LOCUS       sequence-84161-e        7107 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression plasmid for human GTPBP4 with n-terminal 3xFLAG
            epitope fused..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7107)
  AUTHORS   Li S, Wang L, Fu B, Berman MA, Diallo A, Dorf ME
  TITLE     TRIM65 regulates microRNA activity by ubiquitination of TNRC6.
  JOURNAL   Proc Natl Acad Sci U S A. 2014 May 13;111(19):6970-5. doi: 
            10.1073/pnas.1322545111. Epub 2014 Apr 28.
  PUBMED    24778252
REFERENCE   2  (bases 1 to 7107)
  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"; doi: 
            "10.1073/pnas.1322545111"; journalName: "Proc Natl Acad Sci U S A"; 
            date: "2014-05-13- 13"; volume: "111"; issue: "19"; pages: "6970-5"
COMMENT     Sequence Label: pCMV3FLAG_hGTPBP4
FEATURES             Location/Qualifiers
     source          1..7107
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        67..370
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        371..574
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     525..545
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     promoter        620..638
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     regulatory      673..682
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             682..753
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags"
                     /label=3xFLAG
                     /translation="DYKDDDDKDYKDDDDKDYKDDDDK"
     primer_bind     complement(2739..2758)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2740..2758)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2787..2804)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2793..3000
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      complement(3007..3462)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3144..3165)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3356..3375
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3489..3593
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3594..3662,3663..4451)
                     /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(3812..3831)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        4545..4690
                     /label=HSV TK promoter
                     /note="herpes simplex virus thymidine kinase promoter"
     CDS             4733..5770
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPDRE
                     MGEAN"
     primer_bind     complement(5768..5787)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5812..5860
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      6411..6999
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6900..6919
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 atgcattagt tattaatagt aatcaattac ggggtcatta gttcatagcc catatatgga
       61 gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca acgacccccg
      121 cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga ctttccattg
      181 acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc aagtgtatca
      241 tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct ggcattatgc
      301 ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat tagtcatcgc
      361 tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc ggtttgactc
      421 acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt ggcaccaaaa
      481 tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag
      541 gcgtgtacgg tgggaggtct atataagcag agctggttta gtgaaccgtc agatccgcta
      601 gcgattacgc caagctcgaa attaaccctc actaaaggga acaaaagctg gagctccacc
      661 gcggtggcgg ccgccaccat ggattacaag gatgacgacg ataaggacta taaggacgat
      721 gatgacaagg actacaaaga tgatgacgat aaagcccggg cgggatcccc cgggctgcag
      781 gaattcgata tcgcacatta caacttcaag aaaattacgg tggtgccgtc cgccaaggac
      841 ttcatagacc tcacgttgtc gaagactcaa cgaaagactc caaccgttat tcataaacat
      901 taccaaatac atcgcattag acatttttac atgagaaaag tcaaatttac tcaacagaat
      961 taccatgata gactttcaca aattctaaca gatttcccca aattggatga tattcatccg
     1021 ttctatgctg atttgatgaa tattctctac gacaaggatc attacaagtt ggctctgggg
     1081 caaataaata ttgccaaaaa tttagtggac aatgttgcta aagattatgt gcgactgatg
     1141 aagtatggcg actctctcta ccgctgcaaa cagctgaagc gtgcggccct gggacggatg
     1201 tgcacagtga tcaagaggca gaagcagagt ttggagtatt tggagcaagt gcgtcagcat
     1261 ttatcccgtt tgccaaccat tgatccgaat accaggaccc tgcttttgtg tgggtaccca
     1321 aatgttggga agtccagctt cgtcaacaag gtgacgagag cagacgtgga tgtccagccc
     1381 tatgcgttca caaccaagtc tctgtttgtt gggcacatgg attataagta tctacgttgg
     1441 caggttgtag acactcctgg gatcctggac caccctctgg aggataggaa caccatcgag
     1501 atgcaggcca tcactgccct ggcccacctc cgtgctgcgg tcctgtatgt gatggatttg
     1561 tctgagcagt gtgggcatgg gctgagggag cagctagaac tcttccagaa catcagacct
     1621 ctcttcatca acaagcctct catagttgta gccaacaaat gtgatgtgaa gagaatagct
     1681 gaactttctg aagatgatca gaaaatattt acagatttgc agtctgaagg attccctgta
     1741 atagagacca gcaccctgac tgaggaaggt gttattaaag ttaaaacaga ggcttgcgat
     1801 aggcttttgg ctcatcgagt ggaaaccaaa atgaagggaa ataaagtgaa tgaggtgctg
     1861 aatagactgc acctggctat cccaaccagg agggacgata aggagaggcc ccctttcatc
     1921 cctgaaggag tggtggctcg caggaagagg atggaaactg aggagtccag gaagaagagg
     1981 gaacgagatc ttgagctgga aatgggagat gattatattt tggatcttca gaagtactgg
     2041 gatttaatga atttgtctga aaaacatgat aagataccag aaatctggga aggccataat
     2101 atagctgatt atattgatcc agccatcatg aagaaattgg aagaattaga aaaagaagaa
     2161 gagctgagaa cagctgctgg agagtatgac agtgtatctg agagtgaaga cgaagagatg
     2221 ctggaaatcc gacagctggc aaagcaaatt cgagagaaaa agaagttgaa aattctggag
     2281 tccaaagaaa agaatacaca gggacccagg atgccgcgaa ctgctaagaa ggttcagagg
     2341 acagttttgg agaaggagat gcatagtctt ggtgttgaca tggacgataa agacgatgcc
     2401 cattacgcag tccaggcaag aagatcccgg agcatcacta ggaaaagaag gcgggaagac
     2461 tctgctcccc catcctctgt ggcccggagt gggagttgct ctcgaactcc acgtgacgtt
     2521 tctggtctta gggatgtcaa gatggtgaag aaagccaaga ctatgatgaa gaatgctcag
     2581 aagaagatga atcggttggg gaagaaaggg gaggcggata gacacgtgtt tgatatgaag
     2641 cccaagcact tgctgtctgg gaagaggaaa gctggtaaaa aggacaggag ataggatacc
     2701 ttaattaatt aaggtaccag gtaagtgtac ccaattcgcc ctatagtgag tcgtattaca
     2761 attcactcga tcggctcgct gatcagcctc gactgtgcct tctagttgcc agccatctgt
     2821 tgtttgcccc tcccccgtgc cttccttgac cctggaaggt gccactccca ctgtcctttc
     2881 ctaataaaat gaggaaattg catcgcattg tctgagtagg tgtcattcta ttctgggggg
     2941 tggggtgggg caggacagca agggggagga ttgggaagac aatagcaggc atgctgggga
     3001 acgcgtaaat tgtaagcgtt aatattttgt taaaattcgc gttaaatttt tgttaaatca
     3061 gctcattttt taaccaatag gccgaaatcg gcaaaatccc ttataaatca aaagaataga
     3121 ccgagatagg gttgagtgtt gttccagttt ggaacaagag tccactatta aagaacgtgg
     3181 actccaacgt caaagggcga aaaaccgtct atcagggcga tggcccacta cgtgaaccat
     3241 caccctaatc aagttttttg gggtcgaggt gccgtaaagc actaaatcgg aaccctaaag
     3301 ggagcccccg atttagagct tgacggggaa agccggcgaa cgtggcgaga aaggaaggga
     3361 agaaagcgaa aggagcgggc gctagggcgc tggcaagtgt agcggtcacg ctgcgcgtaa
     3421 ccaccacacc cgccgcgctt aatgcgccgc tacagggcgc gtcaggtggc acttttcggg
     3481 gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc
     3541 tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag agtatgagta
     3601 ttcaacattt ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg
     3661 ctcacccaga aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg
     3721 gttacatcga actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac
     3781 gttttccaat gatgagcact tttaaagttc tgctatgtgg cgcggtatta tcccgtattg
     3841 acgccgggca agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgagt
     3901 actcaccagt cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg
     3961 ctgccataac catgagtgat aacactgcgg ccaacttact tctgacaacg atcggaggac
     4021 cgaaggagct aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt
     4081 gggaaccgga gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgtag
     4141 caatggcaac aacgttgcgc aaactattaa ctggcgaact acttactcta gcttcccggc
     4201 aacaattaat agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc
     4261 ttccggctgg ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta
     4321 tcattgcagc actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg
     4381 ggagtcaggc aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga
     4441 ttaagcattg gcctcaggat tagtgcttca tccccgtggc ccgttgctcg cgtttgctgg
     4501 cggtgtcccc ggaagaaata tatttgcatg tctttagttc tatgatgaca caaaccccgc
     4561 ccagcgtctt gtcattggcg aattcgaaca cgcagatgca gtcggggcgg cgcggtccga
     4621 ggtccacttc gcatattaag gtgacgcgtg tggcctcgaa caccgagcga ccctgcagcg
     4681 acccgcttaa cagcgtcaac agcgtgccgc agatcccggg gggcaatgag atatgaaaaa
     4741 gcctgaactc accgcgacgt ctgtcgagaa gtttctgatc gaaaagttcg acagcgtctc
     4801 cgacctgatg cagctctcgg agggcgaaga atctcgtgct ttcagcttcg atgtaggagg
     4861 gcgtggatat gtcctgcggg taaatagctg cgccgatggt ttctacaaag atcgttatgt
     4921 ttatcggcac tttgcatcgg ccgcgctccc gattccggaa gtgcttgaca ttggggaatt
     4981 cagcgagagc ctgacctatt gcatctcccg ccgtgcacag ggtgtcacgt tgcaagacct
     5041 gcctgaaacc gaactgcccg ctgttctgca gccggtcgcg gaggccatgg atgcgatcgc
     5101 tgcggccgat cttagccaga cgagcgggtt cggcccattc ggaccgcaag gaatcggtca
     5161 atacactaca tggcgtgatt tcatatgcgc gattgctgat ccccatgtgt atcactggca
     5221 aactgtgatg gacgacaccg tcagtgcgtc cgtcgcgcag gctctcgatg agctgatgct
     5281 ttgggccgag gactgccccg aagtccggca cctcgtgcac gcggatttcg gctccaacaa
     5341 tgtcctgacg gacaatggcc gcataacagc ggtcattgac tggagcgagg cgatgttcgg
     5401 ggattcccaa tacgaggtcg ccaacatctt cttctggagg ccgtggttgg cttgtatgga
     5461 gcagcagacg cgctacttcg agcggaggca tccggagctt gcaggatcgc cgcggctccg
     5521 ggcgtatatg ctccgcattg gtcttgacca actctatcag agcttggttg acggcaattt
     5581 cgatgatgca gcttgggcgc agggtcgatg cgacgcaatc gtccgatccg gagccgggac
     5641 tgtcgggcgt acacaaatcg cccgcagaag cgcggccgtc tggaccgatg gctgtgtaga
     5701 agtactcgcc gatagtggaa accgacgccc cagcactcgt ccggatcggg agatggggga
     5761 ggctaactga aacacggaag gagacaatac cggaaggaac ccgcgctatg acggcaataa
     5821 aaagacagaa taaaacgcac gggtgttggg tcgtttgttc ataaacgcgg ggttcggtcc
     5881 cagggctggc actctgtcga taccccaccg agaccccatt ggggccaata cgcccgcgtt
     5941 tcttcctttt ccccacccca ccccccaagt tcgggtgaag gcccagggct cgcagccaac
     6001 gtcggggcgg caggccctgc catagccact ggccccgtgg gttagggacg gggtccccca
     6061 tggggaatgg tttatggttc gtgggggtta ttattttggg cgttgcgtgg ggtcaggtcc
     6121 acgactggac tgagcagaca gacccatggt ttttggatgg cctgggcatg gaccgcatgt
     6181 actggcgcga cacgaacacc gggcgtctgt ggctgccaaa cacccccgac ccccaaaaac
     6241 caccgcgcgg atttctggcg ccgccggacg aactaaacct gactacgaca tctctgcccc
     6301 ttcttcgctg gtacgaggag cgctttgcct caggtcatga ccaaaatccc ttaacgtgag
     6361 ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct
     6421 ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt
     6481 tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg
     6541 cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt caagaactct
     6601 gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc
     6661 gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg
     6721 tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa
     6781 ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg
     6841 gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg
     6901 ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga
     6961 tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt
     7021 ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct
     7081 gattctgtgg ataaccgtat taccgcc
//
