LOCUS       sequence-496358-        4356 bp    DNA     circular SYN 07-JAN-2026
DEFINITION  pComb3X plasmid engineered for golden gate insertion of antibodies.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4356)
  TITLE     Structure-based design of antibody repertoires with drug-like 
            properties
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 4356)
  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: 
            "Unpublished"
COMMENT     Sequence Label: pComb3X-goldengatefriendly
FEATURES             Location/Qualifiers
     source          1..4356
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      77..665
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     566..585
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     819..836
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    953..974
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(989..994,995..1012,1013..1019)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1027..1043
                     /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)."
     sig_peptide     1066..1128
                     /codon_start=1
                     /product="signal peptide from the E. coli outer membrane 
                     protein OmpA"
                     /label=OmpA signal peptide
                     /translation="MKKTAIAIAVALAGFATVAQA"
     CDS             join(1140..1142,1143..1145,1146..1859)
                     /codon_start=1
                     /product="enhanced GFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mEGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GCYKTRAEVKFEGDTLVNRIELHGIDFKEDGNILGHKLEYNFNSHNVYIMPDKQNNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDYHYLHTWSELSKDPNEKRDHMVLL
                     EFVEAAGITLGMDELYK"
     primer_bind     complement(1185..1206)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     1793..1814
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             2213..2230
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     rep_origin      complement(2815..3270)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2952..2973)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3164..3183
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3297..3401
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3402..3470,3471..4262)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGTRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(3620..3639)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa
       61 agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa
      121 aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc
      181 cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt
      241 agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc
      301 tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac
      361 gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca
      421 gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg
      481 ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag
      541 gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt
      601 ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat
      661 ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc
      721 acatgttctt tcctgcgtta tcccctgatt ctgtggataa ccgtattacc gcctttgagt
      781 gagctgatac cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg agcgaggaag
      841 cggcagagcg cccaatacgc aaaccgcctc tccccgcgcg ttggccgatt cattaatgca
      901 gctggcacga caggtttccc gactggaaag cgggcagtga gcgcaacgca attaatgtga
      961 gttagctcac tcattaggca ccccaggctt tacactttat gcttccggct cgtatgttgt
     1021 gtggaattgt gagcggataa caattgaatt caggaggaat ttaaaatgaa aaagacagct
     1081 atcgcgattg cagtggcact ggctggtttc gctaccgtgg cccaggctgc caagtcttca
     1141 tggtgagcaa gggcgaggag ctgttcaccg gggtggtgcc catcctggtc gagctggacg
     1201 gcgacgtaaa cggccacaag ttcagcgtgc gcggcgaggg cgagggcgat gccacctacg
     1261 gcaagctgac cctgaagttc atctgcacca ccggcaagct gcccgtgccc tggcccaccc
     1321 tcgtgaccac cctgacctac ggcgtccagt gcttcagccg ctaccccgac cacatgaagc
     1381 agcacgactt cttcaagtcc gccatgcccg aaggctacgt ccaggagcgc accatcttct
     1441 tcaaggacga cggctgctac aagacccgcg ccgaggtgaa gttcgagggc gacaccctgg
     1501 tgaaccgcat cgagctgcac ggcatcgact tcaaggagga cggcaacatc ctggggcaca
     1561 agctggagta caacttcaac agccacaacg tctatatcat gcccgacaag cagaacaacg
     1621 gcatcaaggt gaacttcaag atccgccaca acatcgagga cggcagcgtg cagctcgccg
     1681 accactacca gcagaacacc cccatcggcg acggccccgt gctgctgccc gactaccact
     1741 acctgcacac ctggtccgag ctgagcaaag accccaacga gaagcgcgat cacatggtcc
     1801 tgctggagtt cgtggaggcc gccgggatca ctctcggcat ggacgagctg tacaagtaag
     1861 aagacccgct agcaccaagg gcccatcggt gttccccctg gcaccctcct ccaagagcac
     1921 ctctgggggc acagcggccc tgggctgcct ggtcaaggac tacttccccg aaccggtgac
     1981 ggtgtcgtgg aactcaggcg ccctgaccag cggcgtgcac accttcccgg ctgtcctaca
     2041 gtcctcagga ctctactccc tcagcagcgt ggtgaccgtg ccctccagca gcttgggcac
     2101 ccagacctac atctgcaacg tgaatcacaa gcccagcaac accaaggtgg acaagaaagt
     2161 tgagcccaaa tcttgtgaca aaactagtcc ggactacgct tctggcggat cccatcacca
     2221 tcaccatcac taggagggtg gtggctctga gggtggcggt tctgagggtg gcggctctga
     2281 gggaggcggt tccggtggtg gctctggttc cggtgatttt gattatgaaa agatggcaaa
     2341 cgctaataag ggggctatga ccgaaaatgc cgatgaaaac gtgctacagt ctgacgctaa
     2401 aggcaaactt gattctgtcg ctactgatta cggtgctgct atcgatggtt tcattggtga
     2461 cgtttccggc cttgctaatg gtaatggtgc tactggtgat tttgctggct ctaattccca
     2521 aatggctcaa gtcggtgacg gtgataattc acctttaatg aataatttcc gtcaatattt
     2581 accttccctc cctcaatcgg ttgaatgtcg cccttttgtc tttggcgctg gtaaaccata
     2641 tgaattttct attgattgtg acaaaataaa cttattccgt ggtgtctttg cgtttctttt
     2701 atatgttgcc acctttatgt atgtattttc tacgtttgct aacatactgc gtaataagga
     2761 gtcttaataa ttaatttaag cggccgcaga tctgctctct gaggaggatc tgggaaattg
     2821 taagcgttaa tattttgtta aaattcgcgt taaatttttg ttaaatcagc tcatttttta
     2881 accaataggc cgaaatcggc aaaatccctt ataaatcaaa agaatagacc gagatagggt
     2941 tgagtgttgt tccagtttgg aacaagagtc cactattaaa gaacgtggac tccaacgtca
     3001 aagggcgaaa aaccgtctat cagggcgatg gcccactacg tgaaccatca ccctaatcaa
     3061 gttttttggg gtcgaggtgc cgtaaagcac taaatcggaa ccctaaaggg agcccccgat
     3121 ttagagcttg acggggaaag ccggcgaacg tggcgagaaa ggaagggaag aaagcgaaag
     3181 gagcgggcgc tagggcgctg gcaagtgtag cggtcacgct gcgcgtaacc accacacccg
     3241 ccgcgcttaa tgcgccgcta cagggcgcgt caggtggcac ttttcgggga aatgtgcgcg
     3301 gaacccctat ttgtttattt ttctaaatac attcaaatat gtatccgctc atgagacaat
     3361 aaccctgata aatgcttcaa taatattgaa aaaggaagag tatgagtatt caacatttcc
     3421 gtgtcgccct tattcccttt tttgcggcat tttgccttcc tgtttttgct cacccagaaa
     3481 cgctggtgaa agtaaaagat gctgaagatc agttgggtgc acgagtgggt tacatcgaac
     3541 tggatctcaa cagcggtaag atccttgaga gttttcgccc cgaagaacgt tttccaatga
     3601 tgagcacttt taaagttctg ctatgtggcg cggtattatc ccgtgttgac gccgggcaag
     3661 agcaactcgg tcgccgcata cactattctc agaatgactt ggttgagtac tcaccagtca
     3721 cagaaaagca tcttacggat ggcatgacag taagagaatt atgcagtgct gccataacca
     3781 tgagtgataa cactgcggcc aacttacttc tgacaacgat cggaggaccg aaggagctaa
     3841 ccgctttttt gcacaacatg ggggatcatg taactcgcct tgatcgttgg gaaccggagc
     3901 tgaatgaagc cataccaaac gacgagcgtg acaccacgat gcctgtagca atggcaacaa
     3961 cgttgcgcaa actattaact ggcgaactac ttactctagc ttcccggcaa caattaatag
     4021 actggatgga ggcggataaa gttgcaggac cacttctgcg ctcggccctt ccggctggct
     4081 ggtttattgc tgataaatct ggagccggtg agcgtgggac tcgcggtatc attgcagcac
     4141 tggggccaga tggtaagccc tcccgtatcg tagttatcta cacgacgggg agtcaggcaa
     4201 ctatggatga acgaaataga cagatcgctg agataggtgc ctcactgatt aagcattggt
     4261 aactgtcaga ccaagtttac tcatatatac tttagattga tttaaaactt catttttaat
     4321 ttaaaaggat ctaggtgaag atcctttttg ataatc
//
