LOCUS       sequence-418922-        6843 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6843)
  TITLE     LIC N-terminal fusion vectors for E. coli expression
REFERENCE   2  (bases 1 to 6843)
  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"
COMMENT     Sequence Label: pET His6 NusA TEV LIC cloning vector (1N)
FEATURES             Location/Qualifiers
     source          1..6843
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             303..323
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             389..406
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(459..477)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      473..520
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      557..1012
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(644..663)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     854..875
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1105..1920)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(1105..1920)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     1828..1847
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     1912..1931
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      2042..2630
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2531..2550
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2784..2801
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    2816..2955
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(2881..2900)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3041..3063
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3057..3248)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4057..5139)
                     /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     5101..5120
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5140..5217)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     5423..5442
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     5526..5545
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5526..5544
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    5545..5569
                     /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             5584..5606
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             5625..5642
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             join(5652..6843,1..293)
                     /codon_start=1
                     /gene="nusA"
                     /product="transcription elongation protein (N utilization 
                     substance protein A)"
                     /label=NusA
                     /note="highly soluble in E. coli"
                     /translation="MNKEILAVVEAVSNEKALPREKIFEALESALATATKKKYEQEIDV
                     RVQIDRKSGDFDTFRRWLVVDEVTQPTKEITLEAARYEDESLNLGDYVEDQIESVTFDR
                     ITTQTAKQVIVQKVREAERAMVVDQFREHEGEIITGVVKKVNRDNISLDLGNNAEAVIL
                     REDMLPRENFRPGDRVRGVLYSVRPEARGAQLFVTRSKPEMLIELFRIEVPEIGEEVIE
                     IKAAARDPGSRAKIAVKTNDKRIDPVGACVGMRGARVQAVSTELGGERIDIVLWDDNPA
                     QFVINAMAPADVASIVVDEDKHTMDIAVEAGNLAQAIGRNGQNVRLASQLSGWELNVMT
                     VDDLQAKHQAEAHAAIDTFTKYLDIDEDFATVLVEEGFSTLEELAYVPMKELLEIEGLD
                     EPTVEALRERAKNALATIAQAQEESLGDNKPADDLLNLEGVDRDLAFKLAARGVCTLED
                     LAEQGIDDLADIEGLTDEKAGALIMAARNICWFGDEA"
ORIGIN
        1 ttgatgagcc gaccgttgaa gcactgcgcg agcgtgctaa aaatgcactg gccaccattg
       61 cacaggccca ggaagaaagc ctcggtgata acaaaccggc tgacgatctg ctgaaccttg
      121 aaggggtaga tcgtgatttg gcattcaaac tggccgcccg tggcgtttgt acgctggaag
      181 atctcgccga acagggcatt gatgatctgg ctgatatcga agggttgacc gacgaaaaag
      241 ccggagcact gattatggct gcccgtaaca tttgctggtt cggtgacgaa gcggggatcg
      301 aggaaaacct gtacttccaa tccaatattg gaagtggata acggatccga attcgagcgc
      361 cgtcgacaag cttgcggccg cactcgagca ccaccaccac caccactgag atccggctgc
      421 taacaaagcc cgaaaggaag ctgagttggc tgctgccacc gctgagcaat aactagcata
      481 accccttggg gcctctaaac gggtcttgag gggttttttg ctgaaaggag gaactatatc
      541 cggattggcg aatgggacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt
      601 acgcgcagcg tgaccgctac acttgccagc gccctagcgc ccgctccttt cgctttcttc
      661 ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct
      721 ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga ttagggtgat
      781 ggttcacgta gtgggccatc gccctgatag acggtttttc gccctttgac gttggagtcc
      841 acgttcttta atagtggact cttgttccaa actggaacaa cactcaaccc tatctcggtc
      901 tattcttttg atttataagg gattttgccg atttcggcct attggttaaa aaatgagctg
      961 atttaacaaa aatttaacgc gaattttaac aaactagtaa cgtttacaat ttcaggtggc
     1021 acttttcggg gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat
     1081 atgtatccgc tcatgaatta attcttagaa aaactcatcg agcatcaaat gaaactgcaa
     1141 tttattcata tcaggattat caataccata tttttgaaaa agccgtttct gtaatgaagg
     1201 agaaaactca ccgaggcagt tccataggat ggcaagatcc tggtatcggt ctgcgattcc
     1261 gactcgtcca acatcaatac aacctattaa tttcccctcg tcaaaaataa ggttatcaag
     1321 tgagaaatca ccatgagtga cgactgaatc cggtgagaat ggcaaaagtt tatgcatttc
     1381 tttccagact tgttcaacag gccagccatt acgctcgtca tcaaaatcac tcgcatcaac
     1441 caaaccgtta ttcattcgtg attgcgcctg agcgagacga aatacgcgat cgctgttaaa
     1501 aggacaatta caaacaggaa tcgaatgcaa ccggcgcagg aacactgcca gcgcatcaac
     1561 aatgttttca cctgaatcag gatattcttc taatacctgg aatgctgttt tcccggggat
     1621 cgcagtggtg agtaaccatg catcatcagg agtacggata aaatgcttga tggtcggaag
     1681 aggcataaat tccgtcagcc agtttagtct gaccatctca tctgtaacat cattggcaac
     1741 gctacctttg ccatgtttca gaaacaactc tggcgcatcg ggcttcccat acaatcgata
     1801 gattgtcgca cctgattgcc cgacattatc gcgagcccat ttatacccat ataaatcagc
     1861 atccatgttg gaatttaatc gcggcctaga gcaagacgtt tcccgttgaa tatggctcat
     1921 aacacccctt gtattactgt ttatgtaagc agacagtttt attgttcatg accaaaatcc
     1981 cttaacgtga gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt
     2041 cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac
     2101 cagcggtggt ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct
     2161 tcagcagagc gcagatacca aatactgtcc ttctagtgta gccgtagtta ggccaccact
     2221 tcaagaactc tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg
     2281 ctgccagtgg cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata
     2341 aggcgcagcg gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga
     2401 cctacaccga actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag
     2461 ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg
     2521 agcttccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac
     2581 ttgagcgtcg atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca
     2641 acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg ttctttcctg
     2701 cgttatcccc tgattctgtg gataaccgta ttaccgcctt tgagtgagct gataccgctc
     2761 gccgcagccg aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa gagcgcctga
     2821 tgcggtattt tctccttacg catctgtgcg gtatttcaca ccgcaatggt gcactctcag
     2881 tacaatctgc tctgatgccg catagttaag ccagtataca ctccgctatc gctacgtgac
     2941 tgggtcatgg ctgcgccccg acacccgcca acacccgctg acgcgccctg acgggcttgt
     3001 ctgctcccgg catccgctta cagacaagct gtgaccgtct ccgggagctg catgtgtcag
     3061 aggttttcac cgtcatcacc gaaacgcgcg aggcagctgc ggtaaagctc atcagcgtgg
     3121 tcgtgaagcg attcacagat gtctgcctgt tcatccgcgt ccagctcgtt gagtttctcc
     3181 agaagcgtta atgtctggct tctgataaag cgggccatgt taagggcggt tttttcctgt
     3241 ttggtcactg atgcctccgt gtaaggggga tttctgttca tgggggtaat gataccgatg
     3301 aaacgagaga ggatgctcac gatacgggtt actgatgatg aacatgcccg gttactggaa
     3361 cgttgtgagg gtaaacaact ggcggtatgg atgcggcggg accagagaaa aatcactcag
     3421 ggtcaatgcc agcgcttcgt taatacagat gtaggtgttc cacagggtag ccagcagcat
     3481 cctgcgatgc agatccggaa cataatggtg cagggcgctg acttccgcgt ttccagactt
     3541 tacgaaacac ggaaaccgaa gaccattcat gttgttgctc aggtcgcaga cgttttgcag
     3601 cagcagtcgc ttcacgttcg ctcgcgtatc ggtgattcat tctgctaacc agtaaggcaa
     3661 ccccgccagc ctagccgggt cctcaacgac aggagcacga tcatgcgcac ccgtggggcc
     3721 gccatgccgg cgataatggc ctgcttctcg ccgaaacgtt tggtggcggg accagtgacg
     3781 aaggcttgag cgagggcgtg caagattccg aataccgcaa gcgacaggcc gatcatcgtc
     3841 gcgctccagc gaaagcggtc ctcgccgaaa atgacccaga gcgctgccgg cacctgtcct
     3901 acgagttgca tgataaagaa gacagtcata agtgcggcga cgatagtcat gccccgcgcc
     3961 caccggaagg agctgactgg gttgaaggct ctcaagggca tcggtcgaga tcccggtgcc
     4021 taatgagtga gctaacttac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga
     4081 aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt
     4141 attgggcgcc agggtggttt ttcttttcac cagtgagacg ggcaacagct gattgccctt
     4201 caccgcctgg ccctgagaga gttgcagcaa gcggtccacg ctggtttgcc ccagcaggcg
     4261 aaaatcctgt ttgatggtgg ttaacggcgg gatataacat gagctgtctt cggtatcgtc
     4321 gtatcccact accgagatat ccgcaccaac gcgcagcccg gactcggtaa tggcgcgcat
     4381 tgcgcccagc gccatctgat cgttggcaac cagcatcgca gtgggaacga taccctcatt
     4441 cagcatttgc atggtttgtt gaaaaccgga catggcactc cagtcgcctt cccgttccgc
     4501 tatcggctga atttgattgc gagtgagata tttatgccag ccagccagac gcagacgcgc
     4561 cgagacagaa cttaatgggc ccgctaacag cgcgatttgc tggtgaccca atgcgaccag
     4621 atgctccacg cccagtcgcg taccgtcttc atgggagaaa ataatactgt tgatgggtgt
     4681 ctggtcagag acatcaagaa ataacgccgg aacattagtg caggcagctt ccacagcaat
     4741 ggcatcctgg tcatccagcg gatagttaat gatcagccca ctgacgcgtt gcgcgagaag
     4801 attgtgcacc gccgctttac aggcttcgac gccgcttcgt tctaccatcg acaccaccac
     4861 gctggcaccc agttgatcgg cgcgagattt aatcgccgcg acaatttgcg acggcgcgtg
     4921 cagggccaga ctggaggtgg caacgccaat cagcaacgac tgtttgcccg ccagttgttg
     4981 tgccacgcgg ttgggaatgt aattcagctc cgccatcgcc gcttccactt tttcccgcgt
     5041 tttcgcagaa acgtggctgg cctggttcac cacgcgggaa acggtctgat aagagacacc
     5101 ggcatactct gcgacatcgt ataacgttac tggtttcaca ttcaccaccc tgaattgact
     5161 ctcttccggg cgctatcatg ccataccgcg aaaggttttg cgccattcga tggtgtccgg
     5221 gatctcgacg ctctccctta tgcgactcct gcattaggaa gcagcccagt agtaggttga
     5281 ggccgttgag caccgccgcc gcaaggaatg gtgcatgcaa ggagatggcg cccaacagtc
     5341 ccccggccac ggggcctgcc accataccca cgccgaaaca agcgctcatg agcccgaagt
     5401 ggcgagcccg atcttcccca tcggtgatgt cggcgatata ggcgccagca accgcacctg
     5461 tggcgccggt gatgccggcc acgatgcgtc cggcgtagag gatcgagatc tcgatcccgc
     5521 gaaattaata cgactcacta taggggaatt gtgagcggat aacaattccc ctctagaaat
     5581 aattttgttt aactttaaga aggagatata ccatgggttc ttctcaccat caccatcacc
     5641 atggttcttc tatgaacaaa gaaattttgg ctgtagttga agccgtatcc aatgaaaagg
     5701 cgctacctcg cgagaagatt ttcgaagcat tggaaagcgc gctggcgaca gcaacaaaga
     5761 aaaaatatga acaagagatc gacgtccgcg tacagatcga tcgcaaaagc ggtgattttg
     5821 acactttccg tcgctggtta gttgttgatg aagtcaccca gccgaccaag gaaatcaccc
     5881 ttgaagccgc acgttatgaa gatgaaagcc tgaacctggg cgattacgtt gaagatcaga
     5941 ttgagtctgt tacctttgac cgtatcacta cccagacggc aaaacaggtt atcgtgcaga
     6001 aagtgcgtga agccgaacgt gcgatggtgg ttgatcagtt ccgtgaacac gaaggtgaaa
     6061 tcatcaccgg cgtggtgaaa aaagtaaacc gcgacaacat ctctctggat ctgggcaaca
     6121 acgctgaagc cgtgatcctg cgcgaagata tgctgccgcg tgaaaacttc cgccctggcg
     6181 accgcgttcg tggcgtgctc tattccgttc gcccggaagc gcgtggcgcg caactgttcg
     6241 tcactcgttc caagccggaa atgctgatcg aactgttccg tattgaagtg ccagaaatcg
     6301 gcgaagaagt gattgaaatt aaagcagcgg ctcgcgatcc gggttctcgt gcgaaaatcg
     6361 cggtgaaaac caacgataaa cgtatcgatc cggtaggtgc ttgcgtaggt atgcgtggcg
     6421 cgcgtgttca ggcggtgtct actgaactgg gtggcgagcg tatcgatatc gtcctgtggg
     6481 atgataaccc ggcgcagttc gtgattaacg caatggcacc ggcagacgtt gcttctatcg
     6541 tggtggatga agataaacac accatggata tcgccgttga agccggtaac ctggcgcagg
     6601 cgattggccg taacggtcag aacgtgcgtc tggcttcgca gctgagcggt tgggaactca
     6661 acgtgatgac cgttgacgac ctgcaggcta agcatcaggc ggaagcgcac gcagcgatcg
     6721 acaccttcac caaatatctc gacatcgacg aagacttcgc gactgttctg gtagaagaag
     6781 gcttctcgac gctggaagaa ttggcctatg tgccgatgaa agagctgttg gaaatcgaag
     6841 gcc
//
