LOCUS       sequence-349719-        7273 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression vector encoding POFUT2 and EmGFP-Bsd.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7273)
  AUTHORS   Moyer AJ, Fernandez FX, Li Y, Klinedinst DK, Florea LD, Kazuki Y, 
            Oshimura M, Reeves RH
  TITLE     Overexpression screen of chromosome 21 genes reveals modulators of 
            Sonic hedgehog signaling relevant to Down syndrome.
  JOURNAL   Dis Model Mech. 2023 Apr 1;16(4):dmm049712. doi: 10.1242/dmm.049712.
            Epub 2023 Apr 13.
  PUBMED    36995257
REFERENCE   2  (bases 1 to 7273)
  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: "Dis Model 
            Mech."; date: "2023-04-1"; volume: "16(4):dmm049712. doi"; pages: " 
            10.1242/dmm.049712. Epub 2023 Apr 13"
COMMENT     Sequence Label: POFUT2_pcDNA6.2/EmGFP-Bsd
FEATURES             Location/Qualifiers
     source          1..7273
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..204
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     155..175
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     249..268
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        249..267
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    297..321
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(1615..1639)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             1665..1706
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     primer_bind     complement(1746..1765)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    1790..1838
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      2040..2468
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2127..2146)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2337..2358
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        2483..2982
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(2518..2541)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     2915..2934
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     promoter        2996..3043
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             join(3062..3064,3065..3067,3068..3778)
                     /codon_start=1
                     /product="Emerald GFP"
                     /label=EmGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFTYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHKVYITADKQKNGIK
                     VNFKTRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(3107..3128)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3368..3387)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3715..3736
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             3779..4174
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="AKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTGV
                     NVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGIK
                     AIVKDSDGQPTAVGIRELLPSGYVWEG"
     polyA_signal    4332..4453
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4369..4388)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4423..4442
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4502..4518)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4502..4518)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4515..4537)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4526..4542
                     /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(4550..4556,4557..4574,4575..4580))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4595..4616
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4733..4750)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4904..5492)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4984..5003)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5663..6454,6455..6523))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPKLNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     6286..6305
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6524..6628)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6703..6722)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        6894..7273
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
ORIGIN
        1 gtgatgcggt tttggcagta catcaatggg cgtggatagc ggtttgactc acggggattt
       61 ccaagtctcc accccattga cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac
      121 tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag gcgtgtacgg
      181 tgggaggtct atataagcag agctctctgg ctaactagag aacccactgc ttactggctt
      241 atcgaaatta atacgactca ctatagggag acccaagctg gctagttaag ctatcaacaa
      301 gtttgtacaa aaaagcaggc accatggcga cactcagctt cgtcttcctg ctgctggggg
      361 cagtgtcctg gcctccggct tctgcctccg gccaggagtt ctggcccgga caatcggcgg
      421 ccgatattct gtcgggggcg gcttcccgca gacggtatct tctgtatgac gtcaaccccc
      481 cggaaggctt caacctgcgc agggatgtct atatccgaat cgcctctctc ctgaagactc
      541 tgctgaagac ggaggagtgg gtgcttgtcc tgcctccatg gggccgcctc tatcactggc
      601 agagtcctga catccaccag gtccggattc cctggtctga gttttttgat cttccaagtc
      661 tcaataaaaa catccccgtc atcgagtatg agcagttcat cgcagaatct ggtgggccct
      721 ttattgacca ggtttacgtc ctgcaaagtt acgcagaggg gtggaaagaa gggacctggg
      781 aagagaaggt ggacgagcgg ccgtgtattg atcagctcct gtactcccag gacaagcacg
      841 agtactacag aggatggttt tggggttatg aggagaccag gggtctaaac gtctcctgtc
      901 tgtccgtcca gggctcagcc tccatcgtgg cgcccctgct gctgagaaac acatcagccc
      961 ggtccgtgat gttagacaga gccgagaacc tacttcacga ccactatgga gggaaagaat
     1021 actgggatac ccgtcgcagc atggtgtttg ccaggcacct gcgggaggtg ggagacgagt
     1081 tcaggagcag acatctcaac tccacggacg acgcagacag gatccccttc caggaggact
     1141 ggatgaagat gaaggtcaag ctgggctccg cgctaggggg cccctacctg ggagtccacc
     1201 tgagaagaaa agatttcatc tggggtcaca gacaggatgt acccagtctg gaaggggccg
     1261 tgaggaagat ccgcagcctc atgaagaccc accggctgga caaggtgttt gtggccacag
     1321 atgccgtcag aaaggaatat gaagagctaa aaaagctgtt acccgagatg gtgaggtttg
     1381 aacccacgtg ggaggagctg gagctctaca aggacggagg cgttgcgatt attgaccagt
     1441 ggatctgcgc acacgccagg ttttttattg gcacctcagt ctcaacattt tcttttcgga
     1501 ttcatgagga aagagaaatc ctggggttgg accccaagac gacgtacaac aggttctgcg
     1561 gagaccaaga gaaggcgtgt gagcaaccca cccactggaa gatcacctac tagaacccag
     1621 ctttcttgta caaagtggtt gatctagagg gcccgcggtt cgaaggtaag cctatcccta
     1681 accctctcct cggtctcgat tctacgcgta ccggttagta atgagtttaa acgggggagg
     1741 ctaactgaaa cacggaagga gacaataccg gaaggaaccc gcgctatgac ggcaataaaa
     1801 agacagaata aaacgcacgg gtgttgggtc gtttgttcat aaacgcgggg ttcggtccca
     1861 gggctggcac tctgtcgata ccccaccgag accccattgg ggccaatacg cccgcgtttc
     1921 ttccttttcc ccaccccacc ccccaagttc gggtgaaggc ccagggctcg cagccaacgt
     1981 cggggcggca ggccctgcca tagcagatct gcgcagctgg ggctctaggg ggtatcccca
     2041 cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc
     2101 tacacttgcc agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac
     2161 gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag
     2221 tgctttacgg cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc
     2281 atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg
     2341 actcttgttc caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata
     2401 agggattttg ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa
     2461 cgcgaattat cctcgagcta ccgggtaggg gaggcgcttt tcccaaggca gtctggagca
     2521 tgcgctttag cagccccgct gggcacttgg cgctacacaa gtggcctctg gcctcgcaca
     2581 cattccacat ccaccggtag gcgccaaccg gctccgttct ttggtggccc cttcgcgcca
     2641 ccttctactc ctcccctagt caggaagttc ccccccgccc cgcagctcgc gtcgtgcagg
     2701 acgtgacaaa tggaagtagc acgtctcact agtctcgtgc agatggacag caccgctgag
     2761 caatggaagc gggtaggcct ttggggcagc ggccaatagc agctttgctc cttcgctttc
     2821 tgggctcaga ggctgggaag gggtgggtcc gggggcgggc tcaggggcgg gctcaggggc
     2881 ggggcgggcg cccgaaggtc ctccggaggc ccggcattct gcacgcttca aaagcgcacg
     2941 tctgccgcgc tgttctcctc ttcctcatct ccgggccttt cgactctaga cacgtgttga
     3001 caattaatca tcggcatagt atatcggcat agtataatac gacaaggtga ggaactaaac
     3061 catggtgagc aagggcgagg agctgttcac cggggtggtg cccatcctgg tcgagctgga
     3121 cggcgacgta aacggccaca agttcagcgt gtccggcgag ggcgagggcg atgccaccta
     3181 cggcaagctg accctgaagt tcatctgcac caccggcaag ctgcccgtgc cctggcccac
     3241 cctcgtgacc accttcacct acggcgtgca gtgcttcgcc cgctaccccg accacatgaa
     3301 gcagcacgac ttcttcaagt ccgccatgcc cgaaggctac gtccaggagc gcaccatctt
     3361 cttcaaggac gacggcaact acaagacccg cgccgaggtg aagttcgagg gcgacaccct
     3421 ggtgaaccgc atcgagctga agggcatcga cttcaaggag gacggcaaca tcctggggca
     3481 caagctggag tacaactaca acagccacaa ggtctatatc accgccgaca agcagaagaa
     3541 cggcatcaag gtgaacttca agacccgcca caacatcgag gacggcagcg tgcagctcgc
     3601 cgaccactac cagcagaaca cccccatcgg cgacggcccc gtgctgctgc ccgacaacca
     3661 ctacctgagc acccagtccg ccctgagcaa agaccccaac gagaagcgcg atcacatggt
     3721 cctgctggag ttcgtgaccg ccgccgggat cactctcggc atggacgagc tgtacaaggc
     3781 caagcctttg tctcaagaag aatccaccct cattgaaaga gcaacggcta caatcaacag
     3841 catccccatc tctgaagact acagcgtcgc cagcgcagct ctctctagcg acggccgcat
     3901 cttcactggt gtcaatgtat atcattttac tgggggacct tgtgcagaac tcgtggtgct
     3961 gggcactgct gctgctgcgg cagctggcaa cctgacttgt atcgtcgcga tcggaaatga
     4021 gaacaggggc atcttgagcc cctgcggacg gtgccgacag gtgcttctcg atctgcatcc
     4081 tgggatcaaa gccatagtga aggacagtga tggacagccg acggcagttg ggattcgtga
     4141 attgctgccc tctggttatg tgtgggaggg ctaagcactt cgtggccgag gagcaggact
     4201 gacacgtgct acgagatttc gattccaccg ccgccttcta tgaaaggttg ggcttcggaa
     4261 tcgttttccg ggacgccggc tggatgatcc tccagcgcgg ggatctcatg ctggagttct
     4321 tcgcccaccc caacttgttt attgcagctt ataatggtta caaataaagc aatagcatca
     4381 caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca
     4441 tcaatgtatc ttatcatgtc tgtataccgt cgacctctag ctagagcttg gcgtaatcat
     4501 ggtcatagct gtttcctgtg tgaaattgtt atccgctcac aattccacac aacatacgag
     4561 ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt gagctaactc acattaattg
     4621 cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa
     4681 tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca
     4741 ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg
     4801 taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc
     4861 agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc
     4921 cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac
     4981 tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc
     5041 tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata
     5101 gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc
     5161 acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca
     5221 acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag
     5281 cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta
     5341 gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg
     5401 gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc
     5461 agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt
     5521 ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa
     5581 ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat
     5641 atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga
     5701 tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac
     5761 gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg
     5821 ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg
     5881 caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt
     5941 cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct
     6001 cgtcgtttgg tatggcttca ttcagcttcg gttcccaacg atcaaggcga gttacatgat
     6061 cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta
     6121 agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca
     6181 tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat
     6241 agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac
     6301 atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa
     6361 ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt
     6421 cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg
     6481 caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat
     6541 attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt
     6601 agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtcg
     6661 acggatcggg agatctcccg atcccctatg gtgcactctc agtacaatct gctctgatgc
     6721 cgcatagtta agccagtatc tgctccctgc ttgtgtgttg gaggtcgctg agtagtgcgc
     6781 gagcaaaatt taagctacaa caaggcaagg cttgaccgac aattgcatga agaatctgct
     6841 tagggttagg cgttttgcgc tgcttcgcga tgtacgggcc agatatacgc gttgacattg
     6901 attattgact agttattaat agtaatcaat tacggggtca ttagttcata gcccatatat
     6961 ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc ccaacgaccc
     7021 ccgcccattg acgtcaataa tgacgtatgt tcccatagta acgccaatag ggactttcca
     7081 ttgacgtcaa tgggtggagt atttacggta aactgcccac ttggcagtac atcaagtgta
     7141 tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg cctggcatta
     7201 tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg tattagtcat
     7261 cgctattacc atg
//
