LOCUS       sequence-357749-        6755 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Expresses murine Fc fusion to the first three EGF-like domains of 
            human NOTCH3 WT.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6755)
  TITLE     NOTCH3 N-terminal domain fusions with Fc -- WT and CADASIL mutants
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6755)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: Fc-E3
FEATURES             Location/Qualifiers
     source          1..6755
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        119..498
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        499..702
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     653..673
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     747..766
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        747..765
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      783..792
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     sig_peptide     789..851
                     /codon_start=1
                     /product="leader sequence from mouse immunoglobulin kappa 
                     light chain"
                     /label=Ig-kappa leader
                     /translation="METDTLLLWVLLLWVPGSTGD"
     CDS             1976..2005
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             2021..2038
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2061..2078)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2067..2291
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2337..2765
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2424..2443)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2634..2655
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2774..2794)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2779..3108
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2959..3094
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3021..3040
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3157..4182
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     CDS             3157..4182
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     polyA_signal    4312..4433
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4349..4368)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4403..4422
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4482..4498)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4482..4498)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4495..4517)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4506..4522
                     /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(4530..4536,4537..4554,4555..4560))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     primer_bind     complement(4713..4730)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4884..5472)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4964..4983)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5643..6434,6435..6503))
                     /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     6266..6285
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6504..6608)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6683..6702)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 tgcgcgagca aaatttaagc tacaacaagg caaggcttga ccgacaattg catgaagaat
       61 ctgcttaggg ttaggcgttt tgcgctgctt cgcgatgtac gggccagata tacgcgttga
      121 cattgattat tgactagtta ttaatagtaa tcaattacgg ggtcattagt tcatagccca
      181 tatatggagt tccgcgttac ataacttacg gtaaatggcc cgcctggctg accgcccaac
      241 gacccccgcc cattgacgtc aataatgacg tatgttccca tagtaacgcc aatagggact
      301 ttccattgac gtcaatgggt ggagtattta cggtaaactg cccacttggc agtacatcaa
      361 gtgtatcata tgccaagtac gccccctatt gacgtcaatg acggtaaatg gcccgcctgg
      421 cattatgccc agtacatgac cttatgggac tttcctactt ggcagtacat ctacgtatta
      481 gtcatcgcta ttaccatggt gatgcggttt tggcagtaca tcaatgggcg tggatagcgg
      541 tttgactcac ggggatttcc aagtctccac cccattgacg tcaatgggag tttgttttgg
      601 caccaaaatc aacgggactt tccaaaatgt cgtaacaact ccgccccatt gacgcaaatg
      661 ggcggtaggc gtgtacggtg ggaggtctat ataagcagag ctctctggct aactagagaa
      721 cccactgctt actggcttat cgaaattaat acgactcact atagggagac ccaagctggc
      781 tagccaccat ggagacagac acactcctgc tatgggtact gctgctctgg gttccaggtt
      841 ccactggtga cgcggcccag ccggccaggc gcgcccccag agtgcccata acacagaacc
      901 cctgtcctcc actcaaagag tgtcccccat gcgcagctcc agacctcttg ggtggaccat
      961 ccgtcttcat cttccctcca aagatcaagg atgtactcat gatctccctg agccccatgg
     1021 tcacatgtgt ggtggtggat gtgagcgagg atgacccaga cgtccagatc agctggtttg
     1081 tgaacaacgt ggaagtacac acagctcaga cacaaaccca tagagaggat tacaacagta
     1141 ctctccgggt ggtcagtgcc ctccccatcc agcaccagga ctggatgagt ggcaaggagt
     1201 tcaaatgcaa ggtcaacaac agagccctcc catcccccat cgagaaaacc atctcaaaac
     1261 ccagagggcc agtaagagct ccacaggtat atgtcttgcc tccaccagca gaagagatga
     1321 ctaagaaaga gttcagtctg acctgcatga tcacaggctt cttacctgcc gaaattgctg
     1381 tggactggac cagcaatggg cgtacagagc aaaactacaa gaacaccgca acagtcctgg
     1441 actctgatgg ttcttacttc atgtacagca agctcagagt acaaaagagc acttgggaaa
     1501 gaggaagtct tttcgcctgc tcagtggtcc acgagggtct gcacaatcac cttacgacta
     1561 agaccatctc ccggtctctg ggtaaaaagc ttgccccccc ttgcctggac ggaagcccgt
     1621 gtgcaaatgg aggtcgttgc acccagctgc cctcccggga ggctgcctgc ctgtgcccgc
     1681 ctggctgggt gggtgagcgg tgtcagctgg aggacccctg tcactcaggc ccctgtgctg
     1741 gccgtggtgt ctgccagagt tcagtggtgg ctggcaccgc ccgattctca tgccggtgcc
     1801 cccgtggctt ccgaggccct gactgctccc tgccagatcc ctgcctcagc agcccttgtg
     1861 cccacggtgc ccgctgctca gtggggcccg atggacgctt cctctgctcc tgcccacctg
     1921 gctaccaggg ccgcagctgc cgaagcgacg tggatgagtg actcgaggag ggcccgaaca
     1981 aaaactcatc tcagaagagg atctgaatag cgccgtcgac catcatcatc atcatcattg
     2041 agtttaaacc cgctgatcag cctcgactgt gccttctagt tgccagccat ctgttgtttg
     2101 cccctccccc gtgccttcct tgaccctgga aggtgccact cccactgtcc tttcctaata
     2161 aaatgaggaa attgcatcgc attgtctgag taggtgtcat tctattctgg ggggtggggt
     2221 ggggcaggac agcaaggggg aggattggga agacaatagc aggcatgctg gggatgcggt
     2281 gggctctatg gcttctgagg cggaaagaac cagctggggc tctagggggt atccccacgc
     2341 gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac
     2401 acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt
     2461 cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc
     2521 tttacggcac ctcgacccca aaaaacttga ttagggtgat ggttcacgta gtgggccatc
     2581 gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact
     2641 cttgttccaa actggaacaa cactcaaccc tatctcggtc tattcttttg atttataagg
     2701 gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc
     2761 gaattaattc tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg ctccccagca
     2821 ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca
     2881 ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc aaccatagtc
     2941 ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca ttctccgccc
     3001 catggctgac taattttttt tatttatgca gaggccgagg ccgcctctgc ctctgagcta
     3061 ttccagaagt agtgaggagg cttttttgga ggcctaggct tttgcaaaaa gctcccggga
     3121 gcttgtatat ccattttcgg atctgatcag cacgtgatga aaaagcctga actcaccgcg
     3181 acgtctgtcg agaagtttct gatcgaaaag ttcgacagcg tctccgacct gatgcagctc
     3241 tcggagggcg aagaatctcg tgctttcagc ttcgatgtag gagggcgtgg atatgtcctg
     3301 cgggtaaata gctgcgccga tggtttctac aaagatcgtt atgtttatcg gcactttgca
     3361 tcggccgcgc tcccgattcc ggaagtgctt gacattgggg aattcagcga gagcctgacc
     3421 tattgcatct cccgccgtgc acagggtgtc acgttgcaag acctgcctga aaccgaactg
     3481 cccgctgttc tgcagccggt cgcggaggcc atggatgcga tcgctgcggc cgatcttagc
     3541 cagacgagcg ggttcggccc attcggaccg caaggaatcg gtcaatacac tacatggcgt
     3601 gatttcatat gcgcgattgc tgatccccat gtgtatcact ggcaaactgt gatggacgac
     3661 accgtcagtg cgtccgtcgc gcaggctctc gatgagctga tgctttgggc cgaggactgc
     3721 cccgaagtcc ggcacctcgt gcacgcggat ttcggctcca acaatgtcct gacggacaat
     3781 ggccgcataa cagcggtcat tgactggagc gaggcgatgt tcggggattc ccaatacgag
     3841 gtcgccaaca tcttcttctg gaggccgtgg ttggcttgta tggagcagca gacgcgctac
     3901 ttcgagcgga ggcatccgga gcttgcagga tcgccgcggc tccgggcgta tatgctccgc
     3961 attggtcttg accaactcta tcagagcttg gttgacggca atttcgatga tgcagcttgg
     4021 gcgcagggtc gatgcgacgc aatcgtccga tccggagccg ggactgtcgg gcgtacacaa
     4081 atcgcccgca gaagcgcggc cgtctggacc gatggctgtg tagaagtact cgccgatagt
     4141 ggaaaccgac gccccagcac tcgtccgagg gcaaaggaat agcacgtgct acgagatttc
     4201 gattccaccg ccgccttcta tgaaaggttg ggcttcggaa tcgttttccg ggacgccggc
     4261 tggatgatcc tccagcgcgg ggatctcatg ctggagttct tcgcccaccc caacttgttt
     4321 attgcagctt ataatggtta caaataaagc aatagcatca caaatttcac aaataaagca
     4381 tttttttcac tgcattctag ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc
     4441 tgtataccgt cgacctctag ctagagcttg gcgtaatcat ggtcatagct gtttcctgtg
     4501 tgaaattgtt atccgctcac aattccacac aacatacgag ccggaagcat aaagtgtaaa
     4561 gcctggggtg cctaatgagt gagctaactt acattaattg cgttgcgctc actgcccgct
     4621 ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga
     4681 ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc
     4741 gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa
     4801 tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt
     4861 aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa
     4921 aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt
     4981 ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg
     5041 tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc
     5101 agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc
     5161 gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta
     5221 tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct
     5281 acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc
     5341 tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa
     5401 caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa
     5461 aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa
     5521 aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt
     5581 ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac
     5641 agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc
     5701 atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt accatctggc
     5761 cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata
     5821 aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc
     5881 cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc
     5941 aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca
     6001 ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa
     6061 gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca
     6121 ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt
     6181 tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt
     6241 tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg
     6301 ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga
     6361 tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc
     6421 agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg
     6481 acacggaaat gttgaatact catactcttc ctttttcaat attattgaag catttatcag
     6541 ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg
     6601 gttccgcgca catttccccg aaaagtgcca cctgacgtcg acggatcggg agatctcccg
     6661 atcccctatg gtgcactctc agtacaatct gctctgatgc cgcatagtta agccagtatc
     6721 tgctccctgc ttgtgtgttg gaggtcgctg agtag
//
