LOCUS       sequence-496434-        6360 bp    DNA     circular SYN 07-JAN-2026
DEFINITION  Expresses recombinant fluorescent CYRI-B (Fam49B) protein in 
            mammalian cells, specifically human Cyri-B with internal GFP after 
            proline 17, preserving the proposed N-terminal myristoylation site..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6360)
  AUTHORS   Le AH, Yelland T, Paul NR, Fort L, Nikolaou S, Ismail S, Machesky LM
  TITLE     CYRI-A limits invasive migration through macropinosome formation and
            integrin uptake regulation.
  JOURNAL   J Cell Biol. 2021 Sep 6;220(9). pii: 212437. doi: 
            10.1083/jcb.202012114. Epub 2021 Jun 24.
  PUBMED    34165494
REFERENCE   2  (bases 1 to 6360)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jan 7, 2026 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "J Cell 
            Biol."; date: "2021-09-6"; volume: "220(9). pii: 212437. doi"; 
            pages: " 10.1083/jcb.202012114. Epub 2021 Jun 24"
COMMENT     Sequence Label: p17-GFP-CYRI-B
FEATURES             Location/Qualifiers
     source          1..6360
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      1..589
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     490..509
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     743..760
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    877..898
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(913..918,919..936,937..943)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    951..967
                     /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)."
     primer_bind     956..978
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     975..991
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     975..991
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     enhancer        1020..1399
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1400..1603
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1554..1574
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     1600..1624
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        1834..1852
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     1834..1851
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     1865..1884
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     CDS             join(1988..1990,1991..1993,1994..2704)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(1988..1990,1991..1993,1994..2704)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2033..2054)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2294..2313)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2641..2662
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     regulatory      3666..3675
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     polyA_signal    3679..3813
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3716..3735)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3770..3789
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     promoter        3882..4239
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3882..4073
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3905..3925)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4090..4225
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(4259..4276)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4259..4275)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4268..4290)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      4488..4943
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4575..4594)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4785..4806
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4960..4979)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5079..5101
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5139..5157)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5225..5329
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5330..5398,5399..6190)
                     /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     complement(5548..5567)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
       61 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
      121 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
      181 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
      241 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
      301 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
      361 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgctacgc ttcccgaagg
      421 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
      481 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
      541 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
      601 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
      661 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
      721 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcccaat
      781 acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc acgacaggtt
      841 tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc tcactcatta
      901 ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa ttgtgagcgg
      961 ataacaattt cacacaggaa acagctatga catgattacg aattaattcg agctcgcccg
     1021 acattgatta ttgactagtt attaatagta atcaattacg gggtcattag ttcatagccc
     1081 atatatggag ttccgcgtta cataacttac ggtaaatggc ccgcctggct gaccgcccaa
     1141 cgacccccgc ccattgacgt caataatgac gtatgttccc atagtaacgc caatagggac
     1201 tttccattga cgtcaatggg tggagtattt acggtaaact gcccacttgg cagtacatca
     1261 agtgtatcat atgccaagta cgccccctat tgacgtcaat gacggtaaat ggcccgcctg
     1321 gcattatgcc cagtacatga ccttatggga ctttcctact tggcagtaca tctacgtatt
     1381 agtcatcgct attaccatgg tgatgcggtt ttggcagtac atcaatgggc gtggatagcg
     1441 gtttgactca cggggatttc caagtctcca ccccattgac gtcaatggga gtttgttttg
     1501 gcaccaaaat caacgggact ttccaaaatg tcgtaacaac tccgccccat tgacgcaaat
     1561 gggcggtagg cgtgtacggt gggaggtcta tataagcaga gctcgtttag tgaaccgtca
     1621 gatcgcctgg agacgccatc cacgctgttt tgacctccat agaagacacc gggaccgatc
     1681 cagcctccgc ggccgggaac ggtgcattgg aacgcggatt ccccgtgcca agagtgacgt
     1741 aagtaccgcc tatagagtct ataggcccac ccccttggct tcgttagaac gcggctacaa
     1801 ttaatacata accttatgta tcatacacat acgatttagg tgacactata gaataacatc
     1861 cactttgcct ttctctccac aggtgtccac tcccaggtcc aactgcacct cggttctatc
     1921 gattgaattc atggggaatc ttcttaaagt tttgacatgc acagaccttg agcaggggcc
     1981 actcgagatg gtgagcaagg gcgaggagct gttcaccggg gtggtgccca tcctggtcga
     2041 gctggacggc gacgtaaacg gccacaagtt cagcgtgtcc ggcgagggcg agggcgatgc
     2101 cacctacggc aagctgaccc tgaagttcat ctgcaccacc ggcaagctgc ccgtgccctg
     2161 gcccaccctc gtgaccaccc tgacctacgg cgtgcagtgc ttcagccgct accccgacca
     2221 catgaagcag cacgacttct tcaagtccgc catgcccgaa ggctacgtcc aggagcgcac
     2281 catcttcttc aaggacgacg gcaactacaa gacccgcgcc gaggtgaagt tcgagggcga
     2341 caccctggtg aaccgcatcg agctgaaggg catcgacttc aaggaggacg gcaacatcct
     2401 ggggcacaag ctggagtaca actacaacag ccacaacgtc tatatcatgg ccgacaagca
     2461 gaagaacggc atcaaggtga acttcaagat ccgccacaac atcgaggacg gcagcgtgca
     2521 gctcgccgac cactaccagc agaacacccc catcggcgac ggccccgtgc tgctgcccga
     2581 caaccactac ctgagcaccc agtccgccct gagcaaagac cccaacgaga agcgcgatca
     2641 catggtcctg ctggagttcg tgaccgccgc cgggatcact ctcggcatgg acgagctgta
     2701 caaggctagc aattttttcc ttgattttga aaatgcccag cctacagagt ctgagaagga
     2761 aatttataat caggtgaatg tagtattaaa agatgcagaa ggcatcttgg aggacttgca
     2821 gtcatacaga ggagctggcc acgaaatacg agaggcaatc cagcatccag cagatgagaa
     2881 gttgcaagag aaggcatggg gtgcagttgt tccactagta ggcaaattaa agaaatttta
     2941 cgaattttct cagaggttag aagcagcatt aagaggtctt ctgggagcct taacaagtac
     3001 cccatattct cccacccagc atctagagcg agagcaggct cttgctaaac agtttgcaga
     3061 aattcttcat ttcacactcc ggtttgatga actcaagatg acaaatcctg ccatacagaa
     3121 tgatttcagc tattatagaa gaacattgag tcgtatgagg attaacaatg taccggcaga
     3181 aggagaaaat gaagtaaata atgaattggc aaatcgaatg tctttgtttt atgctgaggc
     3241 aactccaatg ctgaaaacct tgagtgatgc cacaacaaaa tttgtatcag agaataaaaa
     3301 tttaccaata gaaaatacca cagattgttt aagcacaatg gctagtgtat gcagagtcat
     3361 gctggaaaca ccggaataca gaagcagatt tacaaatgaa gagacagtgt cattctgctt
     3421 gagggtaatg gtgggtgtca taatactcta tgaccacgta catccagtgg gagcatttgc
     3481 taaaacttcc aaaattgata tgaaaggttg tatcaaagtt cttaaggacc aacctcctaa
     3541 tagtgtggaa ggtcttctaa atgctctcag gtacacaaca aaacatttga atgatgagac
     3601 tacctccaag caaattaaat ccatgctgca ataaggatcc tctagagtcg acctgcagaa
     3661 gcttggccgc catggcccaa cttgtttatt gcagcttata atggttacaa ataaagcaat
     3721 agcatcacaa atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc
     3781 aaactcatca atgtatctta tcatgtctgg atcgggaatt aattcggcgc agcaccatgg
     3841 cctgaaataa cctctgaaag aggaacttgg ttaggtacct tctgaggcgg aaagaaccag
     3901 ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag gctccccagc aggcagaagt
     3961 atgcaaagca tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc aggctcccca
     4021 gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccatagt cccgccccta
     4081 actccgccca tcccgcccct aactccgccc agttccgccc attctccgcc ccatggctga
     4141 ctaatttttt ttatttatgc agaggccgag gccgcctcgg cctctgagct attccagaag
     4201 tagtgaggag gcttttttgg aggcctaggc ttttgcaaaa agctgttaac agcttggcac
     4261 tggccgtcgt tttacaacgt cgtgactggg aaaaccctgg cgttacccaa cttaatcgcc
     4321 ttgcagcaca tccccctttc gccagctggc gtaatagcga agaggcccgc accgatcgcc
     4381 cttcccaaca gttgcgcagc ctgaatggcg aatggcgcct gatgcggtat tttctcctta
     4441 cgcatctgtg cggtatttca caccgcatac gtcaaagcaa ccatagtacg cgccctgtag
     4501 cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag
     4561 cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt
     4621 tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg ctttacggca
     4681 cctcgacccc aaaaaacttg atttgggtga tggttcacgt agtgggccat cgccctgata
     4741 gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac tcttgttcca
     4801 aactggaaca acactcaacc ctatctcggg ctattctttt gatttataag ggattttgcc
     4861 gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg cgaattttaa
     4921 caaaatatta acgtttacaa ttttatggtg cactctcagt acaatctgct ctgatgccgc
     4981 atagttaagc cagccccgac acccgccaac acccgctgac gcgccctgac gggcttgtct
     5041 gctcccggca tccgcttaca gacaagctgt gaccgtctcc gggagctgca tgtgtcagag
     5101 gttttcaccg tcatcaccga aacgcgcgag acgaaagggc ctcgtgatac gcctattttt
     5161 ataggttaat gtcatgataa taatggtttc ttagacgtca ggtggcactt ttcggggaaa
     5221 tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat
     5281 gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta tgagtattca
     5341 acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca
     5401 cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac gagtgggtta
     5461 catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg aagaacgttt
     5521 tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc gtattgacgc
     5581 cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg ttgagtactc
     5641 accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat gcagtgctgc
     5701 cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg gaggaccgaa
     5761 ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg atcgttggga
     5821 accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc ctgtagcaat
     5881 ggcaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt cccggcaaca
     5941 attaatagac tggatggagg cggataaagt tgcaggacca cttctgcgct cggcccttcc
     6001 ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc gcggtatcat
     6061 tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca cgacggggag
     6121 tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct cactgattaa
     6181 gcattggtaa ctgtcagacc aagtttactc atatatactt tagattgatt taaaacttca
     6241 tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc
     6301 ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc
//
