LOCUS       sequence-446873-        5884 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Membrane scaffold protein (MSP1D1) with Strep-II tag. pMSP1D1, 7xHis
            tag swapped for a Strep-II tag..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5884)
  AUTHORS   Pettersen JM, McCracken O, Robinson AS
  TITLE     Ligand binding kinetics to evaluate the function and stability of 
            A(2A)R in nanodiscs.
  JOURNAL   Biophys J. 2025 Jan 21;124(2):440-457. doi: 
            10.1016/j.bpj.2024.12.018. Epub 2024 Dec 17.
  PUBMED    39690743
REFERENCE   2  (bases 1 to 5884)
  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.bpj.2024.12.018"; journalName: "Biophys J"; date: 
            "2025-01-21- 21"; volume: "124"; issue: "2"; pages: "440-457"
COMMENT     Sequence Label: pMSP1D1-Strep-II
FEATURES             Location/Qualifiers
     source          1..5884
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     55..74
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     terminator      complement(198..245)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     241..259
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(312..329)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(919..939)
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             complement(961..984)
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     RBS             997..1019
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    1034..1058
                     /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)."
     primer_bind     complement(1058..1077)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1059..1077)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1161..1180)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        1386..1463
                     /gene="lacI"
                     /label=lacI promoter
     CDS             1464..2546
                     /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     complement(1483..1502)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             3355..3546
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(3540..3562)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    3648..3787
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     3703..3722
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(3802..3819)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3973..4561)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4053..4072)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(4672..4691)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             4683..5498
                     /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             4683..5498
                     /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     complement(4756..4775)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      complement(join(5590..5884,1..161))
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5727..5748)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
ORIGIN
        1 gagcccccga tttagagctt gacggggaaa gccggcgaac gtggcgagaa aggaagggaa
       61 gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc tgcgcgtaac
      121 caccacaccc gccgcgctta atgcgccgct acagggcgcg tcccattcgc caatccggat
      181 atagttcctc ctttcagcaa aaaacccctc aagacccgtt tagaggcccc aaggggttat
      241 gctagttatt gctcagcggt ggcagcagcc aactcagctt cctttcgggc tttgttagca
      301 gccggatctc agtggtggtg gtggtggtgc tcgagtgcgg ccgcaagctt actgggtatt
      361 cagcttttta gtatattctt ccagagctga cagaaaactg actttaaagc tctcaagaac
      421 aggcaataag ccctggcgta gatcttccag cgccggtttc gctttttcgc tcaaggtgct
      481 caggtgttct gtcgctttcg cgtggtactc agccaagcgg gcacccccgt tttctttcag
      541 cgcctcgaga cgtgcggcca aacgctggcg aagttcatcc gaatacggcg ccaaatgagt
      601 ccggagtgca tcaacatgtg cacgggcgcg atcgcgcatt tcttcgccta atgggctgag
      661 cttctcttgg agctcatgga gtttttggcg tgccccctct tgcagttccg cacgcagcgg
      721 ttccaccttt tgacgatata attccatctc ttcctgccat tttttctgaa agtcatcgag
      781 atatggctga accttggcct tcacctcttc taaatctttg gacatttcct gacgcagtcc
      841 ctcggtttct ttttccaggt tgtcccagaa ttcctgcgtc acggggccca gttgttcgcg
      901 aagtttactg aaggtagaac cctgaaaata caaattctca gtagtaggaa tatcataatc
      961 tttttcaaac tgcggatggc tccaacccat ggtatatctc cttcttaaag ttaaacaaaa
     1021 ttatttctag aggggaattg ttatccgctc acaattcccc tatagtgagt cgtattaatt
     1081 tcgcgggatc gagatctcga tcctctacgc cggacgcatc gtggccggca tcaccggcgc
     1141 cacaggtgcg gttgctggcg cctatatcgc cgacatcacc gatggggaag atcgggctcg
     1201 ccacttcggg ctcatgagcg cttgtttcgg cgtgggtatg gtggcaggcc ccgtggccgg
     1261 gggactgttg ggcgccatct ccttgcatgc accattcctt gcggcggcgg tgctcaacgg
     1321 cctcaaccta ctactgggct gcttcctaat gcaggagtcg cataagggag agcgtcgaga
     1381 tcccggacac catcgaatgg cgcaaaacct ttcgcggtat ggcatgatag cgcccggaag
     1441 agagtcaatt cagggtggtg aatgtgaaac cagtaacgtt atacgatgtc gcagagtatg
     1501 ccggtgtctc ttatcagacc gtttcccgcg tggtgaacca ggccagccac gtttctgcga
     1561 aaacgcggga aaaagtggaa gcggcgatgg cggagctgaa ttacattccc aaccgcgtgg
     1621 cacaacaact ggcgggcaaa cagtcgttgc tgattggcgt tgccacctcc agtctggccc
     1681 tgcacgcgcc gtcgcaaatt gtcgcggcga ttaaatctcg cgccgatcaa ctgggtgcca
     1741 gcgtggtggt gtcgatggta gaacgaagcg gcgtcgaagc ctgtaaagcg gcggtgcaca
     1801 atcttctcgc gcaacgcgtc agtgggctga tcattaacta tccgctggat gaccaggatg
     1861 ccattgctgt ggaagctgcc tgcactaatg ttccggcgtt atttcttgat gtctctgacc
     1921 agacacccat caacagtatt attttctccc atgaagacgg tacgcgactg ggcgtggagc
     1981 atctggtcgc attgggtcac cagcaaatcg cgctgttagc gggcccatta agttctgtct
     2041 cggcgcgtct gcgtctggct ggctggcata aatatctcac tcgcaatcaa attcagccga
     2101 tagcggaacg ggaaggcgac tggagtgcca tgtccggttt tcaacaaacc atgcaaatgc
     2161 tgaatgaggg catcgttccc actgcgatgc tggttgccaa cgatcagatg gcgctgggcg
     2221 caatgcgcgc cattaccgag tccgggctgc gcgttggtgc ggatatctcg gtagtgggat
     2281 acgacgatac cgaagacagc tcatgttata tcccgccgtt aaccaccatc aaacaggatt
     2341 ttcgcctgct ggggcaaacc agcgtggacc gcttgctgca actctctcag ggccaggcgg
     2401 tgaagggcaa tcagctgttg cccgtctcac tggtgaaaag aaaaaccacc ctggcgccca
     2461 atacgcaaac cgcctctccc cgcgcgttgg ccgattcatt aatgcagctg gcacgacagg
     2521 tttcccgact ggaaagcggg cagtgagcgc aacgcaatta atgtaagtta gctcactcat
     2581 taggcaccgg gatctcgacc gatgcccttg agagccttca acccagtcag ctccttccgg
     2641 tgggcgcggg gcatgactat cgtcgccgca cttatgactg tcttctttat catgcaactc
     2701 gtaggacagg tgccggcagc gctctgggtc attttcggcg aggaccgctt tcgctggagc
     2761 gcgacgatga tcggcctgtc gcttgcggta ttcggaatct tgcacgccct cgctcaagcc
     2821 ttcgtcactg gtcccgccac caaacgtttc ggcgagaagc aggccattat cgccggcatg
     2881 gcggccccac gggtgcgcat gatcgtgctc ctgtcgttga ggacccggct aggctggcgg
     2941 ggttgcctta ctggttagca gaatgaatca ccgatacgcg agcgaacgtg aagcgactgc
     3001 tgctgcaaaa cgtctgcgac ctgagcaaca acatgaatgg tcttcggttt ccgtgtttcg
     3061 taaagtctgg aaacgcggaa gtcagcgccc tgcaccatta tgttccggat ctgcatcgca
     3121 ggatgctgct ggctaccctg tggaacacct acatctgtat taacgaagcg ctggcattga
     3181 ccctgagtga tttttctctg gtcccgccgc atccataccg ccagttgttt accctcacaa
     3241 cgttccagta accgggcatg ttcatcatca gtaacccgta tcgtgagcat cctctctcgt
     3301 ttcatcggta tcattacccc catgaacaga aatccccctt acacggaggc atcagtgacc
     3361 aaacaggaaa aaaccgccct taacatggcc cgctttatca gaagccagac attaacgctt
     3421 ctggagaaac tcaacgagct ggacgcggat gaacaggcag acatctgtga atcgcttcac
     3481 gaccacgctg atgagcttta ccgcagctgc ctcgcgcgtt tcggtgatga cggtgaaaac
     3541 ctctgacaca tgcagctccc ggagacggtc acagcttgtc tgtaagcgga tgccgggagc
     3601 agacaagccc gtcagggcgc gtcagcgggt gttggcgggt gtcggggcgc agccatgacc
     3661 cagtcacgta gcgatagcgg agtgtatact ggcttaacta tgcggcatca gagcagattg
     3721 tactgagagt gcaccattgc ggtgtgaaat accgcacaga tgcgtaagga gaaaataccg
     3781 catcaggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg
     3841 gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa
     3901 cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc
     3961 gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc
     4021 aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag
     4081 ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct
     4141 cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta
     4201 ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc
     4261 cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc
     4321 agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt
     4381 gaagtggtgg cctaactacg gctacactag aaggacagta tttggtatct gcgctctgct
     4441 gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc
     4501 tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatttca
     4561 agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta
     4621 agggattttg gtcatgaaca ataaaactgt ctgcttacat aaacagtaat acaaggggtg
     4681 ttatgagcca tattcaacgg gaaacgtctt gctctaggcc gcgattaaat tccaacatgg
     4741 atgctgattt atatgggtat aaatgggctc gcgataatgt cgggcaatca ggtgcgacaa
     4801 tctatcgatt gtatgggaag cccgatgcgc cagagttgtt tctgaaacat ggcaaaggta
     4861 gcgttgccaa tgatgttaca gatgagatgg tcagactaaa ctggctgacg gaatttatgc
     4921 ctcttccgac catcaagcat tttatccgta ctcctgatga tgcatggtta ctcaccactg
     4981 cgatccccgg gaaaacagca ttccaggtat tagaagaata tcctgattca ggtgaaaata
     5041 ttgttgatgc gctggcagtg ttcctgcgcc ggttgcattc gattcctgtt tgtaattgtc
     5101 cttttaacag cgatcgcgta tttcgtctcg ctcaggcgca atcacgaatg aataacggtt
     5161 tggttgatgc gagtgatttt gatgacgagc gtaatggctg gcctgttgaa caagtctgga
     5221 aagaaatgca taaacttttg ccattctcac cggattcagt cgtcactcat ggtgatttct
     5281 cacttgataa ccttattttt gacgagggga aattaatagg ttgtattgat gttggacgag
     5341 tcggaatcgc agaccgatac caggatcttg ccatcctatg gaactgcctc ggtgagtttt
     5401 ctccttcatt acagaaacgg ctttttcaaa aatatggtat tgataatcct gatatgaata
     5461 aattgcagtt tcatttgatg ctcgatgagt ttttctaaga attaattcat gagcggatac
     5521 atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa
     5581 gtgccaccta aattgtaagc gttaatattt tgttaaaatt cgcgttaaat ttttgttaaa
     5641 tcagctcatt ttttaaccaa taggccgaaa tcggcaaaat cccttataaa tcaaaagaat
     5701 agaccgagat agggttgagt gttgttccag tttggaacaa gagtccacta ttaaagaacg
     5761 tggactccaa cgtcaaaggg cgaaaaaccg tctatcaggg cgatggccca ctacgtgaac
     5821 catcacccta atcaagtttt ttggggtcga ggtgccgtaa agcactaaat cggaacccta
     5881 aagg
//
