LOCUS       sequence-445930-        2638 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  4G-cloning donor for C-terminal 3C-Venus-His8 tag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2638)
  AUTHORS   Taschner M, Dickinson JB, Roisne-Hamelin F, Gruber S
  TITLE     4G cloning: rapid gene assembly for expression of multisubunit 
            protein complexes in diverse hosts
  JOURNAL   bioRxiv 2024.06.17.599261
REFERENCE   2  (bases 1 to 2638)
  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: "bioRxiv 
            2024.06.17.599261"
COMMENT     Sequence Label: pD(Ct)_3C-Venus-His8
FEATURES             Location/Qualifiers
     source          1..2638
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             40..63
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             join(76..78,79..81,82..594)
                     /codon_start=1
                     /product="N-terminal fragment of mVenus for use in 
                     bimolecular fluorescence complementation (BiFC) (Kodama and
                     Hu, 2010)"
                     /label=VN173
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KLICTTGKLPVPWPTLVTTLGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIK
                     ANFKIRHNIE"
     primer_bind     complement(121..142)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(382..401)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     CDS             721..744
                     /codon_start=1
                     /product="8xHis affinity tag"
                     /label=8xHis
                     /translation="HHHHHHHH"
     primer_bind     765..786
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     promoter        1010..1112
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     CDS             1113..1772
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(1179..1198)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     rep_origin      2167..2523
                     /label=R6K gamma ori
                     /note="gamma replication origin from E. coli plasmid R6K; 
                     requires the R6K initiator protein pi for replication"
     protein_bind    complement(2576..2609)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
ORIGIN
        1 gattcgcgac ctactccgga atattaggtc tcaggatccc tggaagttct gttccagggg
       61 cccagcggca gcggcatggt gagcaagggc gaggagctgt tcaccggggt ggtgcccatc
      121 ctggtcgagc tggacggcga cgtaaacggc cacaagttca gcgtgtccgg cgagggcgag
      181 ggcgatgcca cctacggcaa gctgaccctg aagctgatct gcaccaccgg caagctgccc
      241 gtgccctggc ccaccctcgt gaccaccctg ggctacggcc tgcagtgctt cgcccgctac
      301 cccgaccaca tgaagcagca cgacttcttc aagtccgcca tgcccgaagg ctacgtccag
      361 gagcgcacca tcttcttcaa ggacgacggc aactacaaga cccgcgccga ggtgaagttc
      421 gagggcgaca ccctggtgaa ccgcatcgag ctgaagggca tcgacttcaa ggaggacggc
      481 aacatcctgg ggcacaagct ggagtacaac tacaacagcc acaacgtcta tatcaccgcc
      541 gacaagcaga agaacggcat caaggccaac ttcaagatcc gccacaacat cgaggacggc
      601 ggcgtgcagc tcgccgacca ctaccagcag aacactccca tcggcgacgg ccccgtgctg
      661 ctgcccgaca accactacct gagctaccag tccaaactga gcaaagaccc caacggcagc
      721 caccatcacc atcaccatca ccatgggagc gagaagcgcg atcacatggt cctgctggag
      781 ttcgtgaccg ccgccgggat cactctcggc atggacgagc tgtacaaggt ctaagggaga
      841 cccactgctt gagcctagaa gatccggctg ctaacaaagc ccgaaaggaa gctgagttgg
      901 ctgctgtcac cgctgagcaa taactatcat aacccctagg tgccatttca ttacctcttt
      961 ctccgcaccc gacatagatc tgggccaact tttggcgaaa atgagacgtt gatcggcacg
     1021 taagagggtc caactttcac cataatgaaa taagatcact accgggcgta ttttttgagt
     1081 tatcgagatt ttcaggagct aaggaagcta aaatggagaa aaaaatcact ggatatacca
     1141 ccgttgatat atcccaatgg catcgtaaag aacattttga ggcatttcag tcagttgctc
     1201 aatgtaccta taaccagacc gttcagctgg atattacggc ctttttaaag accgtaaaga
     1261 aaaataagca caagttttat ccggccttta ttcacattct tgcccgcctg atgaatgctc
     1321 atccggaatt ccgtatggca atgaaagacg gtgagctggt gatatgggat agtgttcacc
     1381 cttgttacac cgttttccat gagcaaactg aaacgttttc atcgctctgg agtgaatacc
     1441 acgacgattt ccggcagttt ctacacatat attcgcaaga tgtggcgtgt tacggtgaaa
     1501 acctggccta tttccctaaa gggtttattg agaatatgtt tttcgtctca gccaatccct
     1561 gggtgagttt caccagtttt gatttaaacg tggccaatat ggacaacttc ttcgcccccg
     1621 ttttcaccat gggcaaatat tatacgcaag gcgacaaggt gctgatgccg ctggcgattc
     1681 aggttcatca tgccgtttgt gatggcttcc atgtcggcag aatgcttaat gaattacaac
     1741 agtactgcga tgagtggcag ggtggggcgt aattttttta aggcagttat tggtgccctt
     1801 aaacgcctgg ttgctacgcc tgaataagtg ataataagcg gatgaatggc agaaattcga
     1861 aagcaaattc gacccggtcg tcggttcagg gcagggtcgt taaatagccg cttatgtcta
     1921 ttgctggttt accggtttat tgactaccgg aagcagtgtg accgtgtgct tctcaaatgc
     1981 ctgaggccag tttgctcagg ctctccccgt ggaggtaata attgacgata tgatcattta
     2041 ttctgcctcc cagctgacat tcatccgggg tcagcaccgt ttctgcggac tggctttcta
     2101 cgtgttccgc ttcctttagc agcccttgcg ccctgagtgc ttgcggcagc gtgaagctaa
     2161 ttcccatgtc agccgttaag tgttcctgtg tcactcaaaa ttgctttgag aggctctaag
     2221 ggcttctcag tgcgttacat ccctggcttg ttgtccacaa ccgttaaacc ttaaaagctt
     2281 taaaagcctt atatattctt ttttttctta taaaacttaa aaccttagag gctatttaag
     2341 ttgctgattt atattaattt tattgttcaa acatgagagc ttagtacgtg aaacatgaga
     2401 gcttagtacg ttagccatga gagcttagta cgttagccat gagggtttag ttcgttaaac
     2461 atgagagctt agtacgttaa acatgagagc ttagtacgtg aaacatgaga gcttagtacg
     2521 tactatcaac aggttgaact gctgatcaac agatcctcta cgcggccgcg gtaccataac
     2581 ttcgtatagc atacattata cgaagttatc tgccaggcac atgggtttta ctagtatc
//
