LOCUS       sequence-396911-        6344 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of mutated Translocon-associated protein 
            subunit alpha.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6344)
  AUTHORS   Karki S, Javanainen M, Rehan S, Tranter D, Kellosalo J, Huiskonen 
            JT, Happonen L, Paavilainen V
  TITLE     Molecular view of ER membrane remodeling by the Sec61/TRAP 
            translocon.
  JOURNAL   EMBO Rep. 2023 Dec 6;24(12):e57910. doi: 10.15252/embr.202357910. 
            Epub 2023 Nov 20.
  PUBMED    37983950
REFERENCE   2  (bases 1 to 6344)
  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: "EMBO Rep.";
            date: "2023-12-6"; volume: "24(12):e57910. doi"; pages: " 
            10.15252/embr.202357910. Epub 2023 Nov 20"
COMMENT     Sequence Label: TRAPa(W255A,N262E,K266E)-3xFLAG-pCDNA
FEATURES             Location/Qualifiers
     source          1..6344
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     5..24
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(243..347)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(422..441)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        613..992
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        993..1196
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1147..1167
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     1241..1260
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1241..1259
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             2200..2265
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     primer_bind     complement(2313..2330)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2319..2543
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2589..3017
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2676..2695)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2886..2907
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3026..3046)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3031..3360
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3211..3346
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3273..3292
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3427..4221
                     /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(3481..3500)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4091..4110
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4395..4516
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4432..4451)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4486..4505
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4565..4581)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4565..4581)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4578..4600)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4589..4605
                     /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(4613..4619,4620..4637,4638..4643))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4658..4679
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4796..4813)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4967..5555)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5047..5066)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5726..6344,1..173,174..242))
                     /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"
ORIGIN
        1 cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct
       61 tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact
      121 cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa
      181 acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc
      241 atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga
      301 tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga
      361 aaagtgccac ctgacgtcga cggatcggga gatctcccga tcccctatgg tgcactctca
      421 gtacaatctg ctctgatgcc gcatagttaa gccagtatct gctccctgct tgtgtgttgg
      481 aggtcgctga gtagtgcgcg agcaaaattt aagctacaac aaggcaaggc ttgaccgaca
      541 attgcatgaa gaatctgctt agggttaggc gttttgcgct gcttcgcgat gtacgggcca
      601 gatatacgcg ttgacattga ttattgacta gttattaata gtaatcaatt acggggtcat
      661 tagttcatag cccatatatg gagttccgcg ttacataact tacggtaaat ggcccgcctg
      721 gctgaccgcc caacgacccc cgcccattga cgtcaataat gacgtatgtt cccatagtaa
      781 cgccaatagg gactttccat tgacgtcaat gggtggagta tttacggtaa actgcccact
      841 tggcagtaca tcaagtgtat catatgccaa gtacgccccc tattgacgtc aatgacggta
      901 aatggcccgc ctggcattat gcccagtaca tgaccttatg ggactttcct acttggcagt
      961 acatctacgt attagtcatc gctattacca tggtgatgcg gttttggcag tacatcaatg
     1021 ggcgtggata gcggtttgac tcacggggat ttccaagtct ccaccccatt gacgtcaatg
     1081 ggagtttgtt ttggcaccaa aatcaacggg actttccaaa atgtcgtaac aactccgccc
     1141 cattgacgca aatgggcggt aggcgtgtac ggtgggaggt ctatataagc agagctctct
     1201 ggctaactag agaacccact gcttactggc ttatcgaaat taatacgact cactataggg
     1261 agacccaagc tggctagcgt ttaaacttaa gcttggtacc gagctcggat ccactagtcc
     1321 agtgtggtgg aattcgccac catgagactc ctcccccgct tgctgctgct tctcttactc
     1381 gtgttccctg ccactgtctt gttccgaggc ggccccagag gcttgttagc agtggcacaa
     1441 gatcttacag aggatgaaga aacagtagaa gattccataa ttgaggatga agatgatgaa
     1501 gccgaggtag aagaagatga acccacagat ttggtagaag ataaagagga agaagatgtg
     1561 tctggtgaac ctgaagcttc accgagtgca gatacaacta tactgtttgt aaaaggagaa
     1621 gattttccag caaataacat tgtgaagttc ctggtaggct ttaccaacaa gggtacagaa
     1681 gattttattg ttgaatcctt agatgcctca ttccgttatc ctcaggacta ccagttttat
     1741 atccagaatt tcacagctct tcctctgaac actgtagtgc caccccagag acaggcaact
     1801 tttgagtact ctttcattcc tgcagagccc atgggcggac gaccatttgg tttggtcatc
     1861 aatctgaact acaaagattt gaacggcaat gtattccaag atgcagtctt caatcaaaca
     1921 gttacagtta ttgaaagaga ggatgggtta gatggagaaa caatctttat gtatatgttc
     1981 cttgctggtc ttgggcttct ggttattgtt ggccttcatc aactcctaga atctagaaag
     2041 cgtaagagac ccatacagaa agtagaaatg ggtacatcaa gtcagaatga tgttgacatg
     2101 agtgcgattc ctcaggaaac attggaacaa atcaatgaag cttcaccaag aaggttgccc
     2161 aggaaacggg cacagaagag atcagtggga tctgatgagg actataaaga tcatgacggt
     2221 gattacaagg atcacgatat cgactataaa gatgacgacg acaagtaggc ggccgctcga
     2281 gtctagaggg cccgtttaaa cccgctgatc agcctcgact gtgccttcta gttgccagcc
     2341 atctgttgtt tgcccctccc ccgtgccttc cttgaccctg gaaggtgcca ctcccactgt
     2401 cctttcctaa taaaatgagg aaattgcatc gcattgtctg agtaggtgtc attctattct
     2461 ggggggtggg gtggggcagg acagcaaggg ggaggattgg gaagacaata gcaggcatgc
     2521 tggggatgcg gtgggctcta tggcttctga ggcggaaaga accagctggg gctctagggg
     2581 gtatccccac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag
     2641 cgtgaccgct acacttgcca gcgccctagc gcccgctcct ttcgctttct tcccttcctt
     2701 tctcgccacg ttcgccggct ttccccgtca agctctaaat cgggggctcc ctttagggtt
     2761 ccgatttagt gctttacggc acctcgaccc caaaaaactt gattagggtg atggttcacg
     2821 tagtgggcca tcgccctgat agacggtttt tcgccctttg acgttggagt ccacgttctt
     2881 taatagtgga ctcttgttcc aaactggaac aacactcaac cctatctcgg tctattcttt
     2941 tgatttataa gggattttgc cgatttcggc ctattggtta aaaaatgagc tgatttaaca
     3001 aaaatttaac gcgaattaat tctgtggaat gtgtgtcagt tagggtgtgg aaagtcccca
     3061 ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc aaccaggtgt
     3121 ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca
     3181 gcaaccatag tcccgcccct aactccgccc atcccgcccc taactccgcc cagttccgcc
     3241 cattctccgc cccatggctg actaattttt tttatttatg cagaggccga ggccgcctct
     3301 gcctctgagc tattccagaa gtagtgagga ggcttttttg gaggcctagg cttttgcaaa
     3361 aagctcccgg gagcttatat atccattttc ggatctgatc aagagacagg atgaggatcg
     3421 tttcgcatga ttgaacaaga tggattgcac gcaggttctc cggccgcttg ggtggagagg
     3481 ctattcggct atgactgggc acaacagaca atcggctgct ctgatgccgc cgtgttccgg
     3541 ctgtcagcgc aggggcgccc ggttcttttt gtcaagaccg acctgtccgg tgccctgaat
     3601 gaactgcagg acgaggcagc gcggctatcg tggctggcca cgacgggcgt tccttgcgca
     3661 gctgtgctcg acgttgtcac tgaagcggga agggactggc tgctattggg cgaagtgccg
     3721 gggcaggatc tcctgtcatc tcaccttgct cctgccgaga aagtatccat catggctgat
     3781 gcaatgcggc ggctgcatac gcttgatccg gctacctgcc cattcgacca ccaagcgaaa
     3841 catcgcatcg agcgagcacg tactcggatg gaagccggtc ttgtcgatca ggatgatctg
     3901 gacgaagagc atcaggggct cgcgccagcc gaactgttcg ccaggctcaa ggcgcgcatg
     3961 cccgacggcg aggatctcgt cgtgacccat ggcgatgcct gcttgccgaa tatcatggtg
     4021 gaaaatggcc gcttttctgg attcatcgac tgtggccggc tgggtgtggc ggaccgctat
     4081 caggacatag cgttggctac ccgtgatatt gctgaagagc ttggcggcga atgggctgac
     4141 cgcttcctcg tgctttacgg tatcgccgct cccgattcgc agcgcatcgc cttctatcgc
     4201 cttcttgacg agttcttctg agcgggactc tggggttcga aatgaccgac caagcgacgc
     4261 ccaacctgcc atcacgagat ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg
     4321 gaatcgtttt ccgggacgcc ggctggatga tcctccagcg cggggatctc atgctggagt
     4381 tcttcgccca ccccaacttg tttattgcag cttataatgg ttacaaataa agcaatagca
     4441 tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt ttgtccaaac
     4501 tcatcaatgt atcttatcat gtctgtatac cgtcgacctc tagctagagc ttggcgtaat
     4561 catggtcata gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacaacatac
     4621 gagccggaag cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa
     4681 ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat
     4741 gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc
     4801 tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg
     4861 cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag
     4921 gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc
     4981 gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag
     5041 gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga
     5101 ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc
     5161 atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg
     5221 tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt
     5281 ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca
     5341 gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca
     5401 ctagaagaac agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag
     5461 ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca
     5521 agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg
     5581 ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa
     5641 aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta
     5701 tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag
     5761 cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga
     5821 tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac
     5881 cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc
     5941 ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta
     6001 gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac
     6061 gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat
     6121 gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa
     6181 gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg
     6241 tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag
     6301 aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aata
//
