LOCUS       sequence-360341-        6987 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Expresses FLAG tagged FZD4 swapped with FZD6 CRD.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6987)
  TITLE     HOLD - Choi lab 2022
REFERENCE   2  (bases 1 to 6987)
  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"
COMMENT     Sequence Label: pcDNA FZD4_6CRD
FEATURES             Location/Qualifiers
     source          1..6987
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(108..128)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        113..442
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      293..428
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     355..374
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             509..1303
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(563..582)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     1173..1192
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    1479..1600
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1516..1535)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1570..1589
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(1649..1665)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1649..1665)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1662..1684)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1673..1689
                     /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(1697..1703,1704..1721,1722..1727))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1742..1763
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1880..1897)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2051..2639)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2131..2150)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2810..3601,3602..3670))
                     /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     3433..3452
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3671..3775)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(3850..3869)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        4041..4420
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        4421..4624
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     4575..4595
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     4669..4688
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        4669..4687
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             4722..4766
                     /codon_start=1
                     /product="leader sequence from mouse preprotrypsin"
                     /label=preprotrypsin leader
                     /translation="MSALLILALVGAAVA"
     CDS             4767..4790
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             6297..6326
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             6342..6359
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(6382..6399)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6388..6612
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      join(6658..6987,1..99)
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6745..6764)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6955..6976
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
ORIGIN
        1 accctatctc ggtctattct tttgatttat aagggatttt gccgatttcg gcctattggt
       61 taaaaaatga gctgatttaa caaaaattta acgcgaatta attctgtgga atgtgtgtca
      121 gttagggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct
      181 caattagtca gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca
      241 aagcatgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc ccatcccgcc
      301 cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt tttttattta
      361 tgcagaggcc gaggccgcct ctgcctctga gctattccag aagtagtgag gaggcttttt
      421 tggaggccta ggcttttgca aaaagctccc gggagcttgt atatccattt tcggatctga
      481 tcaagagaca ggatgaggat cgtttcgcat gattgaacaa gatggattgc acgcaggttc
      541 tccggccgct tgggtggaga ggctattcgg ctatgactgg gcacaacaga caatcggctg
      601 ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt ttgtcaagac
      661 cgacctgtcc ggtgccctga atgaactgca ggacgaggca gcgcggctat cgtggctggc
      721 cacgacgggc gttccttgcg cagctgtgct cgacgttgtc actgaagcgg gaagggactg
      781 gctgctattg ggcgaagtgc cggggcagga tctcctgtca tctcaccttg ctcctgccga
      841 gaaagtatcc atcatggctg atgcaatgcg gcggctgcat acgcttgatc cggctacctg
      901 cccattcgac caccaagcga aacatcgcat cgagcgagca cgtactcgga tggaagccgg
      961 tcttgtcgat caggatgatc tggacgaaga gcatcagggg ctcgcgccag ccgaactgtt
     1021 cgccaggctc aaggcgcgca tgcccgacgg cgaggatctc gtcgtgaccc atggcgatgc
     1081 ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct ggattcatcg actgtggccg
     1141 gctgggtgtg gcggaccgct atcaggacat agcgttggct acccgtgata ttgctgaaga
     1201 gcttggcggc gaatgggctg accgcttcct cgtgctttac ggtatcgccg ctcccgattc
     1261 gcagcgcatc gccttctatc gccttcttga cgagttcttc tgagcgggac tctggggttc
     1321 gcgaaatgac cgaccaagcg acgcccaacc tgccatcacg agatttcgat tccaccgccg
     1381 ccttctatga aaggttgggc ttcggaatcg ttttccggga cgccggctgg atgatcctcc
     1441 agcgcgggga tctcatgctg gagttcttcg cccaccccaa cttgtttatt gcagcttata
     1501 atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc
     1561 attctagttg tggtttgtcc aaactcatca atgtatctta tcatgtctgt ataccgtcga
     1621 cctctagcta gagcttggcg taatcatggt catagctgtt tcctgtgtga aattgttatc
     1681 cgctcacaat tccacacaac atacgagccg gaagcataaa gtgtaaagcc tggggtgcct
     1741 aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa
     1801 acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta
     1861 ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc
     1921 gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg
     1981 caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt
     2041 tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa
     2101 gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct
     2161 ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc
     2221 cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg
     2281 tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct
     2341 tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag
     2401 cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga
     2461 agtggtggcc taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga
     2521 agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg
     2581 gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag
     2641 aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag
     2701 ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat
     2761 gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct
     2821 taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac
     2881 tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa
     2941 tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg
     3001 gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt
     3061 gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca
     3121 ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt
     3181 cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct
     3241 tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg
     3301 cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg
     3361 agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg
     3421 cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa
     3481 aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt
     3541 aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt
     3601 gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt
     3661 gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca
     3721 tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat
     3781 ttccccgaaa agtgccacct gacgtcgacg gatcgggaga tctcccgatc ccctatggtg
     3841 cactctcagt acaatctgct ctgatgccgc atagttaagc cagtatctgc tccctgcttg
     3901 tgtgttggag gtcgctgagt agtgcgcgag caaaatttaa gctacaacaa ggcaaggctt
     3961 gaccgacaat tgcatgaaga atctgcttag ggttaggcgt tttgcgctgc ttcgcgatgt
     4021 acgggccaga tatacgcgtt gacattgatt attgactagt tattaatagt aatcaattac
     4081 ggggtcatta gttcatagcc catatatgga gttccgcgtt acataactta cggtaaatgg
     4141 cccgcctggc tgaccgccca acgacccccg cccattgacg tcaataatga cgtatgttcc
     4201 catagtaacg ccaataggga ctttccattg acgtcaatgg gtggagtatt tacggtaaac
     4261 tgcccacttg gcagtacatc aagtgtatca tatgccaagt acgcccccta ttgacgtcaa
     4321 tgacggtaaa tggcccgcct ggcattatgc ccagtacatg accttatggg actttcctac
     4381 ttggcagtac atctacgtat tagtcatcgc tattaccatg gtgatgcggt tttggcagta
     4441 catcaatggg cgtggatagc ggtttgactc acggggattt ccaagtctcc accccattga
     4501 cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa
     4561 ctccgcccca ttgacgcaaa tgggcggtag gcgtgtacgg tgggaggtct atataagcag
     4621 agctctctgg ctaactagag aacccactgc ttactggctt atcgaaatta atacgactca
     4681 ctatagggag acccaagctg gctagttaag cttggtaccg aatgtctgca cttctgatcc
     4741 tagctcttgt tggagctgca gttgctgact acaaagacga tgacgacaag cttgcggccg
     4801 cacacagtct cttcacctgt gaaccaatta ctgttcccag atgtatgaaa atggcctaca
     4861 acatgacgtt tttccctaat ctgatgggtc attatgacca gagtattgcc gcggtggaaa
     4921 tggagcattt tcttcctctc gcaaatctgg aatgttcacc aaacattgaa actttcctct
     4981 gcaaagcatt tgtaccaacc tgcatagaac aaattcatgt ggttccacct tgtcgtaaac
     5041 tttgtgagaa agtatattct gattgcaaaa aattaattga cacttttggg atccgatggc
     5101 ctgaggagct tgaatgtgac agattacaat actgtgatga gggaccaggt gatgaagagg
     5161 ttcccttgcc ccacaagact cccatccagc ccggggaaga gtgccactcc gtgggaagca
     5221 attctgatca gtacatctgg gtgaagagga gcctgaactg tgttctcaag tgtggctacg
     5281 atgctggctt gtacagccgc tcagctaagg agttcacgga tatttggatg gctgtgtggg
     5341 ccagcctctg cttcatctcc accaccttca ccgtgctgac cttcctgatt gattcatcca
     5401 ggttttctta ccctgagcgc cccatcatat ttctcagtat gtgctataat atttatagca
     5461 ttgcttatat tgttcggctg actgtaggcc gggaaaggat atcctgtgat tttgaagagg
     5521 cggcagagcc cgttctcatc caagaaggac ttaagaacac aggatgtgca ataattttct
     5581 tgctgatgta cttttttgga atggccagct ccatttggtg ggttattctg acactcactt
     5641 ggtttttggc agccggactc aagtggggtc atgaagccat tgaaatgcac agttcttatt
     5701 tccacatcgc agcctgggct attcccgcag tgaaaaccat tgtcatcttg attatgagac
     5761 tagtggatgc cgatgaactg actggcttgt gctatgttgg gaaccaaaac ctagatgccc
     5821 tcactggctt tgtggtggct cctctcttta cgtatttggt gattggaacg ctgttcattg
     5881 cggcgggttt ggtggcctta ttcaaaattc ggtccaatct tcaaaaagac gggacaaaga
     5941 cagacaagtt ggaaaggcta atggtcaaga tcggggtctt ctcagtactg tacacggttc
     6001 ctgcaacctg tgtgattgcc tgttatttct atgaaatctc aaactgggca ctctttcgat
     6061 attctgcaga tgactcaaac atggcagttg aaatgttgaa aatttttatg tctttgctcg
     6121 tgggcatcac ttcaggcatg tggatttggt ctgccaaaac tcttcacacg tggcaaaagt
     6181 gttctaaccg attggtgaat tctgggaagg taaagagaga gaagaggggg aatggttggg
     6241 tgaagccagg aaaaggcaac gagactgtgg tatagctcga gtctagaggg cccttcgaac
     6301 aaaaactcat ctcagaagag gatctgaata tgcataccgg tcatcatcac catcaccatt
     6361 gagtttaaac ccgctgatca gcctcgactg tgccttctag ttgccagcca tctgttgttt
     6421 gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc ctttcctaat
     6481 aaaatgagga aattgcatcg cattgtctga gtaggtgtca ttctattctg gggggtgggg
     6541 tggggcagga cagcaagggg gaggattggg aagacaatag caggcatgct ggggatgcgg
     6601 tgggctctat ggcttctgag gcggaaagaa ccagctgggg ctctaggggg tatccccacg
     6661 cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta
     6721 cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt
     6781 tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg
     6841 ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt agtgggccat
     6901 cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac
     6961 tcttgttcca aactggaaca acactca
//
