccggtg atgccggcca cgatgcgtcc ggcgtagagg cgattagtcc
     6781 aatttgttaa agacaggata tcagtggtcc aggctctagt tttgactcaa caatatcacc
     6841 agctgaagcc tatagagtac gagccataga taaaataaaa gattttattt agtctccaga
     6901 aaaagggggg aa
//
