LOCUS       sequence-352636-        6261 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  ER protein alpha-1-antitrypsin (WT) fused to an HA tag and the 
            eleventh beta-strand of GFP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6261)
  AUTHORS   Cortes Sanchon A, Santhosh Kumar H, Mantovani M, Osinnii I, Mateos 
            JM, Kaech A, Shcherbakov D, Akbergenov R, Bottger EC
  TITLE     ER-misfolded proteins become sequestered with mitochondria and 
            impair mitochondrial function.
  JOURNAL   Commun Biol. 2021 Dec 2;4(1):1350. doi: 10.1038/s42003-021-02873-w.
  PUBMED    34857875
REFERENCE   2  (bases 1 to 6261)
  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: "Commun 
            Biol."; date: "2021-12-2"; volume: "4(1):1350. doi"; pages: " 
            10.1038/s42003-021-02873-w"
COMMENT     Sequence Label: pAT-WT-HA-GFP11
FEATURES             Location/Qualifiers
     source          1..6261
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     23..42
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             141..1178
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     EVGRV"
     RBS             1157..1165
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
     polyA_signal    1202..1250
                     /note="synthetic polyadenylation signal"
     primer_bind     complement(1406..1423)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1577..2165)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1657..1676)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2365..3156,3157..3225))
                     /codon_start=1
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     polyA_signal    3330..3378
                     /note="synthetic polyadenylation signal"
     misc_feature    3392..3483
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     primer_bind     3432..3451
                     /label=RVprimer3
                     /note="pGL3 vector, forward primer"
     enhancer        3634..4013
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        4014..4217
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     4168..4188
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     4214..4238
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             5539..5565
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             5605..5652
                     /codon_start=1
                     /product="11th beta-strand of superfolder GFP"
                     /label=GFP11
                     /note="associates with GFP(1-10), consisting of the first 
                     10 beta-strands, to reconstitute fluorescent GFP (Cabantous
                     et al., 2005; Kamiyama et al., 2016)"
                     /translation="RDHMVLHEYVNAAGIT"
     polyA_signal    5699..5820
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5710..5729)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     5764..5783
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        join(6014..6261,1..110)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        6014..6205
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(6037..6057)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      join(6222..6261,1..96)
                     /label=SV40 ori
                     /note="SV40 origin of replication"
ORIGIN
        1 cccatggctg actaattttt tttatttatg cagaggccga ggccgcctct gcctctgagc
       61 tattccagaa gtagtgagga ggcttttttg gaggcctagg cttttgcaaa aagctcgatt
      121 cttctgacac tagcgccacc atgaagaagc ccgaactcac cgctaccagc gttgaaaaat
      181 ttctcatcga gaagttcgac agtgtgagcg acctgatgca gttgtcggag ggcgaagaga
      241 gccgagcctt cagcttcgat gtcggcggac gcggctatgt actgcgggtg aatagctgcg
      301 ctgatggctt ctacaaagac cgctacgtgt accgccactt cgccagcgct gcactaccca
      361 tccccgaagt gttggacatc ggcgagttca gcgagagcct gacatactgc atcagtagac
      421 gcgcccaagg cgttactctc caagacctcc ccgaaacaga gctgcctgct gtgttacagc
      481 ctgtcgccga agctatggat gctattgccg ccgccgacct cagtcaaacc agcggcttcg
      541 gcccattcgg gccccaaggc atcggccagt acacaacctg gcgggatttc atttgcgcca
      601 ttgctgatcc ccatgtctac cactggcaga ccgtgatgga cgacaccgtg tccgccagcg
      661 tagctcaagc cctggacgaa ctgatgctgt gggccgaaga ctgtcccgag gtgcgccacc
      721 tcgtccatgc cgacttcggc agcaacaacg tcctgaccga caacggccgc atcaccgccg
      781 taatcgactg gtccgaagct atgttcgggg acagtcagta cgaggtggcc aacatcttct
      841 tctggcggcc ctggctggct tgcatggagc agcagactcg ctacttcgag cgccggcatc
      901 ccgagctggc cggcagccct cgtctgcgag cctacatgct gcgcatcggc ctggatcagc
      961 tctaccagag cctcgtggac ggcaacttcg acgatgctgc ctgggctcaa ggccgctgcg
     1021 atgccatcgt ccgcagcggg gccggcaccg tcggtcgcac acaaatcgct cgccggagcg
     1081 cagccgtatg gaccgacggc tgcgtcgagg tgctggccga cagcggcaac cgccggccca
     1141 gtacacgacc gcgcgctaag gaggtaggtc gagtttaaac tctagaaccg gtcatggccg
     1201 caataaaata tctttatttt cattacatct gtgtgttggt tttttgtgtg ttcgaactag
     1261 atgctgtcga ccgatgccct tgagagcctt caacccagtc agctccttcc ggtgggcgcg
     1321 gggcatgact atcgtcgccg cacttatgac tgtcttcttt atcatgcaac tcgtaggaca
     1381 ggtgccggca gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc
     1441 tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg
     1501 ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg
     1561 ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac
     1621 gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg
     1681 gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct
     1741 ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg
     1801 tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct
     1861 gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac
     1921 tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt
     1981 tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc
     2041 tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca
     2101 ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat
     2161 ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac
     2221 gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt
     2281 aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct gacagcggcc
     2341 gcaaatgcta aaccactgca gtggttacca gtgcttgatc agtgaggcac cgatctcagc
     2401 gatctgccta tttcgttcgt ccatagtggc ctgactcccc gtcgtgtaga tcactacgat
     2461 tcgtgagggc ttaccatcag gccccagcgc agcaatgatg ccgcgagagc cgcgttcacc
     2521 ggcccccgat ttgtcagcaa tgaaccagcc agcagggagg gccgagcgaa gaagtggtcc
     2581 tgctactttg tccgcctcca tccagtctat gagctgctgt cgtgatgcta gagtaagaag
     2641 ttcgccagtg agtagtttcc gaagagttgt ggccattgct actggcatcg tggtatcacg
     2701 ctcgtcgttc ggtatggctt cgttcaactc tggttcccag cggtcaagcc gggtcacatg
     2761 atcacccata ttatgaagaa atgcagtcag ctccttaggg cctccgatcg ttgtcagaag
     2821 taagttggcc gcggtgttgt cgctcatggt aatggcagca ctacacaatt ctcttaccgt
     2881 catgccatcc gtaagatgct tttccgtgac cggcgagtac tcaaccaagt cgttttgtga
     2941 gtagtgtata cggcgaccaa gctgctcttg cccggcgtct atacgggaca acaccgcgcc
     3001 acatagcagt actttgaaag tgctcatcat cgggaatcgt tcttcggggc ggaaagactc
     3061 aaggatcttg ccgctattga gatccagttc gatatagccc actcttgcac ccagttgatc
     3121 ttcagcatct tttactttca ccagcgtttc ggggtgtgca aaaacaggca agcaaaatgc
     3181 cgcaaagaag ggaatgagtg cgacacgaaa atgttggatg ctcatactcg tcctttttca
     3241 atattattga agcatttatc agggttacta gtacgtctct caaggataag taagtaatat
     3301 taaggtacgg gaggtattgg acaggccgca ataaaatatc tttattttca ttacatctgt
     3361 gtgttggttt tttgtgtgaa tcgatagtac taacatacgc tctccatcaa aacaaaacga
     3421 aacaaaacaa actagcaaaa taggctgtcc ccagtgcaag tgcaggtgcc agaacatttc
     3481 tctggcctaa ctggcctcaa tattggccat tagccatatt attcattggt tatatagcat
     3541 aaatcaatat tggctattgg ccattgcata cgttgtatct atatcataat atgtacattt
     3601 atattggctc atgtccaata tgaccgccat gttggcattg attattgact agttattaat
     3661 agtaatcaat tacggggtca ttagttcata gcccatatat ggagttccgc gttacataac
     3721 ttacggtaaa tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa
     3781 tgacgtatgt tcccatagta acgccaatag ggactttcca ttgacgtcaa tgggtggagt
     3841 atttacggta aactgcccac ttggcagtac atcaagtgta tcatatgcca agtccgcccc
     3901 ctattgacgt caatgacggt aaatggcccg cctggcatta tgcccagtac atgaccttac
     3961 gggactttcc tacttggcag tacatctacg tattagtcat cgctattacc atggtgatgc
     4021 ggttttggca gtacaccaat gggcgtggat agcggtttga ctcacgggga tttccaagtc
     4081 tccaccccat tgacgtcaat gggagtttgt tttggcacca aaatcaacgg gactttccaa
     4141 aatgtcgtaa taaccccgcc ccgttgacgc aaatgggcgg taggcgtgta cggtgggagg
     4201 tctatataag cagagctcgt ttagtgaacc gtcagatcac tagaagcttc gatgccgtct
     4261 tctgtctcgt ggggcatcct cctgctggca ggcctgtgct gcctggtccc tgtctccctg
     4321 gctgaggatc cccagggaga tgctgcccag aagacagata catcccacca tgatcaggat
     4381 cacccaacct tcaacaagat cacccccaac ctggctgagt tcgccttcag cctataccgc
     4441 cagctggcac accagtccaa cagcaccaat atcttcttct ccccagtgag catcgctaca
     4501 gcctttgcaa tgctctccct ggggaccaag gctgacactc acgatgaaat cctggagggc
     4561 ctgaatttca acctcacgga gattccggag gctcagatcc atgaaggctt ccaggaactc
     4621 ctccgtaccc tcaaccagcc agacagccag ctccagctga ccaccggcaa tggcctgttc
     4681 ctcagcgagg gcctgaagct agtggataag tttttggagg atgttaaaaa gttgtaccac
     4741 tcagaagcct tcactgtcaa cttcggggac accgaagagg ccaagaaaca gatcaacgat
     4801 tacgtggaga agggtactca agggaaaatt gtggatttgg tcaaggagct tgacagagac
     4861 acagtttttg ctctggtgaa ttacatcttc tttaaaggca aatgggagag accctttgaa
     4921 gtcaaggaca ccgaggaaga ggacttccac gtggaccagg tgaccaccgt gaaggtgcct
     4981 atgatgaagc gtttaggcat gtttaacatc cagcactgta agaagctgtc cagctgggtg
     5041 ctgctgatga aatacctggg caatgccacc gccatcttct tcctgcctga tgaggggaaa
     5101 ctacagcacc tggaaaatga actcacccac gatatcatca ccaagttcct ggaaaatgaa
     5161 gacagaaggt ctgccagctt acatttaccc aaactgtcca ttactggaac ctatgatctg
     5221 aagagcgtcc tgggtcaact gggcatcact aaggtcttca gcaatggggc tgacctctcc
     5281 ggggtcacag aggaggcacc cctgaagctc tccaaggccg tgcataaggc tgtgctgacc
     5341 atcgacgaga aagggactga agctgctggg gccatgtttt tagaggccat acccatgtct
     5401 atcccccccg aggtcaagtt caacaaaccc tttgtcttct taatgattga acaaaatacc
     5461 aagtctcccc tcttcatggg aaaagtggtg aatcccaccc aaaaacccgg gggatcgggt
     5521 gacggtccgg tcgccaccta cccatacgat gttccagatt acgctggtcg acggatcggt
     5581 ggtggttctg gaggcggttc tatgagagat catatggttt tgcatgaata tgttaatgct
     5641 gctggtatta cttaataatt ctagagtcgg ggcggccggc cgcttcgagc agacatgata
     5701 agatacattg atgagtttgg acaaaccaca actagaatgc agtgaaaaaa atgctttatt
     5761 tgtgaaattt gtgatgctat tgctttattt gtaaccatta taagctgcaa taaacaagtt
     5821 aacaacaaca attgcattca ttttatgttt caggttcagg gggaggtgtg ggaggttttt
     5881 taaagcaagt aaaacctcta caaatgtggt aaaatcgata aggatccgtt tgcgtattgg
     5941 gcgctcttcc gctgatctgc gcagcaccat ggcctgaaat aacctctgaa agaggaactt
     6001 ggttagctac cttctgaggc ggaaagaacc agctgtggaa tgtgtgtcag ttagggtgtg
     6061 gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag
     6121 caaccaggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc
     6181 tcaattagtc agcaaccata gtcccgcccc taactccgcc catcccgccc ctaactccgc
     6241 ccagttccgc ccattctccg c
//
