LOCUS       sequence-469802-        7369 bp    DNA     circular SYN 23-APR-2025
DEFINITION  Mammalian expression of truncated bovine arrestin-3 (1-393).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7369)
  AUTHORS   Zheng C, Weinstein LD, Nguyen KK, Grewal A, Gurevich EV, Gurevich VV
  TITLE     GPCR Binding and JNK3 Activation by Arrestin-3 Have Different 
            Structural Requirements.
  JOURNAL   Cells. 2023 Jun 6;12(12):1563. doi: 10.3390/cells12121563.
  PUBMED    37371033
REFERENCE   2  (bases 1 to 7369)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 23, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Cells."; 
            date: "2023-06-6"; volume: "12(12):1563. doi"; pages: " 
            10.3390/cells12121563"
COMMENT     Sequence Label: pVEA3-Tr
FEATURES             Location/Qualifiers
     source          1..7369
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     23..42
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     276..293
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    410..431
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(446..451,452..469,470..476)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    484..500
                     /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     489..511
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     508..524
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     508..524
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     polyA_signal    573..694
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(584..603)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     638..657
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(868..1662)
                     /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(979..998)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     1589..1608
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(1729..2058)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      1743..1878
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(1797..1816)
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     primer_bind     2043..2063
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      complement(2072..2500)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2182..2203)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2394..2413
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     polyA_signal    2546..2770
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     2759..2776
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     promoter        2796..2814
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     2796..2813
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     CDS             complement(join(4104..4814,4815..4817,4818..4820))
                     /codon_start=1
                     /product="Venus YFP with monomerizing A206K mutation (Nagai
                     et al., 2002; Kremers et al., 2006)"
                     /label=mVenus
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KLICTTGKLPVPWPTLVTTLGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIK
                     ANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(4146..4167)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     4495..4514
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4754..4775
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(4854..4873)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4855..4873)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        complement(4918..5121)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(4947..4967)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        5122..5501
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     5673..5692
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        5767..5871
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5872..5940,5941..6732)
                     /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(6090..6109)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      join(6903..7369,1..122)
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc
       61 gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga
      121 aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca
      181 tgttctttcc tgcgttatcc cctgattctg tggataaccg tattaccgcc tttgagtgag
      241 ctgataccgc tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc gaggaagcgg
      301 aagagcgccc aatacgcaaa ccgcctctcc ccgcgcgttg gccgattcat taatgcagct
      361 ggcacgacag gtttcccgac tggaaagcgg gcagtgagcg caacgcaatt aatgtgagtt
      421 agctcactca ttaggcaccc caggctttac actttatgct tccggctcgt atgttgtgtg
      481 gaattgtgag cggataacaa tttcacacag gaaacagcta tgaccatgat tacgccaagc
      541 tctagctaga ggtcgacggt atacagacat gataagatac attgatgagt ttggacaaac
      601 cacaactaga atgcagtgaa aaaaatgctt tatttgtgaa atttgtgatg ctattgcttt
      661 atttgtaacc attataagct gcaataaaca agttggggtg ggcgaagaac tccagcatga
      721 gatccccgcg ctggaggatc atccagccgg cgtcccggaa aacgattccg aagcccaacc
      781 tttcatagaa ggcggcggtg gaatcgaaat ctcgtgatgg caggttgggc gtcgcttggt
      841 cggtcatttc gaaccccaga gtcccgctca gaagaactcg tcaagaaggc gatagaaggc
      901 gatgcgctgc gaatcgggag cggcgatacc gtaaagcacg aggaagcggt cagcccattc
      961 gccgccaagc tcttcagcaa tatcacgggt agccaacgct atgtcctgat agcggtccgc
     1021 cacacccagc cggccacagt cgatgaatcc agaaaagcgg ccattttcca ccatgatatt
     1081 cggcaagcag gcatcgccat gggtcacgac gagatcctcg ccgtcgggca tgcgcgcctt
     1141 gagcctggcg aacagttcgg ctggcgcgag cccctgatgc tcttcgtcca gatcatcctg
     1201 atcgacaaga ccggcttcca tccgagtacg tgctcgctcg atgcgatgtt tcgcttggtg
     1261 gtcgaatggg caggtagccg gatcaagcgt atgcagccgc cgcattgcat cagccatgat
     1321 ggatactttc tcggcaggag caaggtgaga tgacaggaga tcctgccccg gcacttcgcc
     1381 caatagcagc cagtcccttc ccgcttcagt gacaacgtcg agcacagctg cgcaaggaac
     1441 gcccgtcgtg gccagccacg atagccgcgc tgcctcgtcc tgcagttcat tcagggcacc
     1501 ggacaggtcg gtcttgacaa aaagaaccgg gcgcccctgc gctgacagcc ggaacacggc
     1561 ggcatcagag cagccgattg tctgttgtgc ccagtcatag ccgaatagcc tctccaccca
     1621 agcggccgga gaacctgcgt gcaatccatc ttgttcaatc atgcgaaacg atcctcatcc
     1681 tgtctcttga tcagatccga aaatggatat acaagctccc gggagctttt tgcaaaagcc
     1741 taggcctcca aaaaagcctc ctcactactt ctggaatagc tcagaggcag aggcggcctc
     1801 ggcctctgca taaataaaaa aaattagtca gccatggggc ggagaatggg cggaactggg
     1861 cggagttagg ggcgggatgg gcggagttag gggcgggact atggttgctg actaattgag
     1921 atgcatgctt tgcatacttc tgcctgctgg ggagcctggg gactttccac acctggttgc
     1981 tgactaattg agatgcatgc tttgcatact tctgcctgct ggggagcctg gggactttcc
     2041 acaccctaac tgacacacat tccacagaat taattcgcgt taaatttttg ttaaatcagc
     2101 tcatttttta accaataggc cgaaatcggc aaaatccctt ataaatcaaa agaatagacc
     2161 gagatagggt tgagtgttgt tccagtttgg aacaagagtc cactattaaa gaacgtggac
     2221 tccaacgtca aagggcgaaa aaccgtctat cagggcgatg gcccactacg tgaaccatca
     2281 ccctaatcaa gttttttggg gtcgaggtgc cgtaaagcac taaatcggaa ccctaaaggg
     2341 agcccccgat ttagagcttg acggggaaag ccggcgaacg tggcgagaaa ggaagggaag
     2401 aaagcgaaag gagcgggcgc tagggcgctg gcaagtgtag cggtcacgct gcgcgtaacc
     2461 accacacccg ccgcgcttaa tgcgccgcta cagggcgcgt ggggataccc cctagagccc
     2521 cagctggttc tttccgcctc agaagccata gagcccaccg catccccagc atgcctgcta
     2581 ttgtcttccc aatcctcccc cttgctgtcc tgccccaccc caccccccag aatagaatga
     2641 cacctactca gacaatgcga tgcaatttcc tcattttatt aggaaaggac agtgggagtg
     2701 gcaccttcca gggtcaagga aggcacgggg gaggggcaaa caacagatgg ctggcaacta
     2761 gaaggcacag tcgaggctga tcagcgagct ctagcattta ggtgacacta tagaataggg
     2821 ccaagcttcc agccctaatc gatacagaac tggtcgtcat agtcctcgtc cttcagcccc
     2881 ttgagccgct accgggcaaa gtcttcgaac acgatgtcgt cgtctgtggc atagttggtt
     2941 tcaaattcaa tgaggttggt gtccacgggg gcatctgttt caggaacagc tgactggggc
     3001 ctgggaaggg cgatgtgatc atggggcttg gggtgcatga gaacaaatgg cagctccact
     3061 gagacatccc cgcctcgtga caccaccagc ttcaccttga ccctgtagga caccagtatc
     3121 cccagcacct ccttgttggc accctccttc acaatggtgc tcgaggccag gttggtgtcc
     3181 tcgtgcttga gcttcccatc tagagcgagg ccgcgcttct cccggttgtt gctgagcagc
     3241 ggggttatgg tgtacacctt acagaacgtg gaactgggcg acacctggtc atcttgttcg
     3301 acctgagcca cggggcactt gtactgggcg gtgctgaaga ggcagatgtc ggcgtactgt
     3361 ctcacagaga ctttgatctt cttgacagtc ttggtggagt tattggtgac gtggacattg
     3421 acattgaggg gctccccgtg gtagtacagc tccttgtcca gtgaagcctc aaggtgcagg
     3481 gaccggtcag acatgaggaa gtggcgtgtg gtttcagctg aaggctgggg ccctggtttc
     3541 tctggggcaa actgcacctt tcggatcacc agacgtacgg agttccgttt gtggcttttc
     3601 tcttctagtg atttggcgca gaaggctcga atctcaaagt cgaccccgca agccttcccc
     3661 gtatcctctg ggcctggctg tagggtgacg gagcagggca ggttctgggg tattgtgaaa
     3721 aaaaaggggt gggcatgctg gcccagcttc ctcagcagcc gctcctgcag acgcgtaggt
     3781 ggtctaggtg ggttaggtgt tggagggaag gcctggtagt tggcgatgaa caggtctttg
     3841 cggaaggaca agcccaggac gtccaggtct tcgcggccat agcgaaaggc gcacgtgagg
     3901 gtcacaaaca ctttgcggtc cttcaagtag tccggatcca ctagcaccac accatctaca
     3961 gggtccactt tgtccaggtg gtccacaaag tcccgcttgc ctaggtacac ggtgagcttg
     4021 cagttaggac tcgatttctt gaagaccctg gtcccgggtt tctcccccat ggagtcgagc
     4081 gctcgccgag acttaagtcc ggacttgtac agctcgtcca tgccgagagt gatcccggcg
     4141 gcggtcacga actccagcag gaccatgtga tcgcgcttct cgttggggtc tttgctcagt
     4201 ttggactggt agctcaggta gtggttgtcg ggcagcagca cggggccgtc gccgatgggg
     4261 gtgttctgct ggtagtggtc ggcgagctgc acgccgccgt cctcgatgtt gtggcggatc
     4321 ttgaagttgg ccttgatgcc gttcttctgc ttgtcggcgg tgatatagac gttgtggctg
     4381 ttgtagttgt actccagctt gtgccccagg atgttgccgt cctccttgaa gtcgatgccc
     4441 ttcagctcga tgcggttcac cagggtgtcg ccctcgaact tcacctcggc gcgggtcttg
     4501 tagttgccgt cgtccttgaa gaagatggtg cgctcctgga cgtagccttc gggcatggcg
     4561 gacttgaaga agtcgtgctg cttcatgtgg tcggggtagc gggcgaagca ctgcaggccg
     4621 tagcccaggg tggtcacgag ggtgggccag ggcacgggca gcttgccggt ggtgcagatc
     4681 agcttcaggg tcagcttgcc gtaggtggca tcgccctcgc cctcgccgga cacgctgaac
     4741 ttgtggccgt ttacgtcgcc gtccagctcg accaggatgg gcaccacccc ggtgaacagc
     4801 tcctcgccct tgctcaccat cgtggccgcg atcgcgaatt ccagcttggg tctccctata
     4861 gtgagtcgta ttaatttcga taagccagta agcagtgggt tctctagtta gccagagagc
     4921 tctgcttata tagacctccc accgtacacg cctaccgccc atttgcgtca atggggcgga
     4981 gttgttacga cattttggaa agtcccgttg attttggtgc caaaacaaac tcccattgac
     5041 gtcaatgggg tggagacttg gaaatccccg tgagtcaaac cgctatccac gcccattgat
     5101 gtactgccaa aaccgcatca ccatggtaat agcgatgact aatacgtaga tgtactgcca
     5161 agtaggaaag tcccataagg tcatgtactg ggcataatgc caggcgggcc atttaccgtc
     5221 attgacgtca atagggggcg tacttggcat atgatacact tgatgtactg ccaagtgggc
     5281 agtttaccgt aaatactcca cccattgacg tcaatggaaa gtccctattg gcgttactat
     5341 gggaacatac gtcattattg acgtcaatgg gcgggggtcg ttgggcggtc agccaggcgg
     5401 gccatttacc gtaagttatg taacgcggaa ctccatatat gggctatgaa ctaatgaccc
     5461 cgtaattgat tactattaat aactagtcaa taatcaatgt caacgcgtat atctggcccg
     5521 tacatcgcga agcagcgcaa aacgcctaac cctaagcaga ttcttcatgc aattgtcggt
     5581 caagccttgc cttgttgtag cttaaatttt gctcgcgcac tactcagcga cctccaacac
     5641 acaagcaggg agcagatact ggcttaacta tgcggcatca gagcagattg tactgagagt
     5701 gcaccatagg ggatcgggag atctcccgat ccgtcgacgt caggtggcac ttttcgggga
     5761 aatgtgcgcg gaacccctat ttgtttattt ttctaaatac attcaaatat gtatccgctc
     5821 atgagacaat aaccctgata aatgcttcaa taatattgaa aaaggaagag tatgagtatt
     5881 caacatttcc gtgtcgccct tattcccttt tttgcggcat tttgccttcc tgtttttgct
     5941 cacccagaaa cgctggtgaa agtaaaagat gctgaagatc agttgggtgc acgagtgggt
     6001 tacatcgaac tggatctcaa cagcggtaag atccttgaga gttttcgccc cgaagaacgt
     6061 tttccaatga tgagcacttt taaagttctg ctatgtggcg cggtattatc ccgtattgac
     6121 gccgggcaag agcaactcgg tcgccgcata cactattctc agaatgactt ggttgagtac
     6181 tcaccagtca cagaaaagca tcttacggat ggcatgacag taagagaatt atgcagtgct
     6241 gccataacca tgagtgataa cactgcggcc aacttacttc tgacaacgat cggaggaccg
     6301 aaggagctaa ccgctttttt gcacaacatg ggggatcatg taactcgcct tgatcgttgg
     6361 gaaccggagc tgaatgaagc cataccaaac gacgagcgtg acaccacgat gcctgtagca
     6421 atggcaacaa cgttgcgcaa actattaact ggcgaactac ttactctagc ttcccggcaa
     6481 caattaatag actggatgga ggcggataaa gttgcaggac cacttctgcg ctcggccctt
     6541 ccggctggct ggtttattgc tgataaatct ggagccggtg agcgtgggtc tcgcggtatc
     6601 attgcagcac tggggccaga tggtaagccc tcccgtatcg tagttatcta cacgacgggg
     6661 agtcaggcaa ctatggatga acgaaataga cagatcgctg agataggtgc ctcactgatt
     6721 aagcattggt aactgtcaga ccaagtttac tcatatatac tttagattga tttaaaactt
     6781 catttttaat ttaaaaggat ctaggtgaag atcctttttg ataatctcat gaccaaaatc
     6841 ccttaacgtg agttttcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct
     6901 tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta
     6961 ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc
     7021 ttcagcagag cgcagatacc aaatactgtt cttctagtgt agccgtagtt aggccaccac
     7081 ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct
     7141 gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat
     7201 aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg
     7261 acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa
     7321 gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggag
//
