LOCUS       sequence-366420-        6028 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Low-level transient expression of delta-lumenal domain HsIRE1 with a
            C-terminal HaloTag..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6028)
  AUTHORS   Belyy V, Zuazo-Gaztelu I, Alamban A, Ashkenazi A, Walter P
  TITLE     Endoplasmic reticulum stress activates human IRE1alpha through 
            reversible assembly of inactive dimers into small oligomers.
  JOURNAL   Elife. 2022 Jun 22;11:e74342. doi: 10.7554/eLife.74342.
  PUBMED    35730415
REFERENCE   2  (bases 1 to 6028)
  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: "Elife."; 
            date: "2022-06-22"; volume: "11:e74342. doi"; pages: " 
            10.7554/eLife.74342"
COMMENT     Sequence Label: pPW3759 : CMVd3-HsIRE1deltaLD-HaloTag
FEATURES             Location/Qualifiers
     source          1..6028
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        155..199
                     /label=CMVd3 promoter
                     /note="truncated human cytomegalovirus (CMV) immediate 
                     early promoter that yields strongly reduced expression 
                     relative to the full-length promoter"
     CDS             1960..2850
                     /codon_start=1
                     /product="modified bacterial dehalogenase that forms 
                     covalent bonds with chloroalkane derivatives"
                     /label=HaloTag(R)
                     /translation="MAEIGTGFPFDPHYVEVLGERMHYVDVGPRDGTPVLFLHGNPTSS
                     YVWRNIIPHVAPTHRCIAPDLIGMGKSDKPDLGYFFDDHVRFMDAFIEALGLEEVVLVI
                     HDWGSALGFHWAKRNPERVKGIAFMEFIRPIPTWDEWPEFARETFQAFRTTDVGRKLII
                     DQNVFIEGTLPMGVVRPLTEVEMDHYREPFLNPVDREPLWRFPNELPIAGEPANIVALV
                     EEYMDWLHQSPVPKLLFWGTPGVLIPPAEAARLAKSLPNCKAVDIGPGLNLLQEDNPDL
                     IGSEIARWLSTLEISG"
     polyA_signal    2982..3103
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3019..3038)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3073..3092
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3110..3565)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3247..3268)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3459..3478
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3592..3696
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(3721..3741)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3765..3975
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     promoter        3787..3983
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      3834..3969
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4018..4812
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             4018..4812
                     /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
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4072..4091)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4682..4701
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5000..5019)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5044..5091
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5420..6008
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5909..5928
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tttacggttc ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc
       61 tgattctgtg gataaccgta ttaccgccat gcattagtta ttaatagtaa tcaattacgg
      121 ggtcattagt tcatagccca tatatggagt tccgatgggc ggtaggcgtg tacggtggga
      181 ggtctatata agcagagctg gtttagtgaa ccgtcagatc cgctagcgct accggtcgcc
      241 accatgccgg cccggcggct gctgctgctg ctgacgctgc tgctgcccgg cctcgggatt
      301 tttggaagta tgcttaagga catggctacc atcatcctga gcaccttcct gctgattggc
      361 tgggtggcct tcatcatcac ctatcccctg agcatgcatc agcagcagca gctccagcac
      421 cagcagttcc agaaggaact ggagaagatc cagctcctgc agcagcagca gcagcagctg
      481 cccttccacc cacctggaga cacggctcag gacggcgagc tcctggacac gtctggcccg
      541 tactcagaga gctcgggcac cagcagcccc agcacgtccc ccagggcctc caaccactcg
      601 ctctgctccg gcagctctgc ctccaaggct ggcagcagcc cctccctgga acaagacgat
      661 ggagatgagg aaaccagcgt ggtgatagtt gggaaaattt ccttctgtcc caaggatgtc
      721 ctgggccatg gagctgaggg cacaattgtg taccggggca tgtttgacaa ccgcgacgtg
      781 gccgtgaaga ggatcctccc cgagtgtttt agcttcgcag accgtgaggt ccagctgttg
      841 cgagaatcgg atgagcaccc gaacgtgatc cgctacttct gcacggagaa ggaccggcaa
      901 ttccagtaca ttgccatcga gctgtgtgca gccaccctgc aagagtatgt ggagcagaag
      961 gactttgcgc atctcggcct ggagcccatc accttgctgc agcagaccac ctcgggcctg
     1021 gcccacctcc actccctcaa catcgttcac agagacctaa agccacacaa catcctcata
     1081 tccatgccca atgcacacgg caagatcaag gccatgatct ccgactttgg cctctgcaag
     1141 aagctggcag tgggcagaca cagtttcagc cgccgatctg gggtgcctgg cacagaaggc
     1201 tggatcgctc cagagatgct gagcgaagac tgtaaggaga accctaccta cacggtggac
     1261 atcttttctg caggctgcgt cttttactac gtaatctctg agggcagcca cccttttggc
     1321 aagtccctgc agcggcaggc caacatcctc ctgggtgcct gcagccttga ctgcttgcac
     1381 ccagagaagc acgaagacgt cattgcacgt gaattgatag agaagatgat tgcgatggat
     1441 cctcagaaac gcccctcagc gaagcacgtg ctcaaacacc cgttcttctg gagcctagag
     1501 aagcagctcc agttcttcca ggacgtgagc gacagaatag aaaaggaatc cctggatggc
     1561 ccgatcgtga agcagttaga gagaggcggg agagccgtgg tgaagatgga ctggcgggag
     1621 aacatcactg tccccctcca gacagacctg cgtaaattca ggacctataa aggtggttct
     1681 gtcagagatc tcctccgagc catgagaaat aagaagcacc actaccggga gctgcctgca
     1741 gaggtgcggg agacgctggg gtccctcccc gacgacttcg tgtgctactt cacgtctcgc
     1801 ttcccccacc tcctcgcaca cacctaccgg gccatggagc tgtgcagcca cgagagactc
     1861 ttccagccct actacttcca cgagccccca gagccccagc ccccagtgac tccagacgcc
     1921 ctcggtggag gcggtagcgg aggaggtggc tctgtcgaca tggcagaaat cggtactggc
     1981 tttccattcg acccccatta tgtggaagtc ctgggcgagc gcatgcacta cgtcgatgtt
     2041 ggtccgcgcg atggcacccc tgtgctgttc ctgcacggta acccgacctc ctcctacgtg
     2101 tggcgcaaca tcatcccgca tgttgcaccg acccatcgct gcattgctcc agacctgatc
     2161 ggtatgggca aatccgacaa accagacctg ggttatttct tcgacgacca cgtccgcttc
     2221 atggatgcct tcatcgaagc cctgggtctg gaagaggtcg tcctggtcat tcacgactgg
     2281 ggctccgctc tgggtttcca ctgggccaag cgcaatccag agcgcgtcaa aggtattgca
     2341 tttatggagt tcatccgccc tatcccgacc tgggacgaat ggccagaatt tgcccgcgag
     2401 accttccagg ccttccgcac caccgacgtc ggccgcaagc tgatcatcga tcagaacgtt
     2461 tttatcgagg gtacgctgcc gatgggtgtc gtccgcccgc tgactgaagt cgagatggac
     2521 cattaccgcg agccgttcct gaatcctgtt gaccgcgagc cactgtggcg cttcccaaac
     2581 gagctgccaa tcgccggtga gccagcgaac atcgtcgcgc tggtcgaaga atacatggac
     2641 tggctgcacc agtcccctgt cccgaagctg ctgttctggg gcaccccagg cgttctgatc
     2701 ccaccggccg aagccgctcg cctggccaaa agcctgccta actgcaaggc tgtggacatc
     2761 ggcccgggtc tgaatctgct gcaagaagac aacccggacc tgatcggcag cgagatcgcg
     2821 cgctggctgt cgacgctcga gatttccggc taagatccac cggatctaga taactgatca
     2881 taatcagcca taccacattt gtagaggttt tacttgcttt aaaaaacctc ccacacctcc
     2941 ccctgaacct gaaacataaa atgaatgcaa ttgttgttgt taacttgttt attgcagctt
     3001 ataatggtta caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac
     3061 tgcattctag ttgtggtttg tccaaactca tcaatgtatc ttaacgcgta aattgtaagc
     3121 gttaatattt tgttaaaatt cgcgttaaat ttttgttaaa tcagctcatt ttttaaccaa
     3181 taggccgaaa tcggcaaaat cccttataaa tcaaaagaat agaccgagat agggttgagt
     3241 gttgttccag tttggaacaa gagtccacta ttaaagaacg tggactccaa cgtcaaaggg
     3301 cgaaaaaccg tctatcaggg cgatggccca ctacgtgaac catcacccta atcaagtttt
     3361 ttggggtcga ggtgccgtaa agcactaaat cggaacccta aagggagccc ccgatttaga
     3421 gcttgacggg gaaagccggc gaacgtggcg agaaaggaag ggaagaaagc gaaaggagcg
     3481 ggcgctaggg cgctggcaag tgtagcggtc acgctgcgcg taaccaccac acccgccgcg
     3541 cttaatgcgc cgctacaggg cgcgtcaggt ggcacttttc ggggaaatgt gcgcggaacc
     3601 cctatttgtt tatttttcta aatacattca aatatgtatc cgctcatgag acaataaccc
     3661 tgataaatgc ttcaataata ttgaaaaagg aagagtcctg aggcggaaag aaccagctgt
     3721 ggaatgtgtg tcagttaggg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc
     3781 aaagcatgca tctcaattag tcagcaacca tagtcccgcc cctaactccg cccatcccgc
     3841 ccctaactcc gcccagttcc gcccattctc cgccccatgg ctgactaatt ttttttattt
     3901 atgcagaggc cgaggccgcc tcggcctctg agctattcca gaagtagtga ggaggctttt
     3961 ttggaggcct aggcttttgc aaagatcgat caagagacag gatgaggatc gtttcgcatg
     4021 attgaacaag atggattgca cgcaggttct ccggccgctt gggtggagag gctattcggc
     4081 tatgactggg cacaacagac aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg
     4141 caggggcgcc cggttctttt tgtcaagacc gacctgtccg gtgccctgaa tgaactgcaa
     4201 gacgaggcag cgcggctatc gtggctggcc acgacgggcg ttccttgcgc agctgtgctc
     4261 gacgttgtca ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc ggggcaggat
     4321 ctcctgtcat ctcaccttgc tcctgccgag aaagtatcca tcatggctga tgcaatgcgg
     4381 cggctgcata cgcttgatcc ggctacctgc ccattcgacc accaagcgaa acatcgcatc
     4441 gagcgagcac gtactcggat ggaagccggt cttgtcgatc aggatgatct ggacgaagag
     4501 catcaggggc tcgcgccagc cgaactgttc gccaggctca aggcgagcat gcccgacggc
     4561 gaggatctcg tcgtgaccca tggcgatgcc tgcttgccga atatcatggt ggaaaatggc
     4621 cgcttttctg gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata
     4681 gcgttggcta cccgtgatat tgctgaagag cttggcggcg aatgggctga ccgcttcctc
     4741 gtgctttacg gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg ccttcttgac
     4801 gagttcttct gagcgggact ctggggttcg aaatgaccga ccaagcgacg cccaacctgc
     4861 catcacgaga tttcgattcc accgccgcct tctatgaaag gttgggcttc ggaatcgttt
     4921 tccgggacgc cggctggatg atcctccagc gcggggatct catgctggag ttcttcgccc
     4981 accctagggg gaggctaact gaaacacgga aggagacaat accggaagga acccgcgcta
     5041 tgacggcaat aaaaagacag aataaaacgc acggtgttgg gtcgtttgtt cataaacgcg
     5101 gggttcggtc ccagggctgg cactctgtcg ataccccacc gagaccccat tggggccaat
     5161 acgcccgcgt ttcttccttt tccccacccc accccccaag ttcgggtgaa ggcccagggc
     5221 tcgcagccaa cgtcggggcg gcaggccctg ccatagcctc aggttactca tatatacttt
     5281 agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc ctttttgata
     5341 atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag
     5401 aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa
     5461 caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt
     5521 ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgttctt ctagtgtagc
     5581 cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa
     5641 tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa
     5701 gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc
     5761 ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa
     5821 gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa
     5881 caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg
     5941 ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc
     6001 tatggaaaaa cgccagcaac gcggcctt
//
