LOCUS       sequence-349696-        7540 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression vector encoding BACE2 and EmGFP-Bsd.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7540)
  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 7540)
  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: BACE2_pcDNA6.2/EmGFP-Bsd
FEATURES             Location/Qualifiers
     source          1..7540
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..584
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     629..648
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        629..647
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    677..701
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(2262..2286)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             2312..2353
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     primer_bind     complement(2393..2412)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    2437..2485
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      2687..3115
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2774..2793)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2984..3005
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        3130..3629
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(3165..3188)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     3562..3581
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     promoter        3643..3690
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             join(3709..3711,3712..3714,3715..4425)
                     /codon_start=1
                     /product="Emerald GFP"
                     /label=EmGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFTYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHKVYITADKQKNGIK
                     VNFKTRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(3754..3775)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(4015..4034)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4362..4383
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             4426..4821
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="AKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTGV
                     NVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGIK
                     AIVKDSDGQPTAVGIRELLPSGYVWEG"
     polyA_signal    4979..5100
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5016..5035)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5070..5089
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5149..5165)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5149..5165)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5162..5184)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5173..5189
                     /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(5197..5203,5204..5221,5222..5227))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5242..5263
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5380..5397)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5551..6139)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5631..5650)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6310..7101,7102..7170))
                     /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     6933..6952
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7171..7275)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(7350..7369)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctctctgg ctaactagag
      601 aacccactgc ttactggctt atcgaaatta atacgactca ctatagggag acccaagctg
      661 gctagttaag ctatcaacaa gtttgtacaa aaaagcaggc accatgggcg cactggcccg
      721 ggcgctgctg ctgcctctgc tggcccagtg gctcctgcgc gccgccccgg agctggcccc
      781 cgcgcccttc acgctgcccc tccgggtggc cgcggccacg aaccgcgtag ttgcgcccac
      841 cccgggaccc gggacccctg ccgagcgcca cgccgacggc ttggcgctcg ccctggagcc
      901 tgccctggcg tcccccgcgg gcgccgccaa cttcttggcc atggtagaca acctgcaggg
      961 ggactctggc cgcggctact acctggagat gctgatcggg acccccccgc agaagctaca
     1021 gattctcgtt gacactggaa gcagtaactt tgccgtggca ggaaccccgc actcctacat
     1081 agacacgtac tttgacacag agaggtctag cacataccgc tccaagggct ttgacgtcac
     1141 agtgaagtac acacaaggaa gctggacggg cttcgttggg gaagacctcg tcaccatccc
     1201 caaaggcttc aatacttctt ttcttgtcaa cattgccact atttttgaat cagagaattt
     1261 ctttttgcct gggattaaat ggaatggaat acttggccta gcttatgcca cacttgccaa
     1321 gccatcaagt tctctggaga ccttcttcga ctccctggtg acacaagcaa acatccccaa
     1381 cgttttctcc atgcagatgt gtggagccgg cttgcccgtt gctggatctg ggaccaacgg
     1441 aggtagtctt gtcttgggtg gaattgaacc aagtttgtat aaaggagaca tctggtatac
     1501 ccctattaag gaagagtggt actaccagat agaaattctg aaattggaaa ttggaggcca
     1561 aagccttaat ctggactgca gagagtataa cgcagacaag gccatcgtgg acagtggcac
     1621 cacgctgctg cgcctgcccc agaaggtgtt tgatgcggtg gtggaagctg tggcccgcgc
     1681 atctctgatt ccagaattct ctgatggttt ctggactggg tcccagctgg cgtgctggac
     1741 gaattcggaa acaccttggt cttacttccc taaaatctcc atctacctga gagatgagaa
     1801 ctccagcagg tcattccgta tcacaatcct gcctcagctt tacattcagc ccatgatggg
     1861 ggccggcctg aattatgaat gttaccgatt cggcatttcc ccatccacaa atgcgctggt
     1921 gatcggtgcc acggtgatgg agggcttcta cgtcatcttc gacagagccc agaagagggt
     1981 gggcttcgca gcgagcccct gtgcagaaat tgcaggtgct gcagtgtctg aaatttccgg
     2041 gcctttctca acagaggatg tagccagcaa ctgtgtcccc gctcagtctt tgagcgagcc
     2101 cattttgtgg attgtgtcct atgcgctcat gagcgtctgt ggagccatcc tccttgtctt
     2161 aatcgtcctg ctgctgctgc cgttccggtg tcagcgtcgc ccccgtgacc ctgaggtcgt
     2221 caatgatgag tcctctctgg tcagacatcg ctggaaatag aacccagctt tcttgtacaa
     2281 agtggttgat ctagagggcc cgcggttcga aggtaagcct atccctaacc ctctcctcgg
     2341 tctcgattct acgcgtaccg gttagtaatg agtttaaacg ggggaggcta actgaaacac
     2401 ggaaggagac aataccggaa ggaacccgcg ctatgacggc aataaaaaga cagaataaaa
     2461 cgcacgggtg ttgggtcgtt tgttcataaa cgcggggttc ggtcccaggg ctggcactct
     2521 gtcgataccc caccgagacc ccattggggc caatacgccc gcgtttcttc cttttcccca
     2581 ccccaccccc caagttcggg tgaaggccca gggctcgcag ccaacgtcgg ggcggcaggc
     2641 cctgccatag cagatctgcg cagctggggc tctagggggt atccccacgc gccctgtagc
     2701 ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc
     2761 gccctagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt
     2821 ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac
     2881 ctcgacccca aaaaacttga ttagggtgat ggttcacgta gtgggccatc gccctgatag
     2941 acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa
     3001 actggaacaa cactcaaccc tatctcggtc tattcttttg atttataagg gattttgccg
     3061 atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattatcct
     3121 cgagctaccg ggtaggggag gcgcttttcc caaggcagtc tggagcatgc gctttagcag
     3181 ccccgctggg cacttggcgc tacacaagtg gcctctggcc tcgcacacat tccacatcca
     3241 ccggtaggcg ccaaccggct ccgttctttg gtggcccctt cgcgccacct tctactcctc
     3301 ccctagtcag gaagttcccc cccgccccgc agctcgcgtc gtgcaggacg tgacaaatgg
     3361 aagtagcacg tctcactagt ctcgtgcaga tggacagcac cgctgagcaa tggaagcggg
     3421 taggcctttg gggcagcggc caatagcagc tttgctcctt cgctttctgg gctcagaggc
     3481 tgggaagggg tgggtccggg ggcgggctca ggggcgggct caggggcggg gcgggcgccc
     3541 gaaggtcctc cggaggcccg gcattctgca cgcttcaaaa gcgcacgtct gccgcgctgt
     3601 tctcctcttc ctcatctccg ggcctttcga ctctagacac gtgttgacaa ttaatcatcg
     3661 gcatagtata tcggcatagt ataatacgac aaggtgagga actaaaccat ggtgagcaag
     3721 ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg agctggacgg cgacgtaaac
     3781 ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg ccacctacgg caagctgacc
     3841 ctgaagttca tctgcaccac cggcaagctg cccgtgccct ggcccaccct cgtgaccacc
     3901 ttcacctacg gcgtgcagtg cttcgcccgc taccccgacc acatgaagca gcacgacttc
     3961 ttcaagtccg ccatgcccga aggctacgtc caggagcgca ccatcttctt caaggacgac
     4021 ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg acaccctggt gaaccgcatc
     4081 gagctgaagg gcatcgactt caaggaggac ggcaacatcc tggggcacaa gctggagtac
     4141 aactacaaca gccacaaggt ctatatcacc gccgacaagc agaagaacgg catcaaggtg
     4201 aacttcaaga cccgccacaa catcgaggac ggcagcgtgc agctcgccga ccactaccag
     4261 cagaacaccc ccatcggcga cggccccgtg ctgctgcccg acaaccacta cctgagcacc
     4321 cagtccgccc tgagcaaaga ccccaacgag aagcgcgatc acatggtcct gctggagttc
     4381 gtgaccgccg ccgggatcac tctcggcatg gacgagctgt acaaggccaa gcctttgtct
     4441 caagaagaat ccaccctcat tgaaagagca acggctacaa tcaacagcat ccccatctct
     4501 gaagactaca gcgtcgccag cgcagctctc tctagcgacg gccgcatctt cactggtgtc
     4561 aatgtatatc attttactgg gggaccttgt gcagaactcg tggtgctggg cactgctgct
     4621 gctgcggcag ctggcaacct gacttgtatc gtcgcgatcg gaaatgagaa caggggcatc
     4681 ttgagcccct gcggacggtg ccgacaggtg cttctcgatc tgcatcctgg gatcaaagcc
     4741 atagtgaagg acagtgatgg acagccgacg gcagttggga ttcgtgaatt gctgccctct
     4801 ggttatgtgt gggagggcta agcacttcgt ggccgaggag caggactgac acgtgctacg
     4861 agatttcgat tccaccgccg ccttctatga aaggttgggc ttcggaatcg ttttccggga
     4921 cgccggctgg atgatcctcc agcgcgggga tctcatgctg gagttcttcg cccaccccaa
     4981 cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa atttcacaaa
     5041 taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca atgtatctta
     5101 tcatgtctgt ataccgtcga cctctagcta gagcttggcg taatcatggt catagctgtt
     5161 tcctgtgtga aattgttatc cgctcacaat tccacacaac atacgagccg gaagcataaa
     5221 gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca ttaattgcgt tgcgctcact
     5281 gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc
     5341 ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg
     5401 ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc
     5461 cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag
     5521 gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca
     5581 tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca
     5641 ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg
     5701 atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag
     5761 gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt
     5821 tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca
     5881 cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg
     5941 cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa gaacagtatt
     6001 tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc
     6061 cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg
     6121 cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg
     6181 gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta
     6241 gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg
     6301 gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg
     6361 ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc
     6421 atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc cagatttatc
     6481 agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc
     6541 ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag
     6601 tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat
     6661 ggcttcattc agcttcggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg
     6721 caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt
     6781 gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag
     6841 atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg
     6901 accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt
     6961 aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct
     7021 gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac
     7081 tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat
     7141 aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat
     7201 ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca
     7261 aataggggtt ccgcgcacat ttccccgaaa agtgccacct gacgtcgacg gatcgggaga
     7321 tctcccgatc ccctatggtg cactctcagt acaatctgct ctgatgccgc atagttaagc
     7381 cagtatctgc tccctgcttg tgtgttggag gtcgctgagt agtgcgcgag caaaatttaa
     7441 gctacaacaa ggcaaggctt gaccgacaat tgcatgaaga atctgcttag ggttaggcgt
     7501 tttgcgctgc ttcgcgatgt acgggccaga tatacgcgtt
//
