LOCUS       sequence-436903-        7442 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses Human Sema4D with N terminal flag after signal peptide 
            containing 497P mutation.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7442)
  AUTHORS   Bessa P, Newman AG, Yan K, Schaub T, Dannenberg R, Lajko D, 
            Eilenberger J, Brunet T, Textoris-Taube K, Kemmler E, Deng P, 
            Banerjee P, Ravindran E, Preissner R, Rosario M, Tarabykin V
  TITLE     Semaphorin heterodimerization in cis regulates membrane targeting 
            and neocortical wiring.
  JOURNAL   Nat Commun. 2024 Aug 16;15(1):7059. doi: 10.1038/s41467-024-51009-1.
  PUBMED    39152101
REFERENCE   2  (bases 1 to 7442)
  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 
            Commun."; date: "2024-08-16"; volume: "15(1):7059. doi"; pages: " 
            10.1038/s41467-024-51009-1"
COMMENT     Sequence Label: pCAG-sp-flag-hSema4D-Q497P
FEATURES             Location/Qualifiers
     source          1..7442
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        4..383
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     intron          686..1702
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1626..1648
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1710..1729
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     CDS             1821..1844
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     primer_bind     complement(4463..4482)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    4528..4583
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(4582..4601)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(4944..4960)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4944..4960)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4957..4979)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4968..4984
                     /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(4992..4998,4999..5016,5017..5022))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5037..5058
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        5117..5312
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      5163..5298
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    5318..5452
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5355..5374)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5409..5428
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5520..5537)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5691..6279)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5771..5790)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6450..7241,7242..7310))
                     /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     7073..7092
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7311..7415)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gtcgacattg attattgact agttattaat agtaatcaat tacggggtca ttagttcata
       61 gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc
      121 ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta acgccaatag
      181 ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac ttggcagtac
      241 atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg
      301 cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg
      361 tattagtcat cgctattacc atggtcgagg tgagccccac gttctgcttc actctcccca
      421 tctccccccc ctccccaccc ccaattttgt atttatttat tttttaatta ttttgtgcag
      481 cgatgggggc gggggggggg gggggggggg ggccaggggg gggggggggg gggggggggg
      541 gggggggggg gggggggggg ggcggggcgg ggcgaggcgg agaggtgcgg cggcagccaa
      601 tcagagcggc gcgctccgaa agtttccttt tatggcgagg cggcggcggc ggcggcccta
      661 taaaaagcga agcgcgcggc gggcgggagt cgctgcgcgc tgccttcgcc ccgtgccccg
      721 ctccgccgcc gcctcgcgcc gcccgccccg gctctgactg accgcgttac tcccacaggt
      781 gagcgggcgg gacggccctt ctcctccggg ctgtaattag cgcttggttt aatgacggct
      841 tgtttctttt ctgtggctgc gtgaaagcct tgaggggctc cgggagggcc ctttgtgcgg
      901 ggggagcggc tcggggggtg cgtgcgtgtg tgtgtgcgtg gggagcgccg cgtgcggctc
      961 cgcgctgccc ggcggctgtg agcgctgcgg gcgcggcgcg gggctttgtg cgctccgcag
     1021 tgtgcgcgag gggagcgcgg ccgggggcgg tgccccgcgg tgcggggggg gctgcgaggg
     1081 gaacaaaggc tgcgtgcggg gtgtgtgcgt gggggggtga gcagggggtg tgggcgcgtc
     1141 ggtcgggctg caaccccccc tgcacccccc tccccgagtt gctgagcacg gcccggcttc
     1201 gggtgcgggg ctccgtacgg ggcgtggcgc ggggctcgcc gtgccgggcg gggggtggcg
     1261 gcaggtgggg gtgccgggcg gggcggggcc gcctcgggcc ggggagggct cgggggaggg
     1321 gcgcggcggc ccccggagcg ccggcggctg tcgaggcgcg gcgagccgca gccattgcct
     1381 tttatggtaa tcgtgcgaga gggcgcaggg acttcctttg tcccaaatct gtgcggagcc
     1441 gaaatctggg aggcgccgcc gcaccccctc tagcgggcgc ggggcgaagc ggtgcggcgc
     1501 cggcaggaag gaaatgggcg gggagggcct tcgtgcgtcg ccgcgccgcc gtccccttct
     1561 ccctctccag cctcggggct gtccgcgggg ggacggctgc cttcgggggg gacggggcag
     1621 ggcggggttc ggcttctggc gtgtgaccgg cggctctaga gcctctgcta accatgttca
     1681 tgccttcttc tttttcctac agctcctggg caacgtgctg gttattgtgc tgtctcatca
     1741 ttttggcaaa gatgaggatg tgcaccccca ttagggggct gctcatggcc cttgcagtga
     1801 tgtttgggac agcgatggca gattataaag atgatgatga taaatttgca cccatacccc
     1861 ggatcacctg ggagcacaga gaggtgcacc tggtgcagtt tcatgagcca gacatctaca
     1921 actactcagc cttgctgctg agcgaggaca aggacacctt gtacataggt gcccgggagg
     1981 cggtcttcgc tgtgaacgca ctcaacatct ccgagaagca gcatgaggtg tattggaagg
     2041 tctcagaaga caaaaaagca aaatgtgcag aaaaggggaa atcaaaacag acagagtgcc
     2101 tcaactacat ccgggtgctg cagccactca gcgccacttc cctttacgtg tgtgggacca
     2161 acgcattcca gccggcctgt gaccacctga acttaacatc ctttaagttt ctggggaaaa
     2221 atgaagatgg caaaggaaga tgtccctttg acccagcaca cagctacaca tccgtcatgg
     2281 ttgatggaga actttattcg gggacgtcgt ataatttttt gggaagtgaa cccatcatct
     2341 cccgaaattc ttcccacagt cctctgagga cagaatatgc aatcccttgg ctgaacgagc
     2401 ctagtttcgt gtttgctgac gtgatccgaa aaagcccaga cagccccgac ggcgaggatg
     2461 acagggtcta cttcttcttc acggaggtgt ctgtggagta tgagtttgtg ttcagggtgc
     2521 tgatcccacg gatagcaaga gtgtgcaagg gggaccaggg cggcctgagg accttgcaga
     2581 agaaatggac ctccttcctg aaagcccgac tcatctgctc ccggccagac agcggcttgg
     2641 tcttcaatgt gctgcgggat gtcttcgtgc tcaggtcccc gggcctgaag gtgcctgtgt
     2701 tctatgcact cttcacccca cagctgaaca acgtggggct gtcggcagtg tgcgcctaca
     2761 acctgtccac agccgaggag gtcttctccc acgggaagta catgcagagc accacagtgg
     2821 agcagtccca caccaagtgg gtgcgctata atggcccggt acccaagccg cggcctggag
     2881 cgtgcatcga cagcgaggca cgggccgcca actacaccag ctccttgaat ttgccagaca
     2941 agacgctgca gttcgttaaa gaccaccctt tgatggatga ctcggtaacc ccaatagaca
     3001 acaggcccag gttaatcaag aaagatgtga actacaccca gatcgtggtg gaccggaccc
     3061 aggccctgga tgggactgtc tatgatgtca tgtttgtcag cacagaccgg ggagctctgc
     3121 acaaagccat cagcctcgag cacgctgttc acatcatcga ggagacccag ctcttccagg
     3181 actttgagcc agtccagacc ctgctgctgt cttcaaagaa gggcaacagg tttgtctatg
     3241 ctggctctaa ctcgggcgtg gtccccgccc cgctggcctt ctgtgggaag cacggcacct
     3301 gcgaggactg tgtgctggcg cgggacccct actgcgcctg gagcccgccc acagcgacct
     3361 gcgtggctct gcaccagacc gagagcccca gcaggggttt gattcaggag atgagcggcg
     3421 atgcttctgt gtgcccggat aaaagtaaag gaagttaccg gcagcatttt ttcaagcacg
     3481 gtggcacagc ggaactgaaa tgctcccaaa aatccaacct ggcccgggtc ttttggaagt
     3541 tccagaatgg cgtgttgaag gccgagagcc ccaagtacgg tcttatgggc agaaaaaact
     3601 tgctcatctt caacttgtca gaaggagaca gtggggtgta ccagtgcctg tcagaggaga
     3661 gggttaagaa caaaacggtc ttccaagtgg tcgccaagca cgtcctggaa gtgaaggtgg
     3721 ttccaaagcc cgtagtggcc cccaccttgt cagttgttca gacagaaggt agtaggattg
     3781 ccaccaaagt gttggtggca tccacccaag ggtcttctcc cccaacccca gccgtgcagg
     3841 ccacctcctc cggggccatc acccttcctc ccaagcctgc gcccaccggc acatcctgcg
     3901 aaccaaagat cgtcatcaac acggtccccc agctccactc ggagaaaacc atgtatctta
     3961 agtccagcga caaccgcctc ctcatgtccc tcttcctctt cttctttgtt ctcttcctct
     4021 gcctcttttt ctacaactgc tataagggat acctgcccag acagtgcttg aaattccgct
     4081 cggccctact aattgggaag aagaagccca agtcagattt ctgtgaccgt gagcagagcc
     4141 tgaaggagac gttagtagag ccagggagct tctcccagca gaatggggag caccccaagc
     4201 cagccctgga caccggctat gagaccgagc aagacaccat caccagcaaa gtccccacgg
     4261 atagggagga ctcacagagg atcgacgacc tttctgccag ggacaagccc tttgacgtca
     4321 agtgtgagct gaagttcgct gactcagacg cagatggaga ctgacctcga ggatatcgcg
     4381 gccgcactcc tcaggtgcag gctgcctatc agaaggtggt ggctggtgtg gccaatgccc
     4441 tggctcacaa ataccactga gatctttttc cctctgccaa aaattatggg gacatcatga
     4501 agccccttga gcatctgact tctggctaat aaaggaaatt tattttcatt gcaatagtgt
     4561 gttggaattt tttgtgtctc tcactcggaa ggacatatgg gagggcaaat catttaaaac
     4621 atcagaatga gtatttggtt tagagtttgg caacatatgc ccatatgctg gctgccatga
     4681 acaaaggttg gctataaaga ggtcatcagt atatgaaaca gccccctgct gtccattcct
     4741 tattccatag aaaagccttg acttgaggtt agattttttt tatattttgt tttgtgttat
     4801 ttttttcttt aacatcccta aaattttcct tacatgtttt actagccaga tttttcctcc
     4861 tctcctgact actcccagtc atagctgtcc ctcttctctt atggagatcc ctcgacctgc
     4921 agcccaagct tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc
     4981 acaattccac acaacatacg agccggaagc ataaagtgta aagcctgggg tgcctaatga
     5041 gtgagctaac tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg
     5101 tcgtgccagc ggatccgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc
     5161 ccatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt
     5221 tttttattta tgcagaggcc gaggccgcct cggcctctga gctattccag aagtagtgag
     5281 gaggcttttt tggaggccta ggcttttgca aaaagctaac ttgtttattg cagcttataa
     5341 tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt tttcactgca
     5401 ttctagttgt ggtttgtcca aactcatcaa tgtatcttat catgtctgga tccgctgcat
     5461 taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc
     5521 tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca
     5581 aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca
     5641 aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg
     5701 ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg
     5761 acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt
     5821 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt
     5881 tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc
     5941 tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt
     6001 gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt
     6061 agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc
     6121 tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa
     6181 agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt
     6241 tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct
     6301 acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta
     6361 tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa
     6421 agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc
     6481 tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact
     6541 acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc
     6601 tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt
     6661 ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta
     6721 agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg
     6781 tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt
     6841 acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc
     6901 agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt
     6961 actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc
     7021 tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc
     7081 gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa
     7141 ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac
     7201 tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa
     7261 aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt
     7321 tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa
     7381 tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct
     7441 gg
//
