LOCUS       sequence-469465-        6749 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression for structure determination. May not contain 
            entire coding region of gene..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6749)
  TITLE     Arrowsmith Lab SGC Plasmids
REFERENCE   2  (bases 1 to 6749)
  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: SARS2_HisSUMOTEV-nsp7/nsp8
FEATURES             Location/Qualifiers
     source          1..6749
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      complement(26..73)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     69..87
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(140..157)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     protein_bind    840..864
                     /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)."
     promoter        complement(865..883)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             complement(1192..1212)
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             complement(1525..1542)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     RBS             1553..1575
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    1590..1614
                     /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)."
     promoter        complement(1615..1633)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1717..1736)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        1942..2019
                     /gene="lacI"
                     /label=lacI promoter
     CDS             2020..3102
                     /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(2039..2058)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             3911..4102
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(4096..4118)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4204..4346
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4259..4278
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4361..4378)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4532..5120)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4612..4631)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5291..6082,6083..6151))
                     /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     5914..5933
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6152..6256)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(6283..6738)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6420..6441)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6632..6651
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 atccggatat agttcctcct ttcagcaaaa aacccctcaa gacccgttta gaggccccaa
       61 ggggttatgc tagttattgc tcagcggtgg cagcagccaa ctcagcttcc tttcgggctt
      121 tgttagcagc cggatctcag tggtggtggt ggtggtgctc gagtgcggcc gcaagcttgt
      181 cgacggagct cgaattctta ctgcagttta accgcagagt tagcacgcag cgcggtaacg
      241 atcaacggcc aagccaggtt cggagagttg tccatagaga tttcagacag ctgaacgatt
      301 ttagagtccg cgtcaacaac ctgctggatt tcccacagcg cagacgcgta ggtgaaggtg
      361 gtaccgtcgc aggtgttttt gtaggtgttg tagtccggga taacaaccat cagtttagca
      421 gcggtggtca gcgggatgat gttcagcgga acgcaaccgt cacgcgcgtt gttgatgatg
      481 ttgttcagcg cgtcgttgtc cagtttacgc agcatggtga acagcatggt ctgcatagca
      541 gaggtaactt tagcacgttt atcttcagaa cgcgcctgtt tgtacatctg ggtcatcgcc
      601 tggtccgcca ttttttccag tttgcgctgc atagccgcat cacggtcgaa ttcagattta
      661 gcaacgttca gagatttttt cagttttttc agaacaactt cagagtcacc gttagcaaca
      721 gcctgttcat aagcttcctg cgcggtagcg aaagcagcat aagacggcag agaagagaat
      781 tcagaagcga tagccatatg tatatctcct tcttatactt aactaatata ctaagatggg
      841 gaattgttat ccgctcacaa ttcccctata gtgagtcgta ttaatttcga ttatgcggcc
      901 gtgtacaata cgattacttt ctgttcgact taagcattac tgcagggtag cacggttgtc
      961 cagcatttct tcgcacagtt tgttgatgtc aacagcaccc tgcatagaca gcagaacaga
     1021 cagcagagaa accatttttt cgaacgcttc ggtggtgtct ttagccagca ggatgtcgtt
     1081 gtgcagctga acgcactgag cccacagttt agaagaagat tcaacacgca gctgctgcag
     1141 aacagacagc agaacaacag aggtgcattt aacgtcagac attttagaca tcccctgaaa
     1201 gtataagttc tcgctggtgc ccccgatttg ttcgcggtgt gcctcgataa tgtcattgtc
     1261 ctccatatcc agatcttcgg gagcctgatc tgcctggatg cgaataccat cataaagaaa
     1321 acgcagacta tccatctctt tcccttggcg cttggcaaac gcctccataa gacgacgaag
     1381 cggtgttgtc ttcttaattt taaagaagat ttcggacgac ccgtccgaca ctttcaagtt
     1441 aatatgtgtt tcgggtttaa cctctggctt cacctccggt ttcgcctcct gattgacttc
     1501 agagtcctga agacttccag atgagtgatg gtgatgatgg tgcatatgta tatctccttc
     1561 ttaaagttaa acaaaattat ttctagaggg gaattgttat ccgctcacaa ttcccctata
     1621 gtgagtcgta ttaatttcgc gggatcgaga tctcgatcct ctacgccgga cgcatcgtgg
     1681 ccggcatcac cggcgccaca ggtgcggttg ctggcgccta tatcgccgac atcaccgatg
     1741 gggaagatcg ggctcgccac ttcgggctca tgagcgcttg tttcggcgtg ggtatggtgg
     1801 caggccccgt ggccggggga ctgttgggcg ccatctcctt gcatgcacca ttccttgcgg
     1861 cggcggtgct caacggcctc aacctactac tgggctgctt cctaatgcag gagtcgcata
     1921 agggagagcg tcgagatccc ggacaccatc gaatggcgca aaacctttcg cggtatggca
     1981 tgatagcgcc cggaagagag tcaattcagg gtggtgaatg tgaaaccagt aacgttatac
     2041 gatgtcgcag agtatgccgg tgtctcttat cagaccgttt cccgcgtggt gaaccaggcc
     2101 agccacgttt ctgcgaaaac gcgggaaaaa gtggaagcgg cgatggcgga gctgaattac
     2161 attcccaacc gcgtggcaca acaactggcg ggcaaacagt cgttgctgat tggcgttgcc
     2221 acctccagtc tggccctgca cgcgccgtcg caaattgtcg cggcgattaa atctcgcgcc
     2281 gatcaactgg gtgccagcgt ggtggtgtcg atggtagaac gaagcggcgt cgaagcctgt
     2341 aaagcggcgg tgcacaatct tctcgcgcaa cgcgtcagtg ggctgatcat taactatccg
     2401 ctggatgacc aggatgccat tgctgtggaa gctgcctgca ctaatgttcc ggcgttattt
     2461 cttgatgtct ctgaccagac acccatcaac agtattattt tctcccatga agacggtacg
     2521 cgactgggcg tggagcatct ggtcgcattg ggtcaccagc aaatcgcgct gttagcgggc
     2581 ccattaagtt ctgtctcggc gcgtctgcgt ctggctggct ggcataaata tctcactcgc
     2641 aatcaaattc agccgatagc ggaacgggaa ggcgactgga gtgccatgtc cggttttcaa
     2701 caaaccatgc aaatgctgaa tgagggcatc gttcccactg cgatgctggt tgccaacgat
     2761 cagatggcgc tgggcgcaat gcgcgccatt accgagtccg ggctgcgcgt tggtgcggat
     2821 atctcggtag tgggatacga cgataccgaa gacagctcat gttatatccc gccgttaacc
     2881 accatcaaac aggattttcg cctgctgggg caaaccagcg tggaccgctt gctgcaactc
     2941 tctcagggcc aggcggtgaa gggcaatcag ctgttgcccg tctcactggt gaaaagaaaa
     3001 accaccctgg cgcccaatac gcaaaccgcc tctccccgcg cgttggccga ttcattaatg
     3061 cagctggcac gacaggtttc ccgactggaa agcgggcagt gagcgcaacg caattaatgt
     3121 aagttagctc actcattagg caccgggatc tcgaccgatg cccttgagag ccttcaaccc
     3181 agtcagctcc ttccggtggg cgcggggcat gactatcgtc gccgcactta tgactgtctt
     3241 ctttatcatg caactcgtag gacaggtgcc ggcagcgctc tgggtcattt tcggcgagga
     3301 ccgctttcgc tggagcgcga cgatgatcgg cctgtcgctt gcggtattcg gaatcttgca
     3361 cgccctcgct caagccttcg tcactggtcc cgccaccaaa cgtttcggcg agaagcaggc
     3421 cattatcgcc ggcatggcgg ccccacgggt gcgcatgatc gtgctcctgt cgttgaggac
     3481 ccggctaggc tggcggggtt gccttactgg ttagcagaat gaatcaccga tacgcgagcg
     3541 aacgtgaagc gactgctgct gcaaaacgtc tgcgacctga gcaacaacat gaatggtctt
     3601 cggtttccgt gtttcgtaaa gtctggaaac gcggaagtca gcgccctgca ccattatgtt
     3661 ccggatctgc atcgcaggat gctgctggct accctgtgga acacctacat ctgtattaac
     3721 gaagcgctgg cattgaccct gagtgatttt tctctggtcc cgccgcatcc ataccgccag
     3781 ttgtttaccc tcacaacgtt ccagtaaccg ggcatgttca tcatcagtaa cccgtatcgt
     3841 gagcatcctc tctcgtttca tcggtatcat tacccccatg aacagaaatc ccccttacac
     3901 ggaggcatca gtgaccaaac aggaaaaaac cgcccttaac atggcccgct ttatcagaag
     3961 ccagacatta acgcttctgg agaaactcaa cgagctggac gcggatgaac aggcagacat
     4021 ctgtgaatcg cttcacgacc acgctgatga gctttaccgc agctgcctcg cgcgtttcgg
     4081 tgatgacggt gaaaacctct gacacatgca gctcccggag acggtcacag cttgtctgta
     4141 agcggatgcc gggagcagac aagcccgtca gggcgcgtca gcgggtgttg gcgggtgtcg
     4201 gggcgcagcc atgacccagt cacgtagcga tagcggagtg tatactggct taactatgcg
     4261 gcatcagagc agattgtact gagagtgcac catatatgcg gtgtgaaata ccgcacagat
     4321 gcgtaaggag aaaataccgc atcaggcgct cttccgcttc ctcgctcact gactcgctgc
     4381 gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat
     4441 ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca
     4501 ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc
     4561 atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc
     4621 aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg
     4681 gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta
     4741 ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg
     4801 ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac
     4861 acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag
     4921 gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga aggacagtat
     4981 ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat
     5041 ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc
     5101 gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt
     5161 ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct
     5221 agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt
     5281 ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc
     5341 gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac
     5401 catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat
     5461 cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg
     5521 cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata
     5581 gtttgcgcaa cgttgttgcc attgctgcag gcatcgtggt gtcacgctcg tcgtttggta
     5641 tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt
     5701 gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag
     5761 tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa
     5821 gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc
     5881 gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt
     5941 taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc
     6001 tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta
     6061 ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa
     6121 taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca
     6181 tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac
     6241 aaataggggt tccgcgcaca tttccccgaa aagtgccacc tgaaattgta aacgttaata
     6301 ttttgttaaa attcgcgtta aatttttgtt aaatcagctc attttttaac caataggccg
     6361 aaatcggcaa aatcccttat aaatcaaaag aatagaccga gatagggttg agtgttgttc
     6421 cagtttggaa caagagtcca ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa
     6481 ccgtctatca gggcgatggc ccactacgtg aaccatcacc ctaatcaagt tttttggggt
     6541 cgaggtgccg taaagcacta aatcggaacc ctaaagggag cccccgattt agagcttgac
     6601 ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa agcgaaagga gcgggcgcta
     6661 gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac cacacccgcc gcgcttaatg
     6721 cgccgctaca gggcgcgtcc cattcgcca
//
