LOCUS       sequence-451048-        5657 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  mEOS3.2-GS-SUMO-1(3-103)-Pex15(1-309)-PSP-GSS-FLAG-6xHis.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5657)
  AUTHORS   Castanzo DT, LaFrance B, Martin A
  TITLE     The AAA+ ATPase Msp1 is a processive protein translocase with robust
            unfoldase activity.
  JOURNAL   Proc Natl Acad Sci U S A. 2020 Jun 30;117(26):14970-14977. doi: 
            10.1073/pnas.1920109117. Epub 2020 Jun 15.
  PUBMED    32541053
REFERENCE   2  (bases 1 to 5657)
  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.1920109117"; journalName: "Proc Natl Acad Sci U S A"; 
            date: "2020-06-30- 30"; volume: "117"; issue: "26"; pages: 
            "14970-14977"
COMMENT     Sequence Label: pAM256
FEATURES             Location/Qualifiers
     source          1..5657
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(125..1207)
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     1169..1188
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(1208..1285)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     1331..1350
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1331..1349
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1350..1374
                     /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)."
     CDS             1418..2095
                     /codon_start=1
                     /product="improved monomeric variant of green-to-red 
                     photoswitchable fluorescent protein EosFP (Zhang et al., 
                     2012)"
                     /label=mEos3.2
                     /translation="MSAIKPDMKIKLRMEGNVNGHHFVIDGDGTGKPFEGKQSMDLEVK
                     EGGPLPFAFDILTTAFHYGNRVFAKYPDNIQDYFKQSFPKGYSWERSLTFEDGGICNAR
                     NDITMEGDTFYNKVRFYGTNFPANGPVMQKKTLKWEPSTEKMYVRDGVLTGDIEMALLL
                     EGNAHYRCDFRTTYKAKEKGVKLPGAHFVDHCIEILSHDKDYNKVKLYEHAVAHSGLPD
                     NARR"
     CDS             3341..3364
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             3374..3397
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             3398..3415
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(3442..3460)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      3456..3503
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(3724..4383)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     4298..4317
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(4384..4486)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     primer_bind     complement(4878..4895)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5012..5557)
                     /direction=LEFT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     complement(5092..5111)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 aattctcatg ttagtcatgc cccgcgccca ccggaaggag ctgactgggt tgaaggctct
       61 caagggcatc ggtcgagatc ccggtgccta atgagtgagc taacttacat taattgcgtt
      121 gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg
      181 ccaacgcgcg gggagaggcg gtttgcgtat tgggcgccag ggtggttttt cttttcacca
      241 gtgagacggg caacagctga ttgcccttca ccgcctggcc ctgagagagt tgcagcaagc
      301 ggtccacgct ggtttgcccc agcaggcgaa aatcctgttt gatggtggtt aacggcggga
      361 tataacatga gctgtcttcg gtatcgtcgt atcccactac cgagatgtcc gcaccaacgc
      421 gcagcccgga ctcggtaatg gcgcgcattg cgcccagcgc catctgatcg ttggcaacca
      481 gcatcgcagt gggaacgatg ccctcattca gcatttgcat ggtttgttga aaaccggaca
      541 tggcactcca gtcgccttcc cgttccgcta tcggctgaat ttgattgcga gtgagatatt
      601 tatgccagcc agccagacgc agacgcgccg agacagaact taatgggccc gctaacagcg
      661 cgatttgctg gtgacccaat gcgaccagat gctccacgcc cagtcgcgta ccgtcttcat
      721 gggagaaaat aatactgttg atgggtgtct ggtcagagac atcaagaaat aacgccggaa
      781 cattagtgca ggcagcttcc acagcaatgg catcctggtc atccagcgga tagttaatga
      841 tcagcccact gacgcgttgc gcgagaagat tgtgcaccgc cgctttacag gcttcgacgc
      901 cgcttcgttc taccatcgac accaccacgc tggcacccag ttgatcggcg cgagatttaa
      961 tcgccgcgac aatttgcgac ggcgcgtgca gggccagact ggaggtggca acgccaatca
     1021 gcaacgactg tttgcccgcc agttgttgtg ccacgcggtt gggaatgtaa ttcagctccg
     1081 ccatcgccgc ttccactttt tcccgcgttt tcgcagaaac gtggctggcc tggttcacca
     1141 cgcgggaaac ggtctgataa gagacaccgg catactctgc gacatcgtat aacgttactg
     1201 gtttcacatt caccaccctg aattgactct cttccgggcg ctatcatgcc ataccgcgaa
     1261 aggttttgcg ccattcgatg gtgtccggga tctcgacgct ctcccttatg cgactcctgc
     1321 attaggaaat taatacgact cactataggg gaattgtgag cggataacaa ttcccctgta
     1381 gaaataattt tgtttaactt taataaggag atataccatg agtgcgatta agccagacat
     1441 gaagatcaaa ctccgtatgg aaggcaacgt aaacgggcac cactttgtga tcgacggaga
     1501 tggtacaggc aagccttttg agggaaaaca gagtatggat cttgaagtca aagagggcgg
     1561 acctctgcct tttgcctttg atatcctgac cactgcattc cattacggca acagggtatt
     1621 cgccaaatat ccagacaaca tacaagacta ttttaagcag tcgtttccta aggggtattc
     1681 gtgggaacga agcttgactt tcgaagacgg gggcatttgc aacgccagaa acgacataac
     1741 aatggaaggg gacactttct ataataaagt tcgattttat ggtaccaact ttcccgccaa
     1801 tggtccagtt atgcagaaga agacgctgaa atgggagccc tccactgaga aaatgtatgt
     1861 gcgtgatgga gtgctgacgg gtgatattga gatggctttg ttgcttgaag gaaatgccca
     1921 ttaccgatgt gacttcagaa ctacttacaa agctaaggag aagggtgtca agttaccagg
     1981 cgcccacttt gtggaccact gcattgagat tttaagccat gacaaagatt acaacaaggt
     2041 taagctgtat gagcatgctg ttgctcattc tggattgcct gacaatgcca gacgagggtc
     2101 catgctcgag atgagcgatc aagaagcgaa accgagcacc gaagacctgg gcgacaagaa
     2161 ggaaggcgaa tacatcaagc tgaaagttat tggtcaagac agcagcgaaa tccacttcaa
     2221 ggtgaaaatg accacccacc tgaagaaact gaaagagagc tactgccagc gtcaaggtgt
     2281 tccgatgaac agcctgcgtt tcctgtttga aggccagcgt atcgcggaca accacacccc
     2341 gaaggagctg ggtatggagg aagaggatgt gattgaagtt tatcaggagc aaaccggtgg
     2401 ccacagcacc gtgatggctg caagtgagat aatgaacaat ctgcctatgc attccctcga
     2461 ttcttctcta agggatttac ttaacgatga cttattcatt gaaagtgatg aatcgaccaa
     2521 gtcggtaaat gatcagagga gtgaagtatt tcaagaatgt gtgaaccttt tcataaaaag
     2581 agatatcaag gattgcctag aaaaaatgtc tgaggttgga tttattgata ttaccgtttt
     2641 caagtccaac cctatgatat tagatttatt tgtcagcgcc tgtgatatta tgcccagctt
     2701 taccaaatta ggtttaacac tgcagagtga gattttgaat attttcacgt tagatacccc
     2761 tcaatgtatt gagacaagaa aaattattct cggtgacctc agtaaacttt tggtcatcaa
     2821 taaattcttt cgatgctgta tcaaggttat tcagtttaac ttaactgatc acaccgaaca
     2881 agaagaaaaa actctggagc ttgaatccat aatgagtgat ttcattttcg tatacataac
     2941 gaaaatgagg acaactatag atgtagttgg cttacaagaa ttaatcgaga tattcatctt
     3001 tcaggtgaag gtaaaactgc atcataaaaa accctcaccg aatatgtatt gggcgctttg
     3061 caaaacatta cctaagttat ctccaaccct aaagggcctt tacttatcta aagatgtttc
     3121 catagaggat gctattctga actcaataga taataaaata caaaaagata aggcgaagtc
     3181 taaaggtaaa caaagaggag tgaaacagaa aatacaccat tttcacgagc ctatgttaca
     3241 caatagtagt gaagaacagg ttaaggttga ggatgcgttt aatcaacgca cctctactga
     3301 tagtagactc cagagtactg gaacggcgcc ccgtaaaaaa ctggaagtcc tgttccaagg
     3361 tccgggcagc agcgattata aggatgacga tgacaagcat caccatcatc accactgatt
     3421 aattaaccta ggctgctgcc accgctgagc aataactagc ataacccctt ggggcctcta
     3481 aacgggtctt gaggggtttt ttgctgaaac ctcaggcatt tgagaagcac acggtcacac
     3541 tgcttccggt agtcaataaa ccggtaaacc agcaatagac ataagcggct atttaacgac
     3601 cctgccctga accgacgacc gggtcgaatt tgctttcgaa tttctgccat tcatccgctt
     3661 attatcactt attcaggcgt agcaaccagg cgtttaaggg caccaataac tgccttaaaa
     3721 aaattacgcc ccgccctgcc actcatcgca gtactgttgt aattcattaa gcattctgcc
     3781 gacatggaag ccatcacaaa cggcatgatg aacctgaatc gccagcggca tcagcacctt
     3841 gtcgccttgc gtataatatt tgcccatagt gaaaacgggg gcgaagaagt tgtccatatt
     3901 ggccacgttt aaatcaaaac tggtgaaact cacccaggga ttggctgaga cgaaaaacat
     3961 attctcaata aaccctttag ggaaataggc caggttttca ccgtaacacg ccacatcttg
     4021 cgaatatatg tgtagaaact gccggaaatc gtcgtggtat tcactccaga gcgatgaaaa
     4081 cgtttcagtt tgctcatgga aaacggtgta acaagggtga acactatccc atatcaccag
     4141 ctcaccgtct ttcattgcca tacggaactc cggatgagca ttcatcaggc gggcaagaat
     4201 gtgaataaag gccggataaa acttgtgctt atttttcttt acggtcttta aaaaggccgt
     4261 aatatccagc tgaacggtct ggttataggt acattgagca actgactgaa atgcctcaaa
     4321 atgttcttta cgatgccatt gggatatatc aacggtggta tatccagtga tttttttctc
     4381 cattttagct tccttagctc ctgaaaatct cgataactca aaaaatacgc ccggtagtga
     4441 tcttatttca ttatggtgaa agttggaacc tcttacgtgc cgatcaacgt ctcattttcg
     4501 ccaaaagttg gcccagggct tcccggtatc aacagggaca ccaggattta tttattctgc
     4561 gaagtgatct tccgtcacag gtatttattc ggcgcaaagt gcgtcgggtg atgctgccaa
     4621 cttactgatt tagtgtatga tggtgttttt gaggtgctcc agtggcttct gtttctatca
     4681 gctgtccctc ctgttcagct actgacgggg tggtgcgtaa cggcaaaagc accgccggac
     4741 atcagcgcta gcggagtgta tactggctta ctatgttggc actgatgagg gtgtcagtga
     4801 agtgcttcat gtggcaggag aaaaaaggct gcaccggtgc gtcagcagaa tatgtgatac
     4861 aggatatatt ccgcttcctc gctcactgac tcgctacgct cggtcgttcg actgcggcga
     4921 gcggaaatgg cttacgaacg gggcggagat ttcctggaag atgccaggaa gatacttaac
     4981 agggaagtga gagggccgcg gcaaagccgt ttttccatag gctccgcccc cctgacaagc
     5041 atcacgaaat ctgacgctca aatcagtggt ggcgaaaccc gacaggacta taaagatacc
     5101 aggcgtttcc ccctggcggc tccctcgtgc gctctcctgt tcctgccttt cggtttaccg
     5161 gtgtcattcc gctgttatgg ccgcgtttgt ctcattccac gcctgacact cagttccggg
     5221 taggcagttc gctccaagct ggactgtatg cacgaacccc ccgttcagtc cgaccgctgc
     5281 gccttatccg gtaactatcg tcttgagtcc aacccggaaa gacatgcaaa agcaccactg
     5341 gcagcagcca ctggtaattg atttagagga gttagtcttg aagtcatgcg ccggttaagg
     5401 ctaaactgaa aggacaagtt ttggtgactg cgctcctcca agccagttac ctcggttcaa
     5461 agagttggta gctcagagaa ccttcgaaaa accgccctgc aaggcggttt tttcgttttc
     5521 agagcaagag attacgcgca gaccaaaacg atctcaagaa gatcatctta ttaatcagat
     5581 aaaatatttc tagatttcag tgcaatttat ctcttcaaat gtagcacctg aagtcagccc
     5641 catacgatat aagttgt
//
