LOCUS       sequence-178981-        8879 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 8879)
  AUTHORS   Reya T, Duncan AW, Ailles L, Domen J, Scherer DC, Willert K, Hintz 
            L, Nusse R, Weissman IL
  TITLE     A role for Wnt signalling in self-renewal of haematopoietic stem 
            cells.
  JOURNAL   Nature. 2003 May 22. 423(6938):409-14.
  PUBMED    12717450
REFERENCE   2  (bases 1 to 8879)
  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. 
            2003 May 22. 423(6938):409-14."
COMMENT     Sequence Label: MSCV-beta-catenin-IRES-GFP
FEATURES             Location/Qualifiers
     source          1..8879
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     LTR             40..556
                     /label=5' LTR
                     /note="5' long terminal repeat from murine embryonic stem 
                     cell virus"
     misc_feature    620..961
                     /label=MESV Psi
                     /note="packaging signal of murine embryonic stem cell 
                     virus"
     CDS             1028..1444
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     1374..1396
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     1412..1428
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     regulatory      1452..1461
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1503..1538
                     /codon_start=1
                     /product="nanobody epitope tag corresponding to amino acids
                     16-27 of human beta-catenin (Traenkle et al., 2015)"
                     /label=BC2 tag
                     /translation="PDRKAAVSHWQQ"
     misc_feature    3880..4429
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(4024..4041)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     4251..4270
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             join(4432..4434,4435..4437,4438..5151)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(4432..4434,4435..4437,4438..5148)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(4477..4498)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(4738..4757)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     5085..5106
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     LTR             5313..5827
                     /label=3' LTR
                     /note="3' long terminal repeat from murine embryonic stem 
                     cell virus"
     protein_bind    5989..6005
                     /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(6013..6019,6020..6037,6038..6043))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6058..6079
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6196..6213)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6367..6955)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6447..6466)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7126..7917,7918..7986))
                     /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     7749..7768
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7987..8091)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     8159..8177
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(8215..8237)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     8337..8356
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     8550..8572
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     8700..8719
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 aaaataaaag attttattta gtctccagaa aaagggggga atgaaagacc ccacctgtag
       61 gtttggcaag ctagcttaag taacgccatt ttgcaaggca tggaaaatac ataactgaga
      121 atagagaagt tcagatcaag gttaggaaca gagagacagc agaatatggg ccaaacagga
      181 tatctgtggt aagcagttcc tgccccggct cagggccaag aacagatggt ccccagatgc
      241 ggtcccgccc tcagcagttt ctagagaacc atcagatgtt tccagggtgc cccaaggacc
      301 tgaaatgacc ctgtgcctta tttgaactaa ccaatcagtt cgcttctcgc ttctgttcgc
      361 gcgcttctgc tccccgagct caataaaaga gcccacaacc cctcactcgg cgcgccagtc
      421 ctccgataga ctgcgtcgcc cgggtacccg tattcccaat aaagcctctt gctgtttgca
      481 tccgaatcgt ggactcgctg atccttggga gggtctcctc agattgattg actgcccacc
      541 tcgggggtct ttcatttgga ggttccaccg agatttggag acccctgccc agggaccacc
      601 gacccccccg ccgggaggta agctggccag cggtcgtttc gtgtctgtct ctgtctttgt
      661 gcgtgtttgt gccggcatct aatgtttgcg cctgcgtctg tactagttag ctaactagct
      721 ctgtatctgg cggacccgtg gtggaactga cgagttctga acacccggcc gcaaccctgg
      781 gagacgtccc agggactttg ggggccgttt ttgtggcccg acctgaggaa gggagtcgat
      841 gtggaatccg accccgtcag gatatgtggt tctggtagga gacgagaacc taaaacagtt
      901 cccgcctccg tctgaatttt tgctttcggt ttggaaccga agccgcgcgt cttgtctgct
      961 gcagcgctgc agcatcgttc tgtgttgtct ctgtctgact gtgtttctgt atttgtctga
     1021 aaattagggc cagactgtta ccactccctt aagtttgacc ttaggtcact ggaaagatgt
     1081 cgagcggatc gctcacaacc agtcggtaga tgtcaagaag agacgttggg ttaccttctg
     1141 ctctgcagaa tggccaacct ttaacgtcgg atggccgcga gacggcacct ttaaccgaga
     1201 cctcatcacc caggttaaga tcaaggtctt ttcacctggc ccgcatggac acccagacca
     1261 ggtcccctac atcgtgacct gggaagcctt ggcttttgac ccccctccct gggtcaagcc
     1321 ctttgtacac cctaagcctc cgcctcctct tcctccatcc gccccgtctc tcccccttga
     1381 acctcctcgt tcgaccccgc ctcgatcctc cctttatcca gccctcactc cttctctagg
     1441 cgccggaatt ggccaccatg gctactcaag ctgacctgat ggagttggac atggccatgg
     1501 agccggacag aaaagctgct gtcagccact ggcagcagca gtcttacttg gatgctggaa
     1561 tccatgctgg tgccaccgcc acagctcctg ccctgagtgg caagggcaac cctgaggaag
     1621 aagatgttga cacctcccaa gtcctttatg aatgggagca aggcttttcc cagtccttca
     1681 cgcaagagca agtagctgat attgacgggc agtatgcaat gactagggct cagagggtcc
     1741 gagctgccat gttccctgag acgctagatg agggcatgca gatcccatcc acgcagtttg
     1801 acgctgctca tcccactaat gtccagcgct tggctgaacc atcacagatg ttgaaacatg
     1861 cagttgtcaa tttgattaac tatcaggatg acgcggaact tgccacacgt gcaattcctg
     1921 agctgacaaa actgctaaac gatgaggacc aggtggtagt taataaagct gctgttatgg
     1981 tccatcagct ttccaaaaag gaagcttcca gacatgccat catgcgctcc cctcagatgg
     2041 tgtctgccat tgtacgcacc atgcagaata caaatgatgt agagacagct cgttgtactg
     2101 ctgggactct gcacaacctt tctcaccacc gcgagggctt gctggccatc tttaagtctg
     2161 gtggcatccc agcgctggtg aaaatgcttg ggtcaccagt ggattctgta ctgttctacg
     2221 ccatcacgac actgcataat ctcctgctcc atcaggaagg agctaaaatg gcagtgcgcc
     2281 tagctggtgg actgcagaaa atggttgctt tgctcaacaa aacaaacgtg aaattcttgg
     2341 ctattacaac agactgcctt cagatcttag cttatggcaa tcaagagagc aagctcatca
     2401 ttctggccag tggtggaccc caagccttag taaacataat gaggacctac acttatgaga
     2461 agcttctgtg gaccacaagc agagtgctga aagtgctgtc tgtctgctct agcaacaagc
     2521 cggccattgt agaagctggt gggatgcagg cactggggct tcatctgaca gacccaagtc
     2581 agcgacttgt tcaaaactgt ctttggactc tcagaaacct ttcagatgca gcgactaagc
     2641 aggaagggat ggaaggcctc cttgggactc tagtgcagct tctgggttcc gatgatataa
     2701 atgtggtcac ctgtgcagct ggaattctct ctaacctcac ttgcaataat tacaaaaaca
     2761 agatgatggt gtgccaagtg ggtggcatag aggctcttgt acgcaccgtc cttcgtgctg
     2821 gtgacaggga agacatcact gagcctgcca tctgtgctct tcgtcatctg accagccggc
     2881 atcaggaagc cgagatggcc cagaatgccg ttcgccttca ttatggactg cctgttgtgg
     2941 ttaaactcct gcacccacca tcccactggc ctctgataaa ggcaactgtt ggattgattc
     3001 gaaaccttgc cctttgccca gcaaatcatg cgcctttgcg ggaacagggt gctattccac
     3061 gactagttca gctgcttgta cgagcacatc aggacaccca acggcgcacc tccatgggtg
     3121 gaacgcagca gcagtttgtg gagggcgtgc gcatggagga gatagtagaa gggtgtactg
     3181 gagctctcca catccttgct cgggacgttc acaaccggat tgtaatccga ggactcaata
     3241 ccattccatt gtttgtgcag ttgctttatt ctcccattga aaatatccaa agagtagctg
     3301 caggggtcct ctgtgaactt gctcaggaca aggaggctgc agaggccatt gaagctgagg
     3361 gagccacagc tcccctgaca gagttactcc actccaggaa tgaaggcgtg gcaacatacg
     3421 cagctgctgt cctattccga atgtctgagg acaagccaca ggattacaag aagcggcttt
     3481 cagtcgagct gaccagttcc ctcttcagga cagagccaat ggcttggaat gagactgcag
     3541 atcttggact ggacattggt gcccagggag aagcccttgg atatcgccag gatgatccca
     3601 gctaccgttc ttttcactct ggtggatacg gccaggatgc cttggggatg gaccctatga
     3661 tggagcatga gatgggtggc caccaccctg gtgctgacta tccagttgat gggctgcctg
     3721 atctgggaca cgcccaggac ctcatggatg ggctgccccc aggtgatagc aatcagctgg
     3781 cctggtttga tactgacctg gatatcaagc ctcctactcc tcctcctgag cccgagacct
     3841 aatctagagt cgagcgggat caattccgcc ccccccctaa cgttactggc cgaagccgct
     3901 tggaataagg ccggtgtgcg tttgtctata tgttattttc caccatattg ccgtcttttg
     3961 gcaatgtgag ggcccggaaa cctggccctg tcttcttgac gagcattcct aggggtcttt
     4021 cccctctcgc caaaggaatg caaggtctgt tgaatgtcgt gaaggaagca gttcctctgg
     4081 aagcttcttg aagacaaaca acgtctgtag cgaccctttg caggcagcgg aaccccccac
     4141 ctggcgacag gtgcctctgc ggccaaaagc cacgtgtata agatacacct gcaaaggcgg
     4201 cacaacccca gtgccacgtt gtgagttgga tagttgtgga aagagtcaaa tggctctcct
     4261 caagcgtatt caacaagggg ctgaaggatg cccagaaggt accccattgt atgggatctg
     4321 atctggggcc tcggtgcaca tgctttacat gtgtttagtc gaggttaaaa aaacgtctag
     4381 gccccccgaa ccacggggac gtggttttcc tttgaaaaac acgataatac catggtgagc
     4441 aagggcgagg agctgttcac cggggtggtg cccatcctgg tcgagctgga cggcgacgta
     4501 aacggccaca agttcagcgt gtccggcgag ggcgagggcg atgccaccta cggcaagctg
     4561 accctgaagt tcatctgcac caccggcaag ctgcccgtgc cctggcccac cctcgtgacc
     4621 accctgacct acggcgtgca gtgcttcagc cgctaccccg accacatgaa gcagcacgac
     4681 ttcttcaagt ccgccatgcc cgaaggctac gtccaggagc gcaccatctt cttcaaggac
     4741 gacggcaact acaagacccg cgccgaggtg aagttcgagg gcgacaccct ggtgaaccgc
     4801 atcgagctga agggcatcga cttcaaggag gacggcaaca tcctggggca caagctggag
     4861 tacaactaca acagccacaa cgtctatatc atggccgaca agcagaagaa cggcatcaag
     4921 gtgaacttca agatccgcca caacatcgag gacggcagcg tgcagctcgc cgaccactac
     4981 cagcagaaca cccccatcgg cgacggcccc gtgctgctgc ccgacaacca ctacctgagc
     5041 acccagtccg ccctgagcaa agaccccaac gagaagcgcg atcacatggt cctgctggag
     5101 ttcgtgaccg ccgccgggat cactctcggc atggacgagc tgtacaagta aagcggccgc
     5161 gactctagag tcgacctgca ggcatgcaag cttcaggtag ccggctaacg ttaacaaccg
     5221 gtacctctag aactatagct agcatgcgca aatttaaagc gctgatatcg ataaaataaa
     5281 agattttatt tagtctccag aaaaaggggg gaatgaaaga ccccacctgt aggtttggca
     5341 agctagctta agtaacgcca ttttgcaagg catggaaaat acataactga gaatagagaa
     5401 gttcagatca aggttaggaa cagagagaca gcagaatatg ggccaaacag gatatctgtg
     5461 gtaagcagtt cctgccccgg ctcagggcca agaacagatg gtccccagat gcggtcccgc
     5521 cctcagcagt ttctagagaa ccatcagatg tttccagggt gccccaagga cctgaaatga
     5581 ccctgtgcct tatttgaact aaccaatcag ttcgcttctc gcttctgttc gcgcgcttct
     5641 gctccccgag ctcaataaaa gagcccacaa cccctcactc ggcgcgccag tcctccgata
     5701 gactgcgtcg cccgggtacc cgtgtatcca ataaaccctc ttgcagttgc atccgacttg
     5761 tggtctcgct gttccttggg agggtctcct ctgagtgatt gactacccgt cagcgggggt
     5821 ctttcatggg taacagtttc ttgaagttgg agaacaacat tctgagggta ggagtcgaat
     5881 attaagtaat cctgactcaa ttagccactg ttttgaatcc acatactcca atactcctga
     5941 aatagttcat tatggacagc gcagaaagag ctggggagaa ttgtgaaatt gttatccgct
     6001 cacaattcca cacaacatac gagccggaag cataaagtgt aaagcctggg gtgcctaatg
     6061 agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct
     6121 gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg
     6181 gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc
     6241 ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg
     6301 aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct
     6361 ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca
     6421 gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct
     6481 cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc
     6541 gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt
     6601 tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc
     6661 cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc
     6721 cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg
     6781 gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc
     6841 agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag
     6901 cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga
     6961 tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat
     7021 tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag
     7081 ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat
     7141 cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc
     7201 cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat
     7261 accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag
     7321 ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg
     7381 ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc
     7441 tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca
     7501 acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg
     7561 tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc
     7621 actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta
     7681 ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc
     7741 aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg
     7801 ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc
     7861 cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc
     7921 aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat
     7981 actcatactc ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag
     8041 cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc
     8101 ccgaaaagtg ccacctgacg tctaagaaac cattattatc atgacattaa cctataaaaa
     8161 taggcgtatc acgaggccct ttcgtctcgc gcgtttcggt gatgacggtg aaaacctctg
     8221 acacatgcag ctcccggaga cggtcacagc ttgtctgtaa gcggatgccg ggagcagaca
     8281 agcccgtcag ggcgcgtcag cgggtgttgg cgggtgtcgg ggctggctta actatgcggc
     8341 atcagagcag attgtactga gagtgcacca tatgcggtgt gaaataccgc acagatgcgt
     8401 aaggagaaaa taccgcatca ggcgccattc gccattcagg ctgcgcaact gttgggaagg
     8461 gcgatcggtg cgggcctctt cgctattacg ccagctggcg aaagggggat gtgctgcaag
     8521 gcgattaagt tgggtaacgc cagggttttc ccagtcacga cgttgtaaaa cgacggcgca
     8581 aggaatggtg catgcaagga gatggcgccc aacagtcccc cggccacggg gcctgccacc
     8641 atacccacgc cgaaacaagc gctcatgagc ccgaagtggc gagcccgatc ttccccatcg
     8701 gtgatgtcgg cgatataggc gccagcaacc gcacctgtgg cgccggtgat gccggccacg
     8761 atgcgtccgg cgtagaggcg attagtccaa tttgttaaag acaggatatc agtggtccag
     8821 gctctagttt tgactcaaca atatcaccag ctgaagccta tagagtacga gccatagat
//
