LOCUS       sequence-450572-        8157 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Less commonly used plasmid for generating ER-Hoxb8 conditionally 
            immortalized myeloid cell lines..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8157)
  AUTHORS   Wang GG, Calvo KR, Pasillas MP, Sykes DB, Hacker H, Kamps MP
  TITLE     Quantitative production of macrophages or neutrophils ex vivo using 
            conditional Hoxb8.
  JOURNAL   Nat Methods. 2006 Apr;3(4):287-93.
  PUBMED    16554834
REFERENCE   2  (bases 1 to 8157)
  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 
            Methods."; date: "2006-04"; volume: "3(4)"; pages: "287-93"
COMMENT     Sequence Label: MSCV-Flag-ER-Hoxb8-IRES-GFP
FEATURES             Location/Qualifiers
     source          1..8157
                     /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"
     regulatory      1422..1431
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1431..1454
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             1467..2402
                     /codon_start=1
                     /product="mutated ligand-binding domain of the human 
                     estrogen receptor (Feil et al., 1997)"
                     /label=ERT2
                     /translation="AGDMRAANLWPSPLMIKRSKKNSLALSLTADQMVSALLDAEPPIL
                     YSEYDPTRPFSEASMMGLLTNLADRELVHMINWAKRVPGFVDLTLHDQVHLLECAWLEI
                     LMIGLVWRSMEHPVKLLFAPNLLLDRNQGKCVEGMVEIFDMLLATSSRFRMMNLQGEEF
                     VCLKSIILLNSGVYTFLSSTLKSLEEKDHIHRVLDKITDTLIHLMAKAGLTLQQQHQRL
                     AQLLLILSHIRHMSNKGMEHLYSMKCKNVVPLYDLLLEMLDAHRLHAPTSRGGASVEET
                     DQSHLATAGSTSSHSLQKYYITGEAEGFPAT"
     CDS             1515..2405
                     /codon_start=1
                     /gene="ESR1"
                     /product="estrogen nuclear receptor alpha ligand-binding 
                     domain"
                     /label=ER-LBD
                     /translation="KRSKKNSLALSLTADQMVSALLDAEPPILYSEYDPTRPFSEASMM
                     GLLTNLADRELVHMINWAKRVPGFVDLTLHDQVHLLECAWLEILMIGLVWRSMEHPVKL
                     LFAPNLLLDRNQGKCVEGMVEIFDMLLATSSRFRMMNLQGEEFVCLKSIILLNSGVYTF
                     LSSTLKSLEEKDHIHRVLDKITDTLIHLMAKAGLTLQQQHQRLAQLLLILSHIRHMSNK
                     GMEHLYSMKCKNVVPLYDLLLEMLDAHRLHAPTSRGGASVEETDQSHLATAGSTSSHSL
                     QKYYITGEAEGFPATV"
     misc_feature    3148..3715
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     misc_feature    3165..3717
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(3309..3326)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     3536..3555
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             join(3729..3731,3732..3734,3735..4448)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     ANFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(3774..3795)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(4035..4054)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4382..4403
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     LTR             4550..5064
                     /label=3' LTR
                     /note="3' long terminal repeat from murine embryonic stem 
                     cell virus"
     protein_bind    5226..5242
                     /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(5250..5256,5257..5274,5275..5280))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5295..5316
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5433..5450)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5604..6192)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5684..5703)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6363..7154,7155..7223))
                     /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     6986..7005
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7224..7328)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7396..7414
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(7452..7474)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     7574..7593
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7787..7809
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     7937..7956
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     LTR             join(8156..8157,1..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 ggccaccatg gactacaagg
     1441 acgacgatga caaaggaacg cgttctgctg gagacatgag agctgccaac ctttggccaa
     1501 gcccgctcat gatcaaacgc tctaagaaga acagcctggc cttgtccctg acggccgacc
     1561 agatggtcag tgccttgttg gatgctgagc cccccatact ctattccgag tatgatccta
     1621 ccagaccctt cagtgaagct tcgatgatgg gcttactgac caacctggca gacagggagc
     1681 tggttcacat gatcaactgg gcgaagaggg tgccaggctt tgtggatttg accctccatg
     1741 atcaggtcca ccttctagaa tgtgcctggc tagagatcct gatgattggt ctcgtctggc
     1801 gctccatgga gcacccagtg aagctactgt ttgctcctaa cttgctcttg gacaggaacc
     1861 agggaaaatg tgtagagggc atggtggaga tcttcgacat gctgctggct acatcatctc
     1921 ggttccgcat gatgaatctg cagggagagg agtttgtgtg cctcaaatct attattttgc
     1981 ttaattctgg agtgtacaca tttctgtcca gcaccctgaa gtctctggaa gagaaggacc
     2041 atatccaccg agtcctggac aagatcacag acactttgat ccacctgatg gccaaggcag
     2101 gcctgaccct gcagcagcag caccagcggc tggcccagct cctcctcatc ctctcccaca
     2161 tcaggcacat gagtaacaaa ggcatggagc atctgtacag catgaagtgc aagaacgtgg
     2221 tgcccctcta tgacctgctg ctggagatgc tggacgccca ccgcctacat gcgcccacta
     2281 gccgtggagg ggcatccgtg gaggagacgg accaaagcca cttggccact gcgggctcta
     2341 cttcatcgca ttccttgcaa aagtattaca tcacggggga ggcagagggt ttccctgcca
     2401 cagtcacgcg tggaagctct tatttcgtca actcactgtt ctccaaatac aaaaccgggg
     2461 agtccctgcg ccccaattat tatgactgcg gcttcgccca ggacctgggc ggccgaccca
     2521 ccgtggtgta cggtcccagc agcggcggca gcttccagca cccttcgcaa atccaggagt
     2581 tctaccacgg gccatcgtcg ctgtccacag ctccctacca gcagaacccg tgcgccgtgg
     2641 cgtgccacgg cgaccccggc aacttctacg gctacgaccc tctgcagcgc cagagcctgt
     2701 tcggtgcgca ggatccagac ctggtgcagt acgcagactg caagctcgcg gcagccagcg
     2761 gcctgggcga ggaggccgag gggtctgagc agagcccgtc gcccacacag ctctttccct
     2821 ggatgcgccc tcaagccgcc ggacgcaggc gaggccgcca gacctacagt cgctaccaga
     2881 ccctggagct ggagaaggag ttcctattta atccctatct gactcgcaag cggaggatcg
     2941 aggtatcgca cgcgctggga ctgacagaga gacaggtcaa aatctggttc cagaatcgga
     3001 gaatgaagtg gaaaaaggag aacaacaaag acaagtttcc cagcagtaaa tgcgagcagg
     3061 aggagctgga gaaagagaag ctggagcggg caccagagac cgccgagcag ggcgatgcgc
     3121 agaagggtga caagaagtag taagaattcc gccccccccc cctaacgtta ctggccgaag
     3181 ccgcttggaa taaggccggt gtgcgtttgt ctatatgtta ttttccacca tattgccgtc
     3241 ttttggcaat gtgagggccc ggaaacctgg ccctgtcttc ttgacgagca ttcctagggg
     3301 tctttcccct ctcgccaaag gaatgcaagg tctgttgaat gtcgtgaagg aagcagttcc
     3361 tctggaagct tcttgaagac aaacaacgtc tgtagcgacc ctttgcaggc agcggaaccc
     3421 cccacctggc gacaggtgcc tctgcggcca aaagccacgt gtataagata cacctgcaaa
     3481 ggcggcacaa ccccagtgcc acgttgtgag ttggatagtt gtggaaagag tcaaatggct
     3541 ctcctcaagc gtattcaaca aggggctgaa ggatgcccag aaggtacccc attgtatggg
     3601 atctgatctg gggcctcggt gcacatgctt tacatgtgtt tagtcgaggt taaaaaaacg
     3661 tctaggcccc ccgaaccacg gggacgtggt tttcctttga aaaacacgat gataatatgg
     3721 ccacaaccat ggtgagcaag ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg
     3781 agctggacgg cgacgtaaac ggccacaagt tcagcgtgtc tggcgagggc gagggcgatg
     3841 ccacctacgg caagctgacc ctgaagttca tctgcaccac cggcaagctg cccgtgccct
     3901 ggcccaccct cgtgaccacc ctgacctacg gcgtgcagtg cttcagccgc taccccgacc
     3961 acatgaagca gcacgacttc ttcaagtccg ccatgcccga aggctacgtc caggagcgca
     4021 ccatcttctt caaggacgac ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg
     4081 acaccctggt gaaccgcatc gagctgaagg gcatcgactt caaggaggac ggcaacatcc
     4141 tggggcacaa gctggagtac aactacaaca gccacaacgt ctatatcatg gccgacaagc
     4201 agaagaacgg catcaaggcg aacttcaaga tccgccacaa catcgaggac ggcagcgtgc
     4261 agctcgccga ccactaccag cagaacaccc ccatcggcga cggccccgtg ctgctgcccg
     4321 acaaccacta cctgagcacc cagtccgccc tgagcaaaga ccccaacgag aagcgcgatc
     4381 acatggtcct gctggagttc gtgaccgccg ccgggatcac tctcggcatg gacgagctgt
     4441 acaagtaatg aattaattaa gaattatcaa gcttatcgat accgtcgacc tgcagccaag
     4501 cttatcgata aaataaaaga ttttatttag tctccagaaa aaggggggaa tgaaagaccc
     4561 cacctgtagg tttggcaagc tagcttaagt aacgccattt tgcaaggcat ggaaaataca
     4621 taactgagaa tagagaagtt cagatcaagg ttaggaacag agagacagca gaatatgggc
     4681 caaacaggat atctgtggta agcagttcct gccccggctc agggccaaga acagatggtc
     4741 cccagatgcg gtcccgccct cagcagtttc tagagaacca tcagatgttt ccagggtgcc
     4801 ccaaggacct gaaatgaccc tgtgccttat ttgaactaac caatcagttc gcttctcgct
     4861 tctgttcgcg cgcttctgct ccccgagctc aataaaagag cccacaaccc ctcactcggc
     4921 gcgccagtcc tccgatagac tgcgtcgccc gggtacccgt gtatccaata aaccctcttg
     4981 cagttgcatc cgacttgtgg tctcgctgtt ccttgggagg gtctcctctg agtgattgac
     5041 tacccgtcag cgggggtctt tcatgggtaa cagtttcttg aagttggaga acaacattct
     5101 gagggtagga gtcgaatatt aagtaatcct gactcaatta gccactgttt tgaatccaca
     5161 tactccaata ctcctgaaat agttcattat ggacagcgca gaaagagctg gggagaattg
     5221 tgaaattgtt atccgctcac aattccacac aacatacgag ccggaagcat aaagtgtaaa
     5281 gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct
     5341 ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga
     5401 ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc
     5461 gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa
     5521 tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt
     5581 aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa
     5641 aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt
     5701 ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg
     5761 tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc
     5821 agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc
     5881 gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta
     5941 tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct
     6001 acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc
     6061 tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa
     6121 caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa
     6181 aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa
     6241 aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt
     6301 ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac
     6361 agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc
     6421 atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt accatctggc
     6481 cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata
     6541 aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc
     6601 cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc
     6661 aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca
     6721 ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa
     6781 gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca
     6841 ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt
     6901 tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt
     6961 tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg
     7021 ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga
     7081 tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc
     7141 agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg
     7201 acacggaaat gttgaatact catactcttc ctttttcaat attattgaag catttatcag
     7261 ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg
     7321 gttccgcgca catttccccg aaaagtgcca cctgacgtct aagaaaccat tattatcatg
     7381 acattaacct ataaaaatag gcgtatcacg aggccctttc gtctcgcgcg tttcggtgat
     7441 gacggtgaaa acctctgaca catgcagctc ccggagacgg tcacagcttg tctgtaagcg
     7501 gatgccggga gcagacaagc ccgtcagggc gcgtcagcgg gtgttggcgg gtgtcggggc
     7561 tggcttaact atgcggcatc agagcagatt gtactgagag tgcaccatat gcggtgtgaa
     7621 ataccgcaca gatgcgtaag gagaaaatac cgcatcaggc gccattcgcc attcaggctg
     7681 cgcaactgtt gggaagggcg atcggtgcgg gcctcttcgc tattacgcca gctggcgaaa
     7741 gggggatgtg ctgcaaggcg attaagttgg gtaacgccag ggttttccca gtcacgacgt
     7801 tgtaaaacga cggcgcaagg aatggtgcat gcaaggagat ggcgcccaac agtcccccgg
     7861 ccacggggcc tgccaccata cccacgccga aacaagcgct catgagcccg aagtggcgag
     7921 cccgatcttc cccatcggtg atgtcggcga tataggcgcc agcaaccgca cctgtggcgc
     7981 cggtgatgcc ggccacgatg cgtccggcgt agaggcgatt agtccaattt gttaaagaca
     8041 ggatatcagt ggtccaggct ctagttttga ctcaacaata tcaccagctg aagcctatag
     8101 agtacgagcc atagataaaa taaaagattt tatttagtct ccagaaaaag gggggaa
//
