LOCUS       sequence-479205-        6561 bp    DNA     circular SYN 22-JUL-2025
DEFINITION  VidN construct that can be bacterially expressed, used to 
            demonstrate intein splicing reaction..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6561)
  AUTHORS   Gharios R, Li A, Kopyeva I, Francis RM, DeForest CA
  TITLE     One-Step Purification and N-Terminal Functionalization of Bioactive 
            Proteins via Atypically Split Inteins.
  JOURNAL   Bioconjug Chem. 2024 Jun 19;35(6):750-757. doi: 
            10.1021/acs.bioconjchem.4c00223. Epub 2024 May 30.
  PUBMED    38815180
REFERENCE   2  (bases 1 to 6561)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jul 22, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1021/acs.bioconjchem.4c00223"; journalName: "Bioconjug Chem"; 
            date: "2024-06-19- 19"; volume: "35"; issue: "6"; pages: "750-757"
COMMENT     Sequence Label: pET-21_MBP-VidN
FEATURES             Location/Qualifiers
     source          1..6561
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(470..1552)
                     /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     1514..1533
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(1553..1630)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     1836..1855
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     1939..1958
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1939..1957
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1958..1982
                     /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             1997..2019
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             3176..3187
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(3297..3315)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      3311..3358
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      3395..3850
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3482..3501)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3692..3713
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        3877..3981
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3982..4050,4051..4842)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(4200..4219)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5013..5601
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5502..5521
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5755..5772
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    5787..5929
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(5855..5874)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6015..6037
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(6031..6222)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
ORIGIN
        1 agacgttttg cagcagcagt cgcttcacgt tcgctcgcgt atcggtgatt cattctgcta
       61 accagtaagg caaccccgcc agcctagccg ggtcctcaac gacaggagca cgatcatgcg
      121 cacccgtggg gccgccatgc cggcgataat ggcctgcttc tcgccgaaac gtttggtggc
      181 gggaccagtg acgaaggctt gagcgagggc gtgcaagatt ccgaataccg caagcgacag
      241 gccgatcatc gtcgcgctcc agcgaaagcg gtcctcgccg aaaatgaccc agagcgctgc
      301 cggcacctgt cctacgagtt gcatgataaa gaagacagtc ataagtgcgg cgacgatagt
      361 catgccccgc gcccaccgga aggagctgac tgggttgaag gctctcaagg gcatcggtcg
      421 agatcccggt gcctaatgag tgagctaact tacattaatt gcgttgcgct cactgcccgc
      481 tttccagtcg ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag
      541 aggcggtttg cgtattgggc gccagggtgg tttttctttt caccagtgag acgggcaaca
      601 gctgattgcc cttcaccgcc tggccctgag agagttgcag caagcggtcc acgctggttt
      661 gccccagcag gcgaaaatcc tgtttgatgg tggttaacgg cgggatataa catgagctgt
      721 cttcggtatc gtcgtatccc actaccgaga tatccgcacc aacgcgcagc ccggactcgg
      781 taatggcgcg cattgcgccc agcgccatct gatcgttggc aaccagcatc gcagtgggaa
      841 cgatgccctc attcagcatt tgcatggttt gttgaaaacc ggacatggca ctccagtcgc
      901 cttcccgttc cgctatcggc tgaatttgat tgcgagtgag atatttatgc cagccagcca
      961 gacgcagacg cgccgagaca gaacttaatg ggcccgctaa cagcgcgatt tgctggtgac
     1021 ccaatgcgac cagatgctcc acgcccagtc gcgtaccgtc ttcatgggag aaaataatac
     1081 tgttgatggg tgtctggtca gagacatcaa gaaataacgc cggaacatta gtgcaggcag
     1141 cttccacagc aatggcatcc tggtcatcca gcggatagtt aatgatcagc ccactgacgc
     1201 gttgcgcgag aagattgtgc accgccgctt tacaggcttc gacgccgctt cgttctacca
     1261 tcgacaccac cacgctggca cccagttgat cggcgcgaga tttaatcgcc gcgacaattt
     1321 gcgacggcgc gtgcagggcc agactggagg tggcaacgcc aatcagcaac gactgtttgc
     1381 ccgccagttg ttgtgccacg cggttgggaa tgtaattcag ctccgccatc gccgcttcca
     1441 ctttttcccg cgttttcgca gaaacgtggc tggcctggtt caccacgcgg gaaacggtct
     1501 gataagagac accggcatac tctgcgacat cgtataacgt tactggtttc acattcacca
     1561 ccctgaattg actctcttcc gggcgctatc atgccatacc gcgaaaggtt ttgcgccatt
     1621 cgatggtgtc cgggatctcg acgctctccc ttatgcgact cctgcattag gaagcagccc
     1681 agtagtaggt tgaggccgtt gagcaccgcc gccgcaagga atggtgcatg caaggagatg
     1741 gcgcccaaca gtcccccggc cacggggcct gccaccatac ccacgccgaa acaagcgctc
     1801 atgagcccga agtggcgagc ccgatcttcc ccatcggtga tgtcggcgat ataggcgcca
     1861 gcaaccgcac ctgtggcgcc ggtgatgccg gccacgatgc gtccggcgta gaggatcgag
     1921 atctcgatcc cgcgaaatta atacgactca ctatagggga attgtgagcg gataacaatt
     1981 cccctctaga aataattttg tttaacttta agaaggagat atacatatga agacagaaga
     2041 aggcaaactc gttatctgga tcaatgggga caagggctac aatggcctgg cggaagtcgg
     2101 caagaaattt gaaaaagata ccgggatcaa agtgaccgtt gaacacccgg ataaattgga
     2161 ggaaaagttc ccgcaagtgg ccgcgaccgg agatggtcct gacataattt tttgggcgca
     2221 cgatcggttt ggtgggtacg ctcaatcggg ccttctagca gaaattacgc cggacaaagc
     2281 gtttcaggat aaactctatc cgtttacctg ggacgcggta cgctataatg gaaaactgat
     2341 tgcatatccg attgcagtgg aagcgctgag ccttatttac aataaagatc tgcttcctaa
     2401 tccgcccaag acctgggaag aaattccggc actggacaaa gaactgaaag ccaaaggtaa
     2461 aagcgcactt atgttcaatc tgcaggagcc gtatttcacc tggccgctaa tcgcggccga
     2521 cggcggttat gcctttaaat atgaaaacgg caagtatgat atcaaagatg ttggtgtaga
     2581 caacgccggt gccaaagctg gcttgacctt cttggtggat ttaattaaaa acaaacacat
     2641 gaacgcagat acggattatt caattgcgga ggccgcgttc aacaagggcg agactgcaat
     2701 gactattaac ggtccatggg cgtggtcaaa catcgacact tctaaagtga actacggtgt
     2761 cactgtgctc ccgacgttta aaggtcagcc gtccaaaccg tttgtgggcg tgctgagtgc
     2821 aggaatcaac gcggcgagtc cgaataaaga attagcgaaa gaatttttag aaaactatct
     2881 gttaacggat gagggtctgg aagctgtcaa caaagacaag cctttggggg ccgtggcact
     2941 gaaatcatat gaggaagagt tagcaaagga tccccgcatc gctgcgacca tggaaaatgc
     3001 ccaaaaaggc gaaattatgc caaatatccc ccagatgagc gctttctggt acgctgtcag
     3061 gacagccgtc attaacgccg cgtccggccg tcaaactgtt gatgaagctt tgaaagatgc
     3121 acagactaat tcctcgagca ataataacaa caataacaac aataataacc tgggaatcga
     3181 ggggcgtgaa agcggctgcc tgccgaaaga agcggtggtg cagattcgcc tgaccaaaaa
     3241 aggctatgat ccggctgcta acaaagcccg aaaggaagct gagttggctg ctgccaccgc
     3301 tgagcaataa ctagcataac cccttggggc ctctaaacgg gtcttgaggg gttttttgct
     3361 gaaaggagga actatatccg gattggcgaa tgggacgcgc cctgtagcgg cgcattaagc
     3421 gcggcgggtg tggtggttac gcgcagcgtg accgctacac ttgccagcgc cctagcgccc
     3481 gctcctttcg ctttcttccc ttcctttctc gccacgttcg ccggctttcc ccgtcaagct
     3541 ctaaatcggg ggctcccttt agggttccga tttagtgctt tacggcacct cgaccccaaa
     3601 aaacttgatt agggtgatgg ttcacgtagt gggccatcgc cctgatagac ggtttttcgc
     3661 cctttgacgt tggagtccac gttctttaat agtggactct tgttccaaac tggaacaaca
     3721 ctcaacccta tctcggtcta ttcttttgat ttataaggga ttttgccgat ttcggcctat
     3781 tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga attttaacaa aatattaacg
     3841 tttacaattt caggtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt
     3901 ttctaaatac attcaaatat gtatccgctc atgagacaat aaccctgata aatgcttcaa
     3961 taatattgaa aaaggaagag tatgagtatt caacatttcc gtgtcgccct tattcccttt
     4021 tttgcggcat tttgccttcc tgtttttgct cacccagaaa cgctggtgaa agtaaaagat
     4081 gctgaagatc agttgggtgc acgagtgggt tacatcgaac tggatctcaa cagcggtaag
     4141 atccttgaga gttttcgccc cgaagaacgt tttccaatga tgagcacttt taaagttctg
     4201 ctatgtggcg cggtattatc ccgtattgac gccgggcaag agcaactcgg tcgccgcata
     4261 cactattctc agaatgactt ggttgagtac tcaccagtca cagaaaagca tcttacggat
     4321 ggcatgacag taagagaatt atgcagtgct gccataacca tgagtgataa cactgcggcc
     4381 aacttacttc tgacaacgat cggaggaccg aaggagctaa ccgctttttt gcacaacatg
     4441 ggggatcatg taactcgcct tgatcgttgg gaaccggagc tgaatgaagc cataccaaac
     4501 gacgagcgtg acaccacgat gcctgcagca atggcaacaa cgttgcgcaa actattaact
     4561 ggcgaactac ttactctagc ttcccggcaa caattaatag actggatgga ggcggataaa
     4621 gttgcaggac cacttctgcg ctcggccctt ccggctggct ggtttattgc tgataaatct
     4681 ggagccggtg agcgtgggtc tcgcggtatc attgcagcac tggggccaga tggtaagccc
     4741 tcccgtatcg tagttatcta cacgacgggg agtcaggcaa ctatggatga acgaaataga
     4801 cagatcgctg agataggtgc ctcactgatt aagcattggt aactgtcaga ccaagtttac
     4861 tcatatatac tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag
     4921 atcctttttg ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg
     4981 tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc
     5041 tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag
     5101 ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtc
     5161 cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac
     5221 ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc
     5281 gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt
     5341 tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt
     5401 gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc
     5461 ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt
     5521 tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca
     5581 ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt
     5641 tgctggcctt ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt
     5701 attaccgcct ttgagtgagc tgataccgct cgccgcagcc gaacgaccga gcgcagcgag
     5761 tcagtgagcg aggaagcgga agagcgcctg atgcggtatt ttctccttac gcatctgtgc
     5821 ggtatttcac accgcatata tggtgcactc tcagtacaat ctgctctgat gccgcatagt
     5881 taagccagta tacactccgc tatcgctacg tgactgggtc atggctgcgc cccgacaccc
     5941 gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg cttacagaca
     6001 agctgtgacc gtctccggga gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg
     6061 cgcgaggcag ctgcggtaaa gctcatcagc gtggtcgtga agcgattcac agatgtctgc
     6121 ctgttcatcc gcgtccagct cgttgagttt ctccagaagc gttaatgtct ggcttctgat
     6181 aaagcgggcc atgttaaggg cggttttttc ctgtttggtc actgatgcct ccgtgtaagg
     6241 gggatttctg ttcatggggg taatgatacc gatgaaacga gagaggatgc tcacgatacg
     6301 ggttactgat gatgaacatg cccggttact ggaacgttgt gagggtaaac aactggcggt
     6361 atggatgcgg cgggaccaga gaaaaatcac tcagggtcaa tgccagcgct tcgttaatac
     6421 agatgtaggt gttccacagg gtagccagca gcatcctgcg atgcagatcc ggaacataat
     6481 ggtgcagggc gctgacttcc gcgtttccag actttacgaa acacggaaac cgaagaccat
     6541 tcatgttgtt gctcaggtcg c
//
