LOCUS       sequence-403142-        5501 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of human GID4 with C-terminal NanoLuciferase 
            fusion.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5501)
  AUTHORS   Owens DGD, Maitland E.R, Yazdi AK, Song X, Schwalm MP, Machado RAC, 
            Bauer N , Wang X, Szewczyk MM, Dong C , Dong A, Loppnau P, Calabrese
            MF, Dowling MS, Lee J, Montgomery JI, O'Connell TN, Subramanyam C, 
            Wang F, Schapira M, Knapp S , Vedadi M, Min J, Lajoie GA, 
            Barsyte-Lovejoy D , Owen DR, Schild-Poulter C, Arrowsmith CH
  TITLE     A chemical probe to modulate human GID4 Pro/N-degron interactions
  JOURNAL   bioRxiv 2023.01.17.524225
REFERENCE   2  (bases 1 to 5501)
  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 
            2023.01.17.524225"
COMMENT     Sequence Label: GID4-Nluc
FEATURES             Location/Qualifiers
     source          1..5501
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        138..517
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        518..721
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     672..692
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     intron          857..989
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     972..991
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     primer_bind     1033..1052
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1033..1051
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1980..2492
                     /codon_start=1
                     /product="NanoLuc(R) luciferase"
                     /label=Nluc
                     /note="human codon-optimized"
                     /translation="VFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQRI
                     VLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGVT
                     PNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGWR
                     LCERILA"
     primer_bind     complement(2580..2598)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     polyA_signal    2635..2756
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2646..2665)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2700..2719
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        3001..3358
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3001..3192
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3024..3044)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3209..3344
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     promoter        3373..3420
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             3452..4246
                     /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(3506..3525)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4116..4135
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4310..4358
                     /note="synthetic polyadenylation signal"
     rep_origin      4515..5060
                     /direction=RIGHT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     4961..4980
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5177..5194
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 tcaatattgg ccattagcca tattattcat tggttatata gcataaatca atattggcta
       61 ttggccattg catacgttgt atctatatca taatatgtac atttatattg gctcatgtcc
      121 aatatgaccg ccatgttggc attgattatt gactagttat taatagtaat caattacggg
      181 gtcattagtt catagcccat atatggagtt ccgcgttaca taacttacgg taaatggccc
      241 gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt atgttcccat
      301 agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac ggtaaactgc
      361 ccacttggca gtacatcaag tgtatcatat gccaagtccg ccccctattg acgtcaatga
      421 cggtaaatgg cccgcctggc attatgccca gtacatgacc ttacgggact ttcctacttg
      481 gcagtacatc tacgtattag tcatcgctat taccatggtg atgcggtttt ggcagtacac
      541 caatgggcgt ggatagcggt ttgactcacg gggatttcca agtctccacc ccattgacgt
      601 caatgggagt ttgttttggc accaaaatca acgggacttt ccaaaatgtc gtaataaccc
      661 cgccccgttg acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata taagcagagc
      721 tggtttagtg aaccgtcaga tcactagaag ctttattgcg gtagtttatc acagttaaat
      781 tgctaacgca gtcagtgctt ctgacacaac agtctcgaac ttaagctgca gaagttggtc
      841 gtgaggcact gggcaggtaa gtatcaaggt tacaagacag gtttaaggag accaatagaa
      901 actgggcttg tcgagacaga gaagactctt gcgtttctga taggcaccta ttggtcttac
      961 tgacatccac tttgcctttc tctccacagg tgtccactcc cagttcaatt acagctctta
     1021 aggctagagt attaatacga ctcactatag ggctagcgat cgccatgtcc tgtgcgcgag
     1081 ggcaagtcgg gagggggacc cagctcagga ctgggaggcc ctgctcgcag gtccctgggt
     1141 cccggtggcg gccggagcgc ttgctccgca ggcagcgggc gggtggtcgc ccctcccgcc
     1201 cccaccccgc gcgtgcgcgc cccggcctct ccctccccgc caccctcctc ggctcccgcg
     1261 cggcggcggc ggttcctctc ccactccccc cagccctggc tccgggggac cccgcgatgc
     1321 cggtccgcac cgagtgtccc ccgccggccg gtgcctccgc tgcctccgcg gcctcactca
     1381 tcccgccgcc gcccatcaac acccagcagc ccggcgtggc caccagcctg ctctacagcg
     1441 gctccaagtt ccgcggccac cagaagagca aggggaactc gtacgacgta gaggtggtgc
     1501 tgcagcacgt ggacacgggg aactcttacc tttgtgggta cttgaagatt aaaggcctta
     1561 ctgaggagta tccaaccctt acaaccttct tcgaaggaga aataatcagc aaaaaacacc
     1621 ctttcttaac tcgcaagtgg gatgcagatg aagatgttga tcggaaacac tggggcaagt
     1681 ttctggcttt ttatcagtat gcaaaatcat ttaactcaga tgactttgat tatgaagagc
     1741 tgaagaatgg agactacgtc ttcatgaggt ggaaggaaca gtttctggtc ccagatcaca
     1801 cgatcaaaga catcagtggt gcttcttttg ccgggttcta ctacatctgc tttcagaagt
     1861 cagcagcctc catagagggc tactactacc ataggagttc agaatggtat cagtccctca
     1921 atctaaccca tgttcctgaa cacagtgcac ccatctatga attccggggc tcgagcggcg
     1981 tcttcacact cgaagatttc gttggggact ggcgacagac agccggctac aacctggacc
     2041 aagtccttga acagggaggt gtgtccagtt tgtttcagaa tctcggggtg tccgtaactc
     2101 cgatccaaag gattgtcctg agcggtgaaa atgggctgaa gatcgacatc catgtcatca
     2161 tcccgtatga aggtctgagc ggcgaccaaa tgggccagat cgaaaaaatt tttaaggtgg
     2221 tgtaccctgt ggatgatcat cactttaagg tgatcctgca ctatggcaca ctggtaatcg
     2281 acggggttac gccgaacatg atcgactatt tcggacggcc gtatgaaggc atcgccgtgt
     2341 tcgacggcaa aaagatcact gtaacaggga ccctgtggaa cggcaacaaa attatcgacg
     2401 agcgcctgat caaccccgac ggctccctgc tgttccgagt aaccatcaac ggagtgaccg
     2461 gctggcggct gtgcgaacgc attctggcgt aaggccgcga ctctagagtc gacctgcagg
     2521 catgcaagct gatccggctg ctaacaaagc ccgaaaggaa gctgagttgg ctgctgccac
     2581 cgctgagcaa taactagcat aaccccttgg ggcggccgct tcgagcagac atgataagat
     2641 acattgatga gtttggacaa accacaacta gaatgcagtg aaaaaaatgc tttatttgtg
     2701 aaatttgtga tgctattgct ttatttgtaa ccattataag ctgcaataaa caagttaaca
     2761 acaacaattg cattcatttt atgtttcagg ttcaggggga gatgtgggag gtttttttaa
     2821 gcaagtaaaa cctctacaaa tgtggtaaaa tcgaatttta acaaaatatt aacgcttaca
     2881 atttcctgat gcggtatttt ctccttacgc atctgtgcgg tatttcacac cgcatacgcg
     2941 gatctgcgca gcaccatggc ctgaaataac ctctgaaaga ggaacttggt taggtacctt
     3001 ctgaggcgga aagaaccagc tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg
     3061 ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg
     3121 aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc
     3181 aaccatagtc ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca
     3241 ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg ccgcctcggc
     3301 ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctaggct tttgcaaaaa
     3361 gcttaattaa ctgttgacaa ttaatcatcg gcatagtata tcggcatagt ataatacgac
     3421 aaggtgagga actaaaccca ggaggcagat catgattgaa caagatggat tgcacgcagg
     3481 ttctccggcc gcttgggtgg agaggctatt cggctatgac tgggcacaac agacaatcgg
     3541 ctgctctgat gccgccgtgt tccggctgtc agcgcagggg cgcccggttc tttttgtcaa
     3601 gaccgacctg tccggtgccc tgaatgaact gcaggacgag gcagcgcggc tatcgtggct
     3661 ggccacgacg ggcgttcctt gcgcagctgt gctcgacgtt gtcactgaag cgggaaggga
     3721 ctggctgcta ttgggcgaag tgccggggca ggatctcctg tcatctcacc ttgctcctgc
     3781 cgagaaagta tccatcatgg ctgatgcaat gcggcggctg catacgcttg atccggctac
     3841 ctgcccattc gaccaccaag cgaaacatcg catcgagcga gcacgtactc ggatggaagc
     3901 cggtcttgtc gatcaggatg atctggacga agagcatcag gggctcgcgc cagccgaact
     3961 gttcgccagg ctcaaggcgc gcatgcccga cggcgaggat ctcgtcgtga cccatggcga
     4021 tgcctgcttg ccgaatatca tggtggaaaa tggccgcttt tctggattca tcgactgtgg
     4081 ccggctgggt gtggcggacc gctatcagga catagcgttg gctacccgtg atattgctga
     4141 agagcttggc ggcgaatggg ctgaccgctt cctcgtgctt tacggtatcg ccgctcccga
     4201 ttcgcagcgc atcgccttct atcgccttct tgacgagttc ttctgagcgg gactctgggg
     4261 ttcgaaatga ccgaccaagc gacgcccaac ctgccatcac gatggccgca ataaaatatc
     4321 tttattttca ttacatctgt gtgttggttt tttgtgtgaa tcgatagcga taaggatccg
     4381 ctcatgcgat gataagctgt caaacatgag aattacaact tatatcgtat ggggctgact
     4441 tcaggtgcta catttgaaga gataaattgc actgaaatct agaaatattt tatctgatta
     4501 ataagatgat cttcttgaga tcgttttggt ctgcgcgtaa tctcttgctc tgaaaacgaa
     4561 aaaaccgcct tgcagggcgg tttttcgaag gttctctgag ctaccaactc tttgaaccga
     4621 ggtaactggc ttggaggagc gcagtcacca aaacttgtcc tttcagttta gccttaaccg
     4681 gcgcatgact tcaagactaa ctcctctaaa tcaattacca gtggctgctg ccagtggtgc
     4741 ttttgcatgt ctttccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc
     4801 ggactgaacg gggggttcgt gcatacagtc cagcttggag cgaactgcct acccggaact
     4861 gagtgtcagg cgtggaatga gacaaacgcg gccataacag cggaatgaca ccggtaaacc
     4921 gaaaggcagg aacaggagag cgcacgaggg agccgccagg gggaaacgcc tggtatcttt
     4981 atagtcctgt cgggtttcgc caccactgat ttgagcgtca gatttcgtga tgcttgtcag
     5041 gggggcggag cctatggaaa aacggctttg ccgcggccct ctcacttccc tgttaagtat
     5101 cttcctggca tcttccagga aatctccgcc ccgttcgtaa gccatttccg ctcgccgcag
     5161 tcgaacgacc gagcgtagcg agtcagtgag cgaggaagcg gaatatatcc tgtatcacat
     5221 attctgctga cgcaccggtg cagccttttt tctcctgcca catgaagcac ttcactgaca
     5281 ccctcatcag tgccaacata gtaagccagt atacactccg ctagcgctga tgtccggcgg
     5341 tgcttttgcc gttacgcacc accccgtcat gagattacgc gttttagcgg tgcgtacaat
     5401 taagggatta tggtaaatcc acttactgtc tgccctcgta gccatcgaga taaaccgcag
     5461 tactccggcc acgatgcgtc cggcgtagag gatcgagatc t
//
