LOCUS       sequence-323836-        6420 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 6420)
  AUTHORS   Grinkova YV, Denisov IG, Sligar SG
  TITLE     Engineering extended membrane scaffold proteins for self-assembly of
            soluble nanoscale lipid bilayers.
  JOURNAL   Protein Eng Des Sel. 2010 Nov . 23(11):843-8.
  PUBMED    20817758
REFERENCE   2  (bases 1 to 6420)
  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: "Protein Eng
            Des Sel. 2010 Nov . 23(11):843-8."
COMMENT     Sequence Label: pMSP2N2
FEATURES             Location/Qualifiers
     source          1..6420
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     939..958
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(978..1055)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     1261..1280
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     1364..1383
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1364..1382
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1383..1407
                     /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             1422..1444
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             1457..1477
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=7xHis
                     /translation="HHHHHHH"
     CDS             1499..1519
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             2649..2666
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2719..2737)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2733..2780
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      2817..3272
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2904..2923)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3114..3135
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(3364..4179)
                     /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(3364..4179)
                     /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     4087..4106
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     4171..4190
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      4301..4889
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4790..4809
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5043..5060
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    5075..5214
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(5140..5159)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5300..5322
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(join(6315..6420,1..977))
                     /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
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLTGWHKYLTRDQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
ORIGIN
        1 cttttcacca gtgagacggg caacagctga ttgcccttca ccgcctggcc ctgagagagt
       61 tgcagcaagc ggtccacgct ggtttgcccc agcaggcgaa aatcctgttt gatggtggtt
      121 aacggcggga tataacatga gctgtcttcg gtatcgtcgt atcccactac cgagatatcc
      181 gcaccaacgc gcagcccgga ctcggtaatg gcgcgcattg cgcccagcgc catctgatcg
      241 ttggcaacca gcatcgcagt gggaacgatg ccctcattca gcatttgcat ggtttgttga
      301 aaaccggaca tggcactcca gtcgccttcc cgttccgcta tcggctgaat ttgatcgcga
      361 gtgagatatt tatgccagcc agtcagacgc agacgcgccg agacagaact taatgggccc
      421 gctaacagcg cgatttgctg gtgacccaat gcgaccagat gctccacgcc cagtcgcgta
      481 ccgtcttcat gggagaaaat aatactgttg atgggtgtct ggtcagagac atcaagaaat
      541 aacgccggaa cattagtgca ggcagcttcc acagcaatgg catcctggtc atccagcgga
      601 tagttaatga tcagcccact gacgcgttgc gcgagaagat tgtgcaccgc cgctttacag
      661 gcttcgacgc cgcttcgttc taccatcgac accaccacgc tggcacccag ttgatcggcg
      721 cgagatttaa tcgccgcgac aatttgcgac ggcgcgtgca gggccagact ggaggtggca
      781 acgccaatca gcaacgactg tttgcccgcc agttgttgtg ccacgcggtt gggaatgtaa
      841 ttcagctccg ccatcgccgc ttccactttt tcccgcgttt tcgcagaaac gtggctggcc
      901 tggttcacca cgcgggaaac ggtctgataa gagacaccgg catactctgc gacatcgtat
      961 aacgttactg gtttcacatt caccaccctg aattgactct cttccgggcg ctatcatgcc
     1021 ataccgcgaa aggttttgcg ccattcgatg gtgtccggga tctcgacgct ctcccttatg
     1081 cgactcctgc attaggaagc agcccagtag taggttgagg ccgttgagca ccgccgccgc
     1141 aaggaatggt gcatgcaagg agatggcgcc caacagtccc ccggccacgg ggcctgccac
     1201 catacccacg ccgaaacaag cgctcatgag cccgaagtgg cgagcccgat cttccccatc
     1261 ggtgatgtcg gcgatatagg cgccagcaac cgcacctgtg gcgccggtga tgccggccac
     1321 gatgcgtccg gcgtagagga tcgagatctc gatcccgcga aattaatacg actcactata
     1381 ggggaattgt gagcggataa caattcccct ctagaaataa ttttgtttaa ctttaagaag
     1441 gagatatacc atgggtcatc atcatcatca tcatcacgat tatgatattc ctactactga
     1501 gaatttgtat tttcagggtt ctaccttcag taaacttcgc gaacaactgg gccccgtgac
     1561 gcaggaattc tgggacaacc tggaaaaaga aaccgaggga ctgcgtcagg aaatgtccaa
     1621 agatttagaa gaggtgaagg ccaaggttca gccatatctc gatgactttc agaaaaaatg
     1681 gcaggaagag atggaattat atcgtcaaaa ggtggaaccg ctgcgtgcgg aactgcaaga
     1741 gggggcacgc caaaaactcc atgagctcca agagaagctc agcccattag gcgaagaaat
     1801 gcgcgatcgc gcccgtgcac atgttgatgc actccggact catttggcgc cgtattcgga
     1861 tgaacttcgc cagcgtttgg ccgcacgtct cgaggcgctg aaagaaaacg ggggtgcccg
     1921 cttggctgag taccacgcga aagcgacaga acacctgagc accttgagcg aaaaagcgaa
     1981 accggcgctg gaagatctac gccagggctt attgcctgtt cttgagagct ttaaagtcag
     2041 ttttctgtca gctctggaag aatatactaa aaagctgaat acccagggta cccccgtgac
     2101 gcaggaattc tgggacaacc tggaaaaaga aaccgaggga ctgcgtcagg aaatgtccaa
     2161 agatttagaa gaggtgaagg ccaaggttca gccatatctc gatgactttc agaaaaaatg
     2221 gcaggaagag atggaattat atcgtcaaaa ggtggaaccg ctgcgtgcgg aactgcaaga
     2281 gggggcacgc caaaaactcc atgagctcca agagaagctc agcccattag gcgaagaaat
     2341 gcgcgatcgc gcccgtgcac atgttgatgc actccggact catttggcgc cgtattcgga
     2401 tgaacttcgc cagcgtttgg ccgcacgtct cgaggcgctg aaagaaaacg ggggtgcccg
     2461 cttggctgag taccacgcga aagcgacaga acacctgagc accttgagcg aaaaagcgaa
     2521 accggcgctg gaagatctac gccagggctt attgcctgtt cttgagagct ttaaagtcag
     2581 ttttctgtca gctctggaag aatatactaa aaagctgaat acccagtaag cttgcggccg
     2641 cactcgagca ccaccaccac caccactgag atccggctgc taacaaagcc cgaaaggaag
     2701 ctgagttggc tgctgccacc gctgagcaat aactagcata accccttggg gcctctaaac
     2761 gggtcttgag gggttttttg ctgaaaggag gaactatatc cggattggcg aatgggacgc
     2821 gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac
     2881 acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt
     2941 cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc
     3001 tttacggcac ctcgacccca aaaaacttga ttagggtgat ggttcacgta gtgggccatc
     3061 gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact
     3121 cttgttccaa actggaacaa cactcaaccc tatctcggtc tattcttttg atttataagg
     3181 gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc
     3241 gaattttaac aaaatattaa cgcttacaat ttaggtggca cttttcgggg aaatgtgcgc
     3301 ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct catgaattaa
     3361 ttcttagaaa aactcatcga gcatcaaatg aaactgcaat ttattcatat caggattatc
     3421 aataccatat ttttgaaaaa gccgtttctg taatgaagga gaaaactcac cgaggcagtt
     3481 ccataggatg gcaagatcct ggtatcggtc tgcgattccg actcgtccaa catcaataca
     3541 acctattaat ttcccctcgt caaaaataag gttatcaagt gagaaatcac catgagtgac
     3601 gactgaatcc ggtgagaatg gcaaaagttt atgcatttct ttccagactt gttcaacagg
     3661 ccagccatta cgctcgtcat caaaatcact cgcatcaacc aaaccgttat tcattcgtga
     3721 ttgcgcctga gcgagacgaa atacgcgatc gctgttaaaa ggacaattac aaacaggaat
     3781 cgaatgcaac cggcgcagga acactgccag cgcatcaaca atattttcac ctgaatcagg
     3841 atattcttct aatacctgga atgctgtttt cccggggatc gcagtggtga gtaaccatgc
     3901 atcatcagga gtacggataa aatgcttgat ggtcggaaga ggcataaatt ccgtcagcca
     3961 gtttagtctg accatctcat ctgtaacatc attggcaacg ctacctttgc catgtttcag
     4021 aaacaactct ggcgcatcgg gcttcccata caatcgatag attgtcgcac ctgattgccc
     4081 gacattatcg cgagcccatt tatacccata taaatcagca tccatgttgg aatttaatcg
     4141 cggcctagag caagacgttt cccgttgaat atggctcata acaccccttg tattactgtt
     4201 tatgtaagca gacagtttta ttgttcatga ccaaaatccc ttaacgtgag ttttcgttcc
     4261 actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgaaatcct ttttttctgc
     4321 gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg
     4381 atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa
     4441 atactgtcct tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc
     4501 ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt
     4561 gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa
     4621 cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc
     4681 tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc
     4741 cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct
     4801 ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat
     4861 gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc
     4921 tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct gattctgtgg
     4981 ataaccgtat taccgccttt gagtgagctg ataccgctcg ccgcagccga acgaccgagc
     5041 gcagcgagtc agtgagcgag gaagcggaag agcgcctgat gcggtatttt ctccttacgc
     5101 atctgtgcgg tatttcacac cgcaatggtg cactctcagt acaatctgct ctgatgccgc
     5161 atagttaagc cagtatacac tccgctatcg ctacgtgact gggtcatggc tgcgccccga
     5221 cacccgccaa cacccgctga cgcgccctga cgggcttgtc tgctcccggc atccgcttac
     5281 agacaagctg tgaccgtctc cgggagctgc atgtgtcaga ggttttcacc gtcatcaccg
     5341 aacgcgcgag gcagctgcgg taaagctcat cagcgtggtc gtgaagcgat tcacagatgt
     5401 ctgcctgttc atccgcgtcc agctcgttga gtttctccag aagcgttaat gtctggcttc
     5461 tgataaagcg ggccatgtta agggcggttt tttcctgttt ggtcactgat gcctccgtgt
     5521 aagggggatt tctgttcatg ggggtaatga taccgatgaa acgagagagg atgctcacga
     5581 tacgggttac tgatgatgaa catgcccggt tactggaacg ttgtgagggt aaacaactgg
     5641 cggtatggat gcggcgggac cagagaaaaa tcactcaggg tcaatgccag cgcttcgtta
     5701 atacagatgt aggtgttcca cagggtagcc agcagcatcc tgcgatgcag atccggaaca
     5761 taatggtgca gggcgctgac ttccgcgttt ccagacttta cgaaacacgg aaaccgaaga
     5821 ccattcatgt tgttgctcag gtcgcagacg ttttgcagca gcagtcgctt cacgttcgct
     5881 cgcgtatcgg tgattcattc tgctaaccag taaggcaacc ccgccagcct agccgggtcc
     5941 tcaacgacag gagcacgatc atgcgcaccc gtggggccgc catgccggcg ataatggcct
     6001 gcttctcgcc gaaacgtttg gtggcgggac cagtgacgaa ggcttgagcg agggcgtgca
     6061 agattccgaa taccgcaagc gacaggccga tcatcgtcgc gctccagcga aagcggtcct
     6121 cgccgaaaat gacccagagc gctgccggca cctgtcctac gagttgcatg ataaagaaga
     6181 cagtcataag tgcggcgacg atagtcatgc cccgcgccca ccggaaggag ctgactgggt
     6241 tgaaggctct caagggcatc ggtcgagatc ccggtgccta atgagtgagc taacttacat
     6301 taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt
     6361 aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgccag ggtggttttt
//
