LOCUS       sequence-328374-        3759 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For purification of Venus fluorophore.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3759)
  AUTHORS   Osterlund EJ, Hirmiz N, Pemberton JM, Nougarede A, Liu Q, Leber B, 
            Fang Q, Andrews DW
  TITLE     Efficacy and specificity of inhibitors of BCL-2 family protein 
            interactions assessed by affinity measurements in live cells.
  JOURNAL   Sci Adv. 2022 Apr 22;8(16):eabm7375. doi: 10.1126/sciadv.abm7375. 
            Epub 2022 Apr 20.
  PUBMED    35442739
REFERENCE   2  (bases 1 to 3759)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Sci Adv."; 
            date: "2022-04-22"; volume: "8(16):eabm7375. doi"; pages: " 
            10.1126/sciadv.abm7375. Epub 2022 Apr 20"
COMMENT     Sequence Label: His6-TEV-Venus-pBluescript
FEATURES             Location/Qualifiers
     source          1..3759
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(33..488)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(170..191)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     382..401
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     615..637
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     629..646
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     630..646
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     653..672
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        653..671
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             716..738
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             749..766
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             794..814
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             join(818..820,821..823,824..1537)
                     /codon_start=1
                     /product="yellow fluorescent protein (YFP) with fast and 
                     efficient maturation (Nagai et al., 2002)"
                     /label=Venus
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KLICTTGKLPVPWPTLVTTLGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIK
                     ANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     ERVTAAGITLGMDELYK"
     primer_bind     complement(863..884)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1124..1143)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     promoter        complement(1601..1619)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(1640..1656)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1640..1656)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1653..1675)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1664..1680
                     /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(1688..1694,1695..1712,1713..1718))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1733..1754
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1871..1888)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2042..2630)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2122..2141)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2801..3592,3593..3661))
                     /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     3424..3443
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(3662..3759,1..7))
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ttccgcgcac atttccccga aaagtgccac ctaaattgta agcgttaata ttttgttaaa
       61 attcgcgtta aatttttgtt aaatcagctc attttttaac caataggccg aaatcggcaa
      121 aatcccttat aaatcaaaag aatagaccga gatagggttg agtgttgttc cagtttggaa
      181 caagagtcca ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa ccgtctatca
      241 gggcgatggc ccactacgtg aaccatcacc ctaatcaagt tttttggggt cgaggtgccg
      301 taaagcacta aatcggaacc ctaaagggag cccccgattt agagcttgac ggggaaagcc
      361 ggcgaacgtg gcgagaaagg aagggaagaa agcgaaagga gcgggcgcta gggcgctggc
      421 aagtgtagcg gtcacgctgc gcgtaaccac cacacccgcc gcgcttaatg cgccgctaca
      481 gggcgcgtcc cattcgccat tcaggctgcg caactgttgg gaagggcgat cggtgcgggc
      541 ctcttcgcta ttacgccagc tggcgaaagg gggatgtgct gcaaggcgat taagttgggt
      601 aacgccaggg ttttcccagt cacgacgttg taaaacgacg gccagtgaat tgtaatacga
      661 ctcactatag ggcgaattgg agctccaccg cggtggcggc cgctctagaa ataattttgt
      721 ttaactttaa gaaggagata taaccatgca tcaccaccac catcacggcg gcagcggcgg
      781 tacgggcggt agcgagaact tatactttca aggcaccatg gtgagcaagg gcgaggagct
      841 gttcaccggg gtggtgccca tcctggtcga gctggacggc gacgtaaacg gccacaagtt
      901 cagcgtgtcc ggcgagggcg agggcgatgc cacctacggc aagctgaccc tgaagctgat
      961 ctgcaccacc ggcaagctgc ccgtgccctg gcccaccctc gtgaccaccc tgggctacgg
     1021 cctgcagtgc ttcgcccgct accccgacca catgaagcag cacgacttct tcaagtccgc
     1081 catgcccgaa ggctacgtcc aggagcgcac catcttcttc aaggacgacg gcaactacaa
     1141 gacccgcgcc gaggtgaagt tcgagggcga caccctggtg aaccgcatcg agctgaaggg
     1201 catcgacttc aaggaggacg gcaacatcct ggggcacaag ctggagtaca actacaacag
     1261 ccacaacgtc tatatcaccg ccgacaagca gaagaacggc atcaaggcca acttcaagat
     1321 ccgccacaac atcgaggacg gcggcgtgca gctcgccgac cactaccagc agaacacccc
     1381 catcggcgac ggccccgtgc tgctgcccga caaccactac ctgagctacc agtccgccct
     1441 gagcaaagac cccaacgaga agcgcgatca catggtcctg ctggaacgcg tgaccgccgc
     1501 cgggatcact ctcggcatgg acgagctgta caagtaagga ctcagatcca cgcgtatcga
     1561 ttgtcgacct cgaggggggg cccggtacca gcttttgttc cctttagtga gggttaattt
     1621 cgagcttggc gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa
     1681 ttccacacaa catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga
     1741 gctaactcac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt
     1801 gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct
     1861 cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat
     1921 cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga
     1981 acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt
     2041 ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt
     2101 ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc
     2161 gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa
     2221 gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct
     2281 ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta
     2341 actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg
     2401 gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc
     2461 ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta
     2521 ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg
     2581 gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt
     2641 tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg
     2701 tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta
     2761 aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg
     2821 aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg
     2881 tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc
     2941 gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg
     3001 agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg
     3061 aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag
     3121 gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat
     3181 caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc
     3241 cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc
     3301 ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa
     3361 ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac
     3421 gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt
     3481 cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc
     3541 gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa
     3601 caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca
     3661 tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat
     3721 acatatttga atgtatttag aaaaataaac aaatagggg
//
