LOCUS       sequence-389414-        6238 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Codon-optimized bacterial expression plasmid. Expresses 
            GB1-His6-TEV-A11(89-196)..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6238)
  AUTHORS   Shihora A, Elias RD, Hammond JA, Ghirlando R, Deshmukh L
  TITLE     ALS Variants of Annexin A11's Proline-Rich Domain Impair Its 
            S100A6-Mediated Fibril Dissolution.
  JOURNAL   ACS Chem Neurosci. 2023 Jul 11. doi: 10.1021/acschemneuro.3c00169.
  PUBMED    37433222
REFERENCE   2  (bases 1 to 6238)
  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: "ACS Chem 
            Neurosci. 2023 Jul 11. doi: 10.1021/acschemneuro.3c00169."
COMMENT     Sequence Label: GB1-A11(89-196)
FEATURES             Location/Qualifiers
     source          1..6238
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     2..19
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    34..173
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(99..118)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     259..281
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(275..466)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(1778..2860)
                     /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     2822..2841
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(2861..2938)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     3144..3163
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     3247..3266
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        3247..3265
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    3266..3290
                     /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)."
     RBS             3305..3327
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             3503..3520
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             3521..3532
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     CDS             3572..3589
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             3590..3610
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     primer_bind     complement(3985..4003)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      3999..4046
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        complement(join(4221..4226,4227..4243,4244..4249))
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     primer_bind     complement(4276..4294)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4362..4466
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4467..4535,4536..5327)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(4685..4704)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5498..6086
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5987..6006
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cagcgagtca gtgagcgagg aagcggaaga gcgcctgatg cggtattttc tccttacgca
       61 tctgtgcggt atttcacacc gcaatggtgc actctcagta caatctgctc tgatgccgca
      121 tagttaagcc agtatacact ccgctatcgc tacgtgactg ggtcatggct gcgccccgac
      181 acccgccaac acccgctgac gcgccctgac gggcttgtct gctcccggca tccgcttaca
      241 gacaagctgt gaccgtctcc gggagctgca tgtgtcagag gttttcaccg tcatcaccga
      301 aacgcgcgag gcagctgcgg taaagctcat cagcgtggtc gtgaagcgat tcacagatgt
      361 ctgcctgttc atccgcgtcc agctcgttga gtttctccag aagcgttaat gtctggcttc
      421 tgataaagcg ggccatgtta agggcggttt tttcctgttt ggtcactgat gcctccgtgt
      481 aagggggatt tctgttcatg ggggtaatga taccgatgaa acgagagagg atgctcacga
      541 tacgggttac tgatgatgaa catgcccggt tactggaacg ttgtgagggt aaacaactgg
      601 cggtatggat gcggcgggac cagagaaaaa tcactcaggg tcaatgccag cgcttcgtta
      661 atacagatgt aggtgttcca cagggtagcc agcagcatcc tgcgatgcag atccggaaca
      721 taatggtgca gggcgctgac ttccgcgttt ccagacttta cgaaacacgg aaaccgaaga
      781 ccattcatgt tgttgctcag gtcgcagacg ttttgcagca gcagtcgctt cacgttcgct
      841 cgcgtatcgg tgattcattc tgctaaccag taaggcaacc ccgccagcct agccgggtcc
      901 tcaacgacag gagcacgatc atgcgcaccc gtggccagga cccaacgctg cccgagatgc
      961 gccgcgtgcg gctgctggag atggcggacg cgatggatat gttctgccaa gggttggttt
     1021 gcgcattcac agttctccgc aagaattgat tggctccaat tcttggagtg gtgaatccgt
     1081 tagcgaggtg ccgccggctt ccattcaggt cgaggtggcc cggctccatg caccgcgacg
     1141 caacgcgggg aggcagacaa ggtatagggc ggcgcctaca atccatgcca acccgttcca
     1201 tgtgctcgcc gaggcggcat aaatcgccgt gacgatcagc ggtccaatga tcgaagttag
     1261 gctggtaaga gccgcgagcg atccttgaag ctgtccctga tggtcgtcat ctacctgcct
     1321 ggacagcatg gcctgcaacg cgggcatccc gatgccgccg gaagcgagaa gaatcataat
     1381 ggggaaggcc atccagcctc gcgtcgcgaa cgccagcaag acgtagccca gcgcgtcggc
     1441 cgccatgccg gcgataatgg cctgcttctc gccgaaacgt ttggtggcgg gaccagtgac
     1501 gaaggcttga gcgagggcgt gcaagattcc gaataccgca agcgacaggc cgatcatcgt
     1561 cgcgctccag cgaaagcggt cctcgccgaa aatgacccag agcgctgccg gcacctgtcc
     1621 tacgagttgc atgataaaga agacagtcat aagtgcggcg acgatagtca tgccccgcgc
     1681 ccaccggaag gagctgactg ggttgaaggc tctcaagggc atcggtcgag atcccggtgc
     1741 ctaatgagtg agctaactta cattaattgc gttgcgctca ctgcccgctt tccagtcggg
     1801 aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg
     1861 tattgggcgc cagggtggtt tttcttttca ccagtgagac gggcaacagc tgattgccct
     1921 tcaccgcctg gccctgagag agttgcagca agcggtccac gctggtttgc cccagcaggc
     1981 gaaaatcctg tttgatggtg gttaacggcg ggatataaca tgagctgtct tcggtatcgt
     2041 cgtatcccac taccgagata tccgcaccaa cgcgcagccc ggactcggta atggcgcgca
     2101 ttgcgcccag cgccatctga tcgttggcaa ccagcatcgc agtgggaacg atgccctcat
     2161 tcagcatttg catggtttgt tgaaaaccgg acatggcact ccagtcgcct tcccgttccg
     2221 ctatcggctg aatttgattg cgagtgagat atttatgcca gccagccaga cgcagacgcg
     2281 ccgagacaga acttaatggg cccgctaaca gcgcgatttg ctggtgaccc aatgcgacca
     2341 gatgctccac gcccagtcgc gtaccgtctt catgggagaa aataatactg ttgatgggtg
     2401 tctggtcaga gacatcaaga aataacgccg gaacattagt gcaggcagct tccacagcaa
     2461 tggcatcctg gtcatccagc ggatagttaa tgatcagccc actgacgcgt tgcgcgagaa
     2521 gattgtgcac cgccgcttta caggcttcga cgccgcttcg ttctaccatc gacaccacca
     2581 cgctggcacc cagttgatcg gcgcgagatt taatcgccgc gacaatttgc gacggcgcgt
     2641 gcagggccag actggaggtg gcaacgccaa tcagcaacga ctgtttgccc gccagttgtt
     2701 gtgccacgcg gttgggaatg taattcagct ccgccatcgc cgcttccact ttttcccgcg
     2761 ttttcgcaga aacgtggctg gcctggttca ccacgcggga aacggtctga taagagacac
     2821 cggcatactc tgcgacatcg tataacgtta ctggtttcac attcaccacc ctgaattgac
     2881 tctcttccgg gcgctatcat gccataccgc gaaaggtttt gcgccattcg atggtgtccg
     2941 ggatctcgac gctctccctt atgcgactcc tgcattagga agcagcccag tagtaggttg
     3001 aggccgttga gcaccgccgc cgcaaggaat ggtgcatgca aggagatggc gcccaacagt
     3061 cccccggcca cggggcctgc caccataccc acgccgaaac aagcgctcat gagcccgaag
     3121 tggcgagccc gatcttcccc atcggtgatg tcggcgatat aggcgccagc aaccgcacct
     3181 gtggcgccgg tgatgccggc cacgatgcgt ccggcgtaga ggatcgagat ctcgatcccg
     3241 cgaaattaat acgactcact ataggggaat tgtgagcgga taacaattcc cctctagaaa
     3301 taattttgtt taactttaag aaggagatat acatatgcag tataaactgg cgctgaacgg
     3361 caaaaccctg aaaggcgaaa ccaccaccga agcggtggat gcggcgaccg cggaaaaagt
     3421 gtttaaacag tatgcgaacg ataacggcgt ggatggcgaa tggacctatg atgatgcgac
     3481 caaaaccttt accgtgaccg aactggtgcc gcgcggcagc attgaaggcc gcattcgcga
     3541 aaacctgaaa gatagcggcc tgtttctgga acatcatcat catcatcatg aaaacctgta
     3601 tttccaaggc tttgggcagc cgccgagcgc gcaacagcct gttccgccgt acgggatgta
     3661 cccgccgccg ggcggcaacc caccgagccg catgccgtca tatccaccgt accctggggc
     3721 accggtccct ggtcaaccga tgccaccgcc tgggcaacaa ccgccgggtg cttacccggg
     3781 tcagccgcct gttacgtatc ctggtcagcc gccggtgccg ctcccgggtc aacagcaacc
     3841 ggtgccatca tacccgggct acccgggcag cggcaccgta actccggcgg tgccgccgac
     3901 gcagttcggc agcagaggca ccatcaccga ctaaggatcc ggctgctaac aaagcccgaa
     3961 aggaagctga gttggctgct gccaccgctg agcaataact agcataaccc cttggggcct
     4021 ctaaacgggt cttgaggggt tttttgctga aaggaggaac tatatccgga tatcccgcaa
     4081 gaggcccggc agtaccggca taaccaagcc tatgcctaca gcatccaggg tgacggtgcc
     4141 gaggatgacg atgagcgcat tgttagattt catacacggt gcctgactgc gttagcaatt
     4201 taactgtgat aaactaccgc attaaagctt atcgatgata agctgtcaaa catgagaatt
     4261 cttgaagacg aaagggcctc gtgatacgcc tatttttata ggttaatgtc atgataataa
     4321 tggtttctta gacgtcaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt
     4381 tatttttcta aatacattca aatatgtatc cgctcatgag acaataaccc tgataaatgc
     4441 ttcaataata ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc gcccttattc
     4501 ccttttttgc ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa
     4561 aagatgctga agatcagttg ggtgcacgag tgggttacat cgaactggat ctcaacagcg
     4621 gtaagatcct tgagagtttt cgccccgaag aacgttttcc aatgatgagc acttttaaag
     4681 ttctgctatg tggcgcggta ttatcccgtg ttgacgccgg gcaagagcaa ctcggtcgcc
     4741 gcatacacta ttctcagaat gacttggttg agtactcacc agtcacagaa aagcatctta
     4801 cggatggcat gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg
     4861 cggccaactt acttctgaca acgatcggag gaccgaagga gctaaccgct tttttgcaca
     4921 acatggggga tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac
     4981 caaacgacga gcgtgacacc acgatgcctg cagcaatggc aacaacgttg cgcaaactat
     5041 taactggcga actacttact ctagcttccc ggcaacaatt aatagactgg atggaggcgg
     5101 ataaagttgc aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata
     5161 aatctggagc cggtgagcgt gggtctcgcg gtatcattgc agcactgggg ccagatggta
     5221 agccctcccg tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa
     5281 atagacagat cgctgagata ggtgcctcac tgattaagca ttggtaactg tcagaccaag
     5341 tttactcata tatactttag attgatttaa aacttcattt ttaatttaaa aggatctagg
     5401 tgaagatcct ttttgataat ctcatgacca aaatccctta acgtgagttt tcgttccact
     5461 gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg
     5521 taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc
     5581 aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata
     5641 ctgtccttct agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta
     5701 catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc
     5761 ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg
     5821 ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac
     5881 agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg
     5941 taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt
     6001 atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct
     6061 cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg
     6121 ccttttgctg gccttttgct cacatgttct ttcctgcgtt atcccctgat tctgtggata
     6181 accgtattac cgcctttgag tgagctgata ccgctcgccg cagccgaacg accgagcg
//
