LOCUS       sequence-360351-        7829 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Expresses FLAG tagged FZD5 swapped with FZD4 CRDlinker domain.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7829)
  TITLE     HOLD - Choi lab 2022
REFERENCE   2  (bases 1 to 7829)
  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 FZD5_4CRDlinker
FEATURES             Location/Qualifiers
     source          1..7829
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(120..140)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        125..454
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      305..440
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     367..386
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             521..1315
                     /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(575..594)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     1185..1204
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    1491..1612
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1528..1547)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1582..1601
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(1661..1677)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1661..1677)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1674..1696)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1685..1701
                     /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(1709..1715,1716..1733,1734..1739))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1754..1775
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1892..1909)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2063..2651)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2143..2162)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2822..3613,3614..3682))
                     /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     3445..3464
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3683..3787)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(3862..3881)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        4053..4432
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        4433..4636
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     4587..4607
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     4681..4700
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        4681..4699
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             4733..4777
                     /codon_start=1
                     /product="leader sequence from mouse preprotrypsin"
                     /label=preprotrypsin leader
                     /translation="MSALLILALVGAAVA"
     CDS             4778..4801
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             join(6419..6421,6422..6424,6425..7135)
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIKLKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSHQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(6464..6485)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(6725..6744)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     7072..7093
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             7151..7180
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             7196..7213
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(7236..7253)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    7242..7466
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      join(7512..7829,1..111)
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(7599..7618)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     join(7809..7829,1)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
ORIGIN
        1 gaacaacact caaccctatc tcggtctatt cttttgattt ataagggatt ttgccgattt
       61 cggcctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat taattctgtg
      121 gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca
      181 aagcatgcat ctcaattagt cagcaaccag gtgtggaaag tccccaggct ccccagcagg
      241 cagaagtatg caaagcatgc atctcaatta gtcagcaacc atagtcccgc ccctaactcc
      301 gcccatcccg cccctaactc cgcccagttc cgcccattct ccgccccatg gctgactaat
      361 tttttttatt tatgcagagg ccgaggccgc ctctgcctct gagctattcc agaagtagtg
      421 aggaggcttt tttggaggcc taggcttttg caaaaagctc ccgggagctt gtatatccat
      481 tttcggatct gatcaagaga caggatgagg atcgtttcgc atgattgaac aagatggatt
      541 gcacgcaggt tctccggccg cttgggtgga gaggctattc ggctatgact gggcacaaca
      601 gacaatcggc tgctctgatg ccgccgtgtt ccggctgtca gcgcaggggc gcccggttct
      661 ttttgtcaag accgacctgt ccggtgccct gaatgaactg caggacgagg cagcgcggct
      721 atcgtggctg gccacgacgg gcgttccttg cgcagctgtg ctcgacgttg tcactgaagc
      781 gggaagggac tggctgctat tgggcgaagt gccggggcag gatctcctgt catctcacct
      841 tgctcctgcc gagaaagtat ccatcatggc tgatgcaatg cggcggctgc atacgcttga
      901 tccggctacc tgcccattcg accaccaagc gaaacatcgc atcgagcgag cacgtactcg
      961 gatggaagcc ggtcttgtcg atcaggatga tctggacgaa gagcatcagg ggctcgcgcc
     1021 agccgaactg ttcgccaggc tcaaggcgcg catgcccgac ggcgaggatc tcgtcgtgac
     1081 ccatggcgat gcctgcttgc cgaatatcat ggtggaaaat ggccgctttt ctggattcat
     1141 cgactgtggc cggctgggtg tggcggaccg ctatcaggac atagcgttgg ctacccgtga
     1201 tattgctgaa gagcttggcg gcgaatgggc tgaccgcttc ctcgtgcttt acggtatcgc
     1261 cgctcccgat tcgcagcgca tcgccttcta tcgccttctt gacgagttct tctgagcggg
     1321 actctggggt tcgcgaaatg accgaccaag cgacgcccaa cctgccatca cgagatttcg
     1381 attccaccgc cgccttctat gaaaggttgg gcttcggaat cgttttccgg gacgccggct
     1441 ggatgatcct ccagcgcggg gatctcatgc tggagttctt cgcccacccc aacttgttta
     1501 ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat
     1561 ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct tatcatgtct
     1621 gtataccgtc gacctctagc tagagcttgg cgtaatcatg gtcatagctg tttcctgtgt
     1681 gaaattgtta tccgctcaca attccacaca acatacgagc cggaagcata aagtgtaaag
     1741 cctggggtgc ctaatgagtg agctaactca cattaattgc gttgcgctca ctgcccgctt
     1801 tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag
     1861 gcggtttgcg tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg
     1921 ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat
     1981 caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta
     2041 aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa
     2101 atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc
     2161 cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt
     2221 ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca
     2281 gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg
     2341 accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat
     2401 cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta
     2461 cagagttctt gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct
     2521 gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac
     2581 aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa
     2641 aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa
     2701 actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt
     2761 taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca
     2821 gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca
     2881 tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc
     2941 ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa
     3001 accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc
     3061 agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca
     3121 acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat
     3181 tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag
     3241 cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac
     3301 tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt
     3361 ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt
     3421 gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc
     3481 tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat
     3541 ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca
     3601 gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga
     3661 cacggaaatg ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg
     3721 gttattgtct catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg
     3781 ttccgcgcac atttccccga aaagtgccac ctgacgtcga cggatcggga gatctcccga
     3841 tcccctatgg tgcactctca gtacaatctg ctctgatgcc gcatagttaa gccagtatct
     3901 gctccctgct tgtgtgttgg aggtcgctga gtagtgcgcg agcaaaattt aagctacaac
     3961 aaggcaaggc ttgaccgaca attgcatgaa gaatctgctt agggttaggc gttttgcgct
     4021 gcttcgcgat gtacgggcca gatatacgcg ttgacattga ttattgacta gttattaata
     4081 gtaatcaatt acggggtcat tagttcatag cccatatatg gagttccgcg ttacataact
     4141 tacggtaaat ggcccgcctg gctgaccgcc caacgacccc cgcccattga cgtcaataat
     4201 gacgtatgtt cccatagtaa cgccaatagg gactttccat tgacgtcaat gggtggagta
     4261 tttacggtaa actgcccact tggcagtaca tcaagtgtat catatgccaa gtacgccccc
     4321 tattgacgtc aatgacggta aatggcccgc ctggcattat gcccagtaca tgaccttatg
     4381 ggactttcct acttggcagt acatctacgt attagtcatc gctattacca tggtgatgcg
     4441 gttttggcag tacatcaatg ggcgtggata gcggtttgac tcacggggat ttccaagtct
     4501 ccaccccatt gacgtcaatg ggagtttgtt ttggcaccaa aatcaacggg actttccaaa
     4561 atgtcgtaac aactccgccc cattgacgca aatgggcggt aggcgtgtac ggtgggaggt
     4621 ctatataagc agagctctct ggctaactag agaacccact gcttactggc ttatcgaaat
     4681 taatacgact cactataggg agacccaagc tggctagtta agcttggtac caatgtctgc
     4741 acttctgatc ctagctcttg ttggagctgc agttgctgac tacaaagacg atgacgacaa
     4801 gcttgcggcc gcagaggagc ggcgctgcga ccccatccgc atcgccatgt gccagaacct
     4861 cggctacaac gtgaccaaga tgcccaactt agtgggacac gagctgcaga cagacgccga
     4921 gctgcagctg acaactttca cgccgctcat ccagtacggc tgctccagcc agctgcagtt
     4981 cttcctttgt tcggtttatg tgccaatgtg cacagagaag atcaacatcc ccatcggccc
     5041 gtgcggtggc atgtgccttt cagtcaagag acgctgtgaa ccagtcctga gagaatttgg
     5101 gtttgcctgg cccgacaccc tgaactgcag caagttcccg ccccagaacg accacaacca
     5161 catgtgcatg gaaggaccag gtgatgaaga ggttcccttg ccccacaaga ctcccatcca
     5221 gcccggggaa gagtgccact ccgtgggaag caattctgat cagtacatct gggtgaagag
     5281 gagcctgaac tgtgttctca agtgctacca gccgtccttc agtgccgacg agcgcacgtt
     5341 cgccaccttc tggataggcc tgtggtcggt gctgtgcttc atctccacgt ccaccacagt
     5401 ggccaccttc ctcatcgaca tggaacgctt ccgctatcct gagcgcccca tcatcttcct
     5461 gtcagcctgc tacctgtgcg tgtcgctggg cttcctggtg cgtctggtcg tgggccatgc
     5521 cagcgtggcc tgcagccgcg agcacaacca catccactac gagaccacgg gccctgcact
     5581 gtgcaccatc gtcttcctcc tggtctactt cttcggcatg gccagctcca tctggtgggt
     5641 catcctgtcg ctcacctggt tcctggccgc cggcatgaag tggggcaacg aggccatcgc
     5701 gggctacgcg cagtacttcc acctggctgc gtggctcatc cccagcgtca agtccatcac
     5761 ggcactggcg ctgagctccg tggacgggga cccagtggcc ggcatctgct acgtgggcaa
     5821 ccagaacctg aactcgctgc gcggcttcgt gctgggcccg ctggtgctct acctgctggt
     5881 gggcacgctc ttcctgctgg cgggcttcgt gtcgctcttc cgcatccgca gcgtcatcaa
     5941 gcagggcggc accaagacgg acaagctgga gaagctcatg atccgcatcg gcatcttcac
     6001 gctgctctat acggtccccg ccagcattgt ggtggcctgc tacctgtacg agcagcacta
     6061 ccgcgagagc tgggaggcgg cgctcacctg cgcctgcccg ggccacgaca ccggccagcc
     6121 gcgcgccaag cccgagtact gggtgctcat gctcaagtac ttcatgtgcc tggtggtggg
     6181 catcacgtcg ggcgtctgga tctggtcggg caagacggtg gagtcgtggc ggcgtttcac
     6241 cagccgctgc tgctgccgcc cgcggcgcgg ccacaagagc gggggcgcca tggccgcagg
     6301 ggactacccc gaggcgagcg ccgcgctcac aggcaggacc gggccgccgg gccccgccgc
     6361 cacctaccac aagcaggtgt ccctgtcgca cgtgtaagaa ttcggtggtg gtggttcaat
     6421 ggtgagcaag ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg agctggacgg
     6481 cgacgtaaac ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg ccacctacgg
     6541 caagctgacc ctgaagttca tctgcaccac cggcaagctg cccgtgccct ggcccaccct
     6601 cgtgaccacc ttcggctacg gcctgcagtg cttcgcccgc taccccgacc acatgaagca
     6661 gcacgacttc ttcaagtccg ccatgcccga aggctacgtc caggagcgca ccatcttctt
     6721 caaggacgac ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg acaccctggt
     6781 gaaccgcatc aagctgaagg gcatcgactt caaggaggac ggcaacatcc tggggcacaa
     6841 gctggagtac aactacaaca gccacaacgt ctatatcatg gccgacaagc agaagaacgg
     6901 catcaaggtg aacttcaaga tccgccacaa catcgaggac ggcagcgtgc agctcgccga
     6961 ccactaccag cagaacaccc ccatcggcga cggccccgtg ctgctgcccg acaaccacta
     7021 cctgagccac cagtccgccc tgagcaaaga ccccaacgag aagcgcgatc acatggtcct
     7081 gctggagttc gtgaccgccg ccgggatcac tctcggcatg gacgagctgt acaagtctag
     7141 agggcccttc gaacaaaaac tcatctcaga agaggatctg aatatgcata ccggtcatca
     7201 tcaccatcac cattgagttt aaacccgctg atcagcctcg actgtgcctt ctagttgcca
     7261 gccatctgtt gtttgcccct cccccgtgcc ttccttgacc ctggaaggtg ccactcccac
     7321 tgtcctttcc taataaaatg aggaaattgc atcgcattgt ctgagtaggt gtcattctat
     7381 tctggggggt ggggtggggc aggacagcaa gggggaggat tgggaagaca atagcaggca
     7441 tgctggggat gcggtgggct ctatggcttc tgaggcggaa agaaccagct ggggctctag
     7501 ggggtatccc cacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg
     7561 cagcgtgacc gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc
     7621 ctttctcgcc acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg
     7681 gttccgattt agtgctttac ggcacctcga ccccaaaaaa cttgattagg gtgatggttc
     7741 acgtagtggg ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt
     7801 ctttaatagt ggactcttgt tccaaactg
//
