LOCUS       sequence-366419-        5755 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Low-level transient expression of HsIRE1 lacking the linker, kinase,
            and RNase domains, with a C-terminal HaloTag..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5755)
  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 5755)
  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: pPW3763 : CMVd3-HsIRE1dLKR-HaloTag
FEATURES             Location/Qualifiers
     source          1..5755
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        83..127
                     /label=CMVd3 promoter
                     /note="truncated human cytomegalovirus (CMV) immediate 
                     early promoter that yields strongly reduced expression 
                     relative to the full-length promoter"
     CDS             1615..2505
                     /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    2637..2758
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2674..2693)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2728..2747
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2765..3220)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2902..2923)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3114..3133
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3247..3351
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(3376..3396)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3420..3630
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     promoter        3442..3638
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      3489..3624
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3673..4467
                     /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             3673..4467
                     /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(3727..3746)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4337..4356
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4655..4674)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4699..4746
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5075..5663
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5564..5583
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 taaccgtatt accgccatgc attagttatt aatagtaatc aattacgggg tcattagttc
       61 atagcccata tatggagttc cgatgggcgg taggcgtgta cggtgggagg tctatataag
      121 cagagctggt ttagtgaacc gtcagatccg ctagcgctac cggtcgccac catgccggcc
      181 cggcggctgc tgctgctgct gacgctgctg ctgcccggcc tcgggatttt tggaagtacc
      241 agcacagtga cgcttcctga aaccttgttg tttgtgtcaa cgctggatgg aagtttgcat
      301 gctgtcagca agaggacagg ctcaatcaaa tggactttaa aagaagatcc agtcctgcag
      361 gtcccaacac atgtggaaga gcctgccttt ctcccagatc ctaatgatgg cagcctgtat
      421 acgcttggaa gcaagaataa tgaaggcctg acgaaacttc cttttaccat cccagaattg
      481 gtgcaggcat ccccatgccg aagttcagat ggaatcctct acatgggtaa aaagcaggac
      541 atctggtatg ttattgacct cctgaccgga gagaagcagc agactttgtc atcggccttt
      601 gcagatagtc tctgcccatc aacctctctt ctgtatcttg ggcgaacaga atacaccatc
      661 accatgtacg acaccaaaac ccgagagctc cggtggaatg ccacctactt tgactatgcg
      721 gcctcactgc ctgaggacga cgtggactac aagatgtccc actttgtgtc caatggtgat
      781 gggctggtgg tgactgtgga cagtgaatct ggggacgtcc tgtggatcca aaactacgcc
      841 tcccctgtgg tggcctttta tgtctggcag cgggagggtc tgaggaaggt gatgcacatc
      901 aatgtcgctg tggagaccct gcgctatctg accttcatgt ctggggaggt ggggcgcatc
      961 acaaagtgga agtacccgtt ccccaaggag acagaggcca agagcaagct gacgcccact
     1021 ctgtatgttg ggaaatactc taccagcctc tatgcctctc cctcaatggt acacgagggg
     1081 gttgctgtcg tgccccgcgg cagcacactt cctttgctgg aagggcccca gactgatggc
     1141 gtcaccatcg gggacaaggg ggagtgtgtg atcacgccca gcacggacgt caagtttgat
     1201 cccggactca aaagcaagaa caagctcaac tacttgagga attactggct tctgatagga
     1261 caccatgaaa ccccactgtc tgcgtctacc aagatgctgg agagatttcc caacaatcta
     1321 cccaaacatc gggaaaatgt gattcctgct gattcagaga aaaagagctt tgaggaagtt
     1381 atcaacctgg ttgaccagac ttcagaaaac gcacctacca ccgtgtctcg ggatgtggag
     1441 gagaagcccg cccatgcccc tgcccggccc gaggcccccg tggactccat gcttaaggac
     1501 atggctacca tcatcctgag caccttcctg ctgattggct gggtggcctt catcatcacc
     1561 tatcccctga gcatgcatgg tggaggcggt agcggaggag gtggctctgt cgacatggca
     1621 gaaatcggta ctggctttcc attcgacccc cattatgtgg aagtcctggg cgagcgcatg
     1681 cactacgtcg atgttggtcc gcgcgatggc acccctgtgc tgttcctgca cggtaacccg
     1741 acctcctcct acgtgtggcg caacatcatc ccgcatgttg caccgaccca tcgctgcatt
     1801 gctccagacc tgatcggtat gggcaaatcc gacaaaccag acctgggtta tttcttcgac
     1861 gaccacgtcc gcttcatgga tgccttcatc gaagccctgg gtctggaaga ggtcgtcctg
     1921 gtcattcacg actggggctc cgctctgggt ttccactggg ccaagcgcaa tccagagcgc
     1981 gtcaaaggta ttgcatttat ggagttcatc cgccctatcc cgacctggga cgaatggcca
     2041 gaatttgccc gcgagacctt ccaggccttc cgcaccaccg acgtcggccg caagctgatc
     2101 atcgatcaga acgtttttat cgagggtacg ctgccgatgg gtgtcgtccg cccgctgact
     2161 gaagtcgaga tggaccatta ccgcgagccg ttcctgaatc ctgttgaccg cgagccactg
     2221 tggcgcttcc caaacgagct gccaatcgcc ggtgagccag cgaacatcgt cgcgctggtc
     2281 gaagaataca tggactggct gcaccagtcc cctgtcccga agctgctgtt ctggggcacc
     2341 ccaggcgttc tgatcccacc ggccgaagcc gctcgcctgg ccaaaagcct gcctaactgc
     2401 aaggctgtgg acatcggccc gggtctgaat ctgctgcaag aagacaaccc ggacctgatc
     2461 ggcagcgaga tcgcgcgctg gctgtcgacg ctcgagattt ccggctaaga tccaccggat
     2521 ctagataact gatcataatc agccatacca catttgtaga ggttttactt gctttaaaaa
     2581 acctcccaca cctccccctg aacctgaaac ataaaatgaa tgcaattgtt gttgttaact
     2641 tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata
     2701 aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttaac
     2761 gcgtaaattg taagcgttaa tattttgtta aaattcgcgt taaatttttg ttaaatcagc
     2821 tcatttttta accaataggc cgaaatcggc aaaatccctt ataaatcaaa agaatagacc
     2881 gagatagggt tgagtgttgt tccagtttgg aacaagagtc cactattaaa gaacgtggac
     2941 tccaacgtca aagggcgaaa aaccgtctat cagggcgatg gcccactacg tgaaccatca
     3001 ccctaatcaa gttttttggg gtcgaggtgc cgtaaagcac taaatcggaa ccctaaaggg
     3061 agcccccgat ttagagcttg acggggaaag ccggcgaacg tggcgagaaa ggaagggaag
     3121 aaagcgaaag gagcgggcgc tagggcgctg gcaagtgtag cggtcacgct gcgcgtaacc
     3181 accacacccg ccgcgcttaa tgcgccgcta cagggcgcgt caggtggcac ttttcgggga
     3241 aatgtgcgcg gaacccctat ttgtttattt ttctaaatac attcaaatat gtatccgctc
     3301 atgagacaat aaccctgata aatgcttcaa taatattgaa aaaggaagag tcctgaggcg
     3361 gaaagaacca gctgtggaat gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag
     3421 caggcagaag tatgcaaagc atgcatctca attagtcagc aaccatagtc ccgcccctaa
     3481 ctccgcccat cccgccccta actccgccca gttccgccca ttctccgccc catggctgac
     3541 taattttttt tatttatgca gaggccgagg ccgcctcggc ctctgagcta ttccagaagt
     3601 agtgaggagg cttttttgga ggcctaggct tttgcaaaga tcgatcaaga gacaggatga
     3661 ggatcgtttc gcatgattga acaagatgga ttgcacgcag gttctccggc cgcttgggtg
     3721 gagaggctat tcggctatga ctgggcacaa cagacaatcg gctgctctga tgccgccgtg
     3781 ttccggctgt cagcgcaggg gcgcccggtt ctttttgtca agaccgacct gtccggtgcc
     3841 ctgaatgaac tgcaagacga ggcagcgcgg ctatcgtggc tggccacgac gggcgttcct
     3901 tgcgcagctg tgctcgacgt tgtcactgaa gcgggaaggg actggctgct attgggcgaa
     3961 gtgccggggc aggatctcct gtcatctcac cttgctcctg ccgagaaagt atccatcatg
     4021 gctgatgcaa tgcggcggct gcatacgctt gatccggcta cctgcccatt cgaccaccaa
     4081 gcgaaacatc gcatcgagcg agcacgtact cggatggaag ccggtcttgt cgatcaggat
     4141 gatctggacg aagagcatca ggggctcgcg ccagccgaac tgttcgccag gctcaaggcg
     4201 agcatgcccg acggcgagga tctcgtcgtg acccatggcg atgcctgctt gccgaatatc
     4261 atggtggaaa atggccgctt ttctggattc atcgactgtg gccggctggg tgtggcggac
     4321 cgctatcagg acatagcgtt ggctacccgt gatattgctg aagagcttgg cggcgaatgg
     4381 gctgaccgct tcctcgtgct ttacggtatc gccgctcccg attcgcagcg catcgccttc
     4441 tatcgccttc ttgacgagtt cttctgagcg ggactctggg gttcgaaatg accgaccaag
     4501 cgacgcccaa cctgccatca cgagatttcg attccaccgc cgccttctat gaaaggttgg
     4561 gcttcggaat cgttttccgg gacgccggct ggatgatcct ccagcgcggg gatctcatgc
     4621 tggagttctt cgcccaccct agggggaggc taactgaaac acggaaggag acaataccgg
     4681 aaggaacccg cgctatgacg gcaataaaaa gacagaataa aacgcacggt gttgggtcgt
     4741 ttgttcataa acgcggggtt cggtcccagg gctggcactc tgtcgatacc ccaccgagac
     4801 cccattgggg ccaatacgcc cgcgtttctt ccttttcccc accccacccc ccaagttcgg
     4861 gtgaaggccc agggctcgca gccaacgtcg gggcggcagg ccctgccata gcctcaggtt
     4921 actcatatat actttagatt gatttaaaac ttcattttta atttaaaagg atctaggtga
     4981 agatcctttt tgataatctc atgaccaaaa tcccttaacg tgagttttcg ttccactgag
     5041 cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa
     5101 tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag
     5161 agctaccaac tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg
     5221 ttcttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat
     5281 acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta
     5341 ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg
     5401 gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc
     5461 gtgagctatg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa
     5521 gcggcagggt cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc
     5581 tttatagtcc tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt
     5641 caggggggcg gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct
     5701 tttgctggcc ttttgctcac atgttctttc ctgcgttatc ccctgattct gtgga
//
