LOCUS       sequence-331345-        7751 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AvTnsC expression construct for protein purification..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7751)
  AUTHORS   Saito M, Ladha A, Strecker J, Faure G, Neumann E, Altae-Tran H, 
            Macrae RK, Zhang F
  TITLE     Dual modes of CRISPR-associated transposon homing.
  JOURNAL   Cell. 2021 Apr 29;184(9):2441-2453.e18. doi: 
            10.1016/j.cell.2021.03.006. Epub 2021 Mar 25.
  PUBMED    33770501
REFERENCE   2  (bases 1 to 7751)
  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: 
            "10.1016/j.cell.2021.03.006"; journalName: "Cell"; date: 
            "2021-04-29- 29"; volume: "184"; issue: "9"; pages: "2441-2453.e18"
COMMENT     Sequence Label: pMS16: pTwinStrep-SUMO-AvTnsC
FEATURES             Location/Qualifiers
     source          1..7751
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     932..951
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(971..1048)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     1254..1273
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     1357..1376
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1357..1375
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1376..1400
                     /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             1415..1437
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             1456..1473
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1483..1500
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             join(1513..1536,1537..1572,1573..1596)
                     /codon_start=1
                     /product="two Strep-Tag(R)II moieties connected by a linker
                     for enhanced binding to Strep-Tactin(R), an engineered form
                     of streptavidin (Schmidt et al., 2013)"
                     /label=Twin-Strep-tag
                     /translation="WSHPQFEKGGGSGGGSGGSAWSHPQFEK"
     CDS             1597..1890
                     /codon_start=1
                     /gene="S. cerevisiae SMT3 (truncated)"
                     /product="cleavable ubiquitin-like protein tag"
                     /label=SUMO
                     /translation="MSDSEVNQEAKPEVKPEVKPETHINLKVSDGSSEIFFKIKKTTPL
                     RRLMEAFAKRQGKEMDSLRFLYDGIRIQADQTPEDLDMEDNDIIEAHREQIGG"
     primer_bind     complement(3608..3626)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      3622..3669
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        complement(join(3844..3849,3850..3866,3867..3872))
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     primer_bind     complement(3899..3917)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3985..4089
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4090..4158,4159..4950)
                     /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(4308..4327)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5121..5709
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5610..5629
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5863..5880
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    5895..6034
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(5960..5979)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6120..6142
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(6136..6327)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(join(7639..7751,1..970))
                     /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"
ORIGIN
        1 ccagtgagac gggcaacagc tgattgccct tcaccgcctg gccctgagag agttgcagca
       61 agcggtccac gctggtttgc cccagcaggc gaaaatcctg tttgatggtg gttaacggcg
      121 ggatataaca tgagctgtct tcggtatcgt cgtatcccac taccgagata tccgcaccaa
      181 cgcgcagccc ggactcggta atggcgcgca ttgcgcccag cgccatctga tcgttggcaa
      241 ccagcatcgc agtgggaacg atgccctcat tcagcatttg catggtttgt tgaaaaccgg
      301 acatggcact ccagtcgcct tcccgttccg ctatcggctg aatttgattg cgagtgagat
      361 atttatgcca gccagccaga cgcagacgcg ccgagacaga acttaatggg cccgctaaca
      421 gcgcgatttg ctggtgaccc aatgcgacca gatgctccac gcccagtcgc gtaccgtctt
      481 catgggagaa aataatactg ttgatgggtg tctggtcaga gacatcaaga aataacgccg
      541 gaacattagt gcaggcagct tccacagcaa tggcatcctg gtcatccagc ggatagttaa
      601 tgatcagccc actgacgcgt tgcgcgagaa gattgtgcac cgccgcttta caggcttcga
      661 cgccgcttcg ttctaccatc gacaccacca cgctggcacc cagttgatcg gcgcgagatt
      721 taatcgccgc gacaatttgc gacggcgcgt gcagggccag actggaggtg gcaacgccaa
      781 tcagcaacga ctgtttgccc gccagttgtt gtgccacgcg gttgggaatg taattcagct
      841 ccgccatcgc cgcttccact ttttcccgcg ttttcgcaga aacgtggctg gcctggttca
      901 ccacgcggga aacggtctga taagagacac cggcatactc tgcgacatcg tataacgtta
      961 ctggtttcac attcaccacc ctgaattgac tctcttccgg gcgctatcat gccataccgc
     1021 gaaaggtttt gcgccattcg atggtgtccg ggatctcgac gctctccctt atgcgactcc
     1081 tgcattagga agcagcccag tagtaggttg aggccgttga gcaccgccgc cgcaaggaat
     1141 ggtgcatgca aggagatggc gcccaacagt cccccggcca cggggcctgc caccataccc
     1201 acgccgaaac aagcgctcat gagcccgaag tggcgagccc gatcttcccc atcggtgatg
     1261 tcggcgatat aggcgccagc aaccgcacct gtggcgccgg tgatgccggc cacgatgcgt
     1321 ccggcgtaga ggatcgagat ctcgatcccg cgaaattaat acgactcact ataggggaat
     1381 tgtgagcgga taacaattcc cctctagaaa taattttgtt taactttaag aaggagatat
     1441 accatgggca gcagccatca tcatcatcat cacagcagcg gcctggtgcc gcgcggcagc
     1501 catatggcta gctggagcca tccgcagttt gaaaaaggtg gtggtagcgg tggtggttca
     1561 ggtggtagtg catggtcaca ccctcagttt gagaaaatgt cggactcaga agtcaatcaa
     1621 gaagctaagc cagaggtcaa gccagaagtc aagcctgaga ctcacatcaa tttaaaggtg
     1681 tccgatggat cttcagagat cttcttcaag atcaaaaaga ccactccttt aagaaggctg
     1741 atggaagcgt tcgctaaaag acagggtaag gaaatggact ccttaagatt cttgtacgac
     1801 ggtattagaa tccaagctga tcagacccct gaagatttgg acatggagga taacgatatt
     1861 attgaggctc acagagaaca gattggtgga tccgacagaa ttgagagtga actggttaaa
     1921 attcctaatg gagaactagc agttattgcc agatatatag atcaaaaact gccagaatac
     1981 aacactaatc cattaattca ggcattacct cctatcctat cagcagcaga atttatagat
     2041 aaagtagcca gagaaccaaa ttttgatgaa caagagagag agctagaacc acattataga
     2101 tttcattgta ttgagaggtt atctagatat tttgaccctc aaaataaaac tcttgaatta
     2161 caaaaagtta tttgtacgct tatcatgaca ggctatctgt caaggaacct tttaaaacct
     2221 gaatatgcaa atcgctctag gcaaatttat aatgcaatag gtaatggagg aggtaaaaat
     2281 ttagaaaact atgttaatct tcctacatct gcgtccggct taactctaat tggcccatca
     2341 ggtattggaa agactacaaa tttcaaaaat attcttgagc tttatcccca aataattttt
     2401 catcctgatt atagtgttta ccaaatagtt tggctgcggg tagattgccc taaaacttta
     2461 aaggggttat gtatagatac ttttgcaggt atcgataaat tgttaggaac taatttatta
     2521 aagaaattta acccagcacg cacatctgaa gattatctat tggctcaact tgcacaaaca
     2581 gcacacactc atcatctagg agttttggtg attgatgaaa tacaaaattt agcaactgca
     2641 aaaagaaata gagacgaaat attaaatttt ttagtaaagc tcgataatac tattggaatt
     2701 ccagttattc gtgtaggtac gaatgaagcc attagttttc ttcaaggtaa tcttagaaat
     2761 gctagaagaa atactggaga aggaagtaga atgtgggata gaatggtaaa taatgatgaa
     2821 tggtgttact ttttagaagg aatatgggaa taccaatgga caaaaacacc tgttcatttc
     2881 tcggatgaaa taaaagaagt tttttatgat gaaagccagg gaattattga tattgctata
     2941 aaactttaca aaatggttca atggagagcg atttcaatag gtgaccaaga aattattacg
     3001 gttgatctga ttcgtcaagt agctaaagaa ggattgtatt tagtaaagcc aatgctagat
     3061 gcaattaggt taggggacag agagtggatg attaaatata aggatattgc tcctctcgat
     3121 attacggaat atcacaatag atgtgtatct gagattgatg ctaaagattt gacagagatt
     3181 aagagattag ctaaaaagca gcagaatcag cagaaagtat ctccgaagct caattatgta
     3241 attagtgagc tgttgaaatt ggaagtagaa tctggtaaag caaaggaatg tgcagaacaa
     3301 attgtttata gtcatgaaga tccagacatt ccaattccat ctttagttaa taaggcatat
     3361 gctcttgctt tgcaaggatg tcaagtaaat gagagtcttg ctaaaacaag taccaggaaa
     3421 gctaaattaa agacaaagta ccaagagaat gatattcggc gaattgtcga gaatgccaag
     3481 aaagacaaaa aatctgcata cgaggatttg aaattagcag gtataatcaa aaatccagtt
     3541 gaagatttta tcaatattta ggcggccgca ctcgaggccc gaaaggaagc tgagttggct
     3601 gctgccaccg ctgagcaata actagcataa ccccttgggg cctctaaacg ggtcttgagg
     3661 ggttttttgc tgaaaggagg aactatatcc ggatatcccg caagaggccc ggcagtaccg
     3721 gcataaccaa gcctatgcct acagcatcca gggtgacggt gccgaggatg acgatgagcg
     3781 cattgttaga tttcatacac ggtgcctgac tgcgttagca atttaactgt gataaactac
     3841 cgcattaaag cttatcgatg ataagctgtc aaacatgaga attcttgaag acgaaagggc
     3901 ctcgtgatac gcctattttt ataggttaat gtcatgataa taatggtttc ttagacgtca
     3961 ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat
     4021 tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa
     4081 aggaagagta tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt
     4141 tgccttcctg tttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag
     4201 ttgggtgcac gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt
     4261 tttcgccccg aagaacgttt tccaatgatg agcactttta aagttctgct atgtggcgcg
     4321 gtattatccc gtgttgacgc cgggcaagag caactcggtc gccgcataca ctattctcag
     4381 aatgacttgg ttgagtactc accagtcaca gaaaagcatc ttacggatgg catgacagta
     4441 agagaattat gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg
     4501 acaacgatcg gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta
     4561 actcgccttg atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac
     4621 accacgatgc ctgcagcaat ggcaacaacg ttgcgcaaac tattaactgg cgaactactt
     4681 actctagctt cccggcaaca attaatagac tggatggagg cggataaagt tgcaggacca
     4741 cttctgcgct cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag
     4801 cgtgggtctc gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta
     4861 gttatctaca cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag
     4921 ataggtgcct cactgattaa gcattggtaa ctgtcagacc aagtttactc atatatactt
     4981 tagattgatt taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat
     5041 aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta
     5101 gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa
     5161 acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt
     5221 tttccgaagg taactggctt cagcagagcg cagataccaa atactgtcct tctagtgtag
     5281 ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta
     5341 atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca
     5401 agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag
     5461 cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa
     5521 agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga
     5581 acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc
     5641 gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc
     5701 ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt
     5761 gctcacatgt tctttcctgc gttatcccct gattctgtgg ataaccgtat taccgccttt
     5821 gagtgagctg ataccgctcg ccgcagccga acgaccgagc gcagcgagtc agtgagcgag
     5881 gaagcggaag agcgcctgat gcggtatttt ctccttacgc atctgtgcgg tatttcacac
     5941 cgcaatggtg cactctcagt acaatctgct ctgatgccgc atagttaagc cagtatacac
     6001 tccgctatcg ctacgtgact gggtcatggc tgcgccccga cacccgccaa cacccgctga
     6061 cgcgccctga cgggcttgtc tgctcccggc atccgcttac agacaagctg tgaccgtctc
     6121 cgggagctgc atgtgtcaga ggttttcacc gtcatcaccg aaacgcgcga ggcagctgcg
     6181 gtaaagctca tcagcgtggt cgtgaagcga ttcacagatg tctgcctgtt catccgcgtc
     6241 cagctcgttg agtttctcca gaagcgttaa tgtctggctt ctgataaagc gggccatgtt
     6301 aagggcggtt ttttcctgtt tggtcactga tgcctccgtg taagggggat ttctgttcat
     6361 gggggtaatg ataccgatga aacgagagag gatgctcacg atacgggtta ctgatgatga
     6421 acatgcccgg ttactggaac gttgtgaggg taaacaactg gcggtatgga tgcggcggga
     6481 ccagagaaaa atcactcagg gtcaatgcca gcgcttcgtt aatacagatg taggtgttcc
     6541 acagggtagc cagcagcatc ctgcgatgca gatccggaac ataatggtgc agggcgctga
     6601 cttccgcgtt tccagacttt acgaaacacg gaaaccgaag accattcatg ttgttgctca
     6661 ggtcgcagac gttttgcagc agcagtcgct tcacgttcgc tcgcgtatcg gtgattcatt
     6721 ctgctaacca gtaaggcaac cccgccagcc tagccgggtc ctcaacgaca ggagcacgat
     6781 catgcgcacc cgtggccagg acccaacgct gcccgagatg cgccgcgtgc ggctgctgga
     6841 gatggcggac gcgatggata tgttctgcca agggttggtt tgcgcattca cagttctccg
     6901 caagaattga ttggctccaa ttcttggagt ggtgaatccg ttagcgaggt gccgccggct
     6961 tccattcagg tcgaggtggc ccggctccat gcaccgcgac gcaacgcggg gaggcagaca
     7021 aggtataggg cggcgcctac aatccatgcc aacccgttcc atgtgctcgc cgaggcggca
     7081 taaatcgccg tgacgatcag cggtccaatg atcgaagtta ggctggtaag agccgcgagc
     7141 gatccttgaa gctgtccctg atggtcgtca tctacctgcc tggacagcat ggcctgcaac
     7201 gcgggcatcc cgatgccgcc ggaagcgaga agaatcataa tggggaaggc catccagcct
     7261 cgcgtcgcga acgccagcaa gacgtagccc agcgcgtcgg ccgccatgcc ggcgataatg
     7321 gcctgcttct cgccgaaacg tttggtggcg ggaccagtga cgaaggcttg agcgagggcg
     7381 tgcaagattc cgaataccgc aagcgacagg ccgatcatcg tcgcgctcca gcgaaagcgg
     7441 tcctcgccga aaatgaccca gagcgctgcc ggcacctgtc ctacgagttg catgataaag
     7501 aagacagtca taagtgcggc gacgatagtc atgccccgcg cccaccggaa ggagctgact
     7561 gggttgaagg ctctcaaggg catcggtcga gatcccggtg cctaatgagt gagctaactt
     7621 acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg
     7681 cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ccagggtggt
     7741 ttttcttttc a
//
