LOCUS       sequence-151428-        7876 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  expresses the N-terminal fragment of manduca sexta soluble guanylyl 
            cyclase subunits alpha and beta.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7876)
  AUTHORS   Fritz BG, Roberts SA, Ahmed A, Breci L, Li W, Weichsel A, Brailey 
            JL, Wysocki VH, Tama F, Montfort WR
  TITLE     Molecular model of a soluble guanylyl cyclase fragment determined by
            small-angle X-ray scattering and chemical cross-linking.
  JOURNAL   Biochemistry. 2013 Mar 5;52(9):1568-82. doi: 10.1021/bi301570m. Epub
            2013 Feb 15.
  PUBMED    23363317
REFERENCE   2  (bases 1 to 7876)
  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: "10.1021/bi301570m";
            journalName: "Biochemistry"; date: "2013-03-5- 5"; volume: "52"; 
            issue: "9"; pages: "1568-82"
COMMENT     Sequence Label: pETDuet-msGC-NT21
FEATURES             Location/Qualifiers
     source          1..7876
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    3..27
                     /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             42..64
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             83..100
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     promoter        1465..1483
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1484..1508
                     /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)."
     CDS             2822..2866
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     primer_bind     complement(2904..2922)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2918..2965
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      3002..3457
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3089..3108)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3299..3320
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(join(3572..4363,4364..4432))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     4195..4214
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4433..4525)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      4606..5194
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5095..5114
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5348..5365
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    5380..5522
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(5448..5467)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5608..5630
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(5624..5815)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(6387..7469)
                     /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     7431..7450
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(7470..7547)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     7753..7772
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        join(7860..7876,1..2)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 ggggaattgt gagcggataa caattcccct ctagaaataa ttttgtttaa ctttaagaag
       61 gagatatacc atgggcagca gccatcacca tcatcaccac agccaggatc cgaccaaagt
      121 gacagatttg aagatcggcg tggctagctt ctgcaaagcg tttccatggc atttcatcac
      181 tgataaaagg ctggaactcg ttcaattggg agctggattc atgcgcctct tcggaaccca
      241 cttggcaaca cacgggtctt cgctgggcac gtacttcaga ctcctgcgtc caagaggagt
      301 gccgctggac tttcgcgaga tcctcaagcg tgtcaacaca ccgttcatgt tttgcctaaa
      361 gatgcctggt tcgacagcac ttgccgaggg tttggagatc aaaggacaaa tggtgttctg
      421 tgccgagtca gattctttgc tgtttgtcgg gtccccgttc ttggacggtc tcgaaggtct
      481 gactggaagg ggactcttca tatccgatat acctcttcat gatgcgacga gagatgtcat
      541 cttagtcgga gaacaggcta gggctcagga tggtttgcga agacgaatgg acaagctgaa
      601 gaactcaata gaagaagcga gcaaagcagt agacaaggaa cgagagaagt aagtcagcct
      661 gctgcattta atattcccac cccacatcgc caaaaggctt tggctaggcg agaaaatcga
      721 agctaaaagc cacgatgatg tgacaatgct atttagtgat atcgtcggct tcacttccat
      781 ttgtgccact gccacaccca tgatggttat tgctatgctt gaagatctgt atagtgtttt
      841 cgatatattc tgcgaggaac ttgatgttta caaggtggaa acaataggcg atgcgtactg
      901 cgtggctagc gggcttcatc gcaaagtgga gactcacgcg ccgcagattg cgtggatggc
      961 tctgcgcatg gtggaaacat gtgctcagca tctcacgcat gaggggaatc ccattaagat
     1021 gcgcataggc ctgcatacgg gcacagtgtt agccggagtg gttggcaaga cgatgctcaa
     1081 atattgcctc ttcggacaca acgtgacact cgcaaacaag tttgaatctg ggtcggaacc
     1141 gctcaaaatc aacgttagcc caaccacata cgagtggttg attaaattcc ccggattcga
     1201 catggagcca cgcgaccggt cttgtctgcc gaattccttc cccaaggaca tacacggaac
     1261 atgttatttt cttcacaagt acacgcaccc cggaaccgac cccggagagc cgcaggtaaa
     1321 acatatacga gaggccttga aagattacgg cattggacaa gcaaattcaa ctgatgtgga
     1381 cacagaagaa ccaacttagg cggccgcata atgcttaagt cgaacagaaa gtaatcgtat
     1441 tgtacacggc cgcataatcg aaattaatac gactcactat aggggaattg tgagcggata
     1501 acaattcccc atcttagtat attagttaag tataagaagg agatatacat atgtacgggt
     1561 ttgtgaacta tgccctcgag ctccttgtga tgaaaacttt tgacgaagaa acatgggaga
     1621 cgataaagaa aaaggccgac gtggctatgg aaggcagttt cctagtgaga cagatttatg
     1681 aggatgagat cacttataat ttgatcaccg ctgccgttga ggtcttgcaa atacccgctg
     1741 acgctatatt ggagctattt ggaaaaacgt tcttcgagtt ttgccaagat tccgggtatg
     1801 ataaaattct gcaagtactt ggagccacgc cacgagattt tctacagaat ctggacggcc
     1861 tgcacgatca tctcgggaca ttatatcctg gcatgagatc cccctcgttc cgatgcaccg
     1921 agcgtcctga agatggcgcc cttgtgcttc attattactc cgacaggcct ggtctggagc
     1981 atatcgttat tggaattgta aagactgtgg cgagcaaact ccataacacc gaagtgaagg
     2041 tggaaatcct aaagaccaaa gaggaatgtg accacgtcca gttcttgatc actgaaactt
     2101 ccaccaccgg tagggtgtct gcacctgaga tagcagagat tgaaacgttg tctcttgaac
     2161 caaaagtgag tccagcgaca ttctgccgag tgtttccatt ccatttgatg tttgatcgtg
     2221 atctgaacat cgtccaagcg ggacgtactg tgtctcgtct actcccaaga gttaccagac
     2281 caggctgtaa gattactgat gtcttggata ctgtgcgtcc ccacttagag atgacgtttg
     2341 ctaacgtcct ggctcatatt aacacggtct acgttttgaa aaccaaacct gaagagatga
     2401 gtgtcactga tcctcatgag gaaatcgctt ctcttcgctt gaaaggtcaa atgctgtaca
     2461 ttccagaaac agacgtggtt gtattccagt gttatccaag tgtgaccaat ttggacgatc
     2521 tgactcgccg cggcctatgc attgctgaca ttccacttca cgatgcgaca cgagacctgg
     2581 ttctcatgtc cgagcaattt gaagctgatt ataagctgac ccagaatctg gaagttttga
     2641 ctgacaaact tcaacaaaca ttccgagagc tggaattgga aaaacagaag taggacaggc
     2701 ttctttactc agtgctgcct ataagcgtgg ccactgaact tcgtcaccgt cgctgaaagg
     2761 gcgaattctg cagatatcgg ccggccacgc gatcgctgac gtcggtaccc tcgagtctgg
     2821 taaagaaacc gctgctgcga aatttgaacg ccagcacatg gactcgtcta ctagcgcagc
     2881 ttaattaacc taggctgctg ccaccgctga gcaataacta gcataacccc ttggggcctc
     2941 taaacgggtc ttgaggggtt ttttgctgaa aggaggaact atatccggat tggcgaatgg
     3001 gacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc
     3061 gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc ctttctcgcc
     3121 acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt
     3181 agtgctttac ggcacctcga ccccaaaaaa cttgattagg gtgatggttc acgtagtggg
     3241 ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt ctttaatagt
     3301 ggactcttgt tccaaactgg aacaacactc aaccctatct cggtctattc ttttgattta
     3361 taagggattt tgccgatttc ggcctattgg ttaaaaaatg agctgattta acaaaaattt
     3421 aacgcgaatt ttaacaaaat attaacgttt acaatttctg gcggcacgat ggcatgagat
     3481 tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct
     3541 aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta
     3601 tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa
     3661 ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac
     3721 gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa
     3781 gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag
     3841 taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg
     3901 tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag
     3961 ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg
     4021 tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc
     4081 ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat
     4141 tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata
     4201 ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa
     4261 aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca
     4321 actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc
     4381 aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc
     4441 tttttcaatc atgattgaag catttatcag ggttattgtc tcatgagcgg atacatattt
     4501 gaatgtattt agaaaaataa acaaataggt catgaccaaa atcccttaac gtgagttttc
     4561 gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt
     4621 tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt
     4681 gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat
     4741 accaaatact gtccttctag tgtagccgta gttaggccac cacttcaaga actctgtagc
     4801 accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa
     4861 gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg
     4921 ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag
     4981 atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag
     5041 gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa
     5101 cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt
     5161 gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg
     5221 gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc
     5281 tgtggataac cgtattaccg cctttgagtg agctgatacc gctcgccgca gccgaacgac
     5341 cgagcgcagc gagtcagtga gcgaggaagc ggaagagcgc ctgatgcggt attttctcct
     5401 tacgcatctg tgcggtattt cacaccgcat atatggtgca ctctcagtac aatctgctct
     5461 gatgccgcat agttaagcca gtatacactc cgctatcgct acgtgactgg gtcatggctg
     5521 cgccccgaca cccgccaaca cccgctgacg cgccctgacg ggcttgtctg ctcccggcat
     5581 ccgcttacag acaagctgtg accgtctccg ggagctgcat gtgtcagagg ttttcaccgt
     5641 catcaccgaa acgcgcgagg cagctgcggt aaagctcatc agcgtggtcg tgaagcgatt
     5701 cacagatgtc tgcctgttca tccgcgtcca gctcgttgag tttctccaga agcgttaatg
     5761 tctggcttct gataaagcgg gccatgttaa gggcggtttt ttcctgtttg gtcactgatg
     5821 cctccgtgta agggggattt ctgttcatgg gggtaatgat accgatgaaa cgagagagga
     5881 tgctcacgat acgggttact gatgatgaac atgcccggtt actggaacgt tgtgagggta
     5941 aacaactggc ggtatggatg cggcgggacc agagaaaaat cactcagggt caatgccagc
     6001 gcttcgttaa tacagatgta ggtgttccac agggtagcca gcagcatcct gcgatgcaga
     6061 tccggaacat aatggtgcag ggcgctgact tccgcgtttc cagactttac gaaacacgga
     6121 aaccgaagac cattcatgtt gttgctcagg tcgcagacgt tttgcagcag cagtcgcttc
     6181 acgttcgctc gcgtatcggt gattcattct gctaaccagt aaggcaaccc cgccagccta
     6241 gccgggtcct caacgacagg agcacgatca tgctagtcat gccccgcgcc caccggaagg
     6301 agctgactgg gttgaaggct ctcaagggca tcggtcgaga tcccggtgcc taatgagtga
     6361 gctaacttac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt
     6421 gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgcc
     6481 agggtggttt ttcttttcac cagtgagacg ggcaacagct gattgccctt caccgcctgg
     6541 ccctgagaga gttgcagcaa gcggtccacg ctggtttgcc ccagcaggcg aaaatcctgt
     6601 ttgatggtgg ttaacggcgg gatataacat gagctgtctt cggtatcgtc gtatcccact
     6661 accgagatgt ccgcaccaac gcgcagcccg gactcggtaa tggcgcgcat tgcgcccagc
     6721 gccatctgat cgttggcaac cagcatcgca gtgggaacga tgccctcatt cagcatttgc
     6781 atggtttgtt gaaaaccgga catggcactc cagtcgcctt cccgttccgc tatcggctga
     6841 atttgattgc gagtgagata tttatgccag ccagccagac gcagacgcgc cgagacagaa
     6901 cttaatgggc ccgctaacag cgcgatttgc tggtgaccca atgcgaccag atgctccacg
     6961 cccagtcgcg taccgtcttc atgggagaaa ataatactgt tgatgggtgt ctggtcagag
     7021 acatcaagaa ataacgccgg aacattagtg caggcagctt ccacagcaat ggcatcctgg
     7081 tcatccagcg gatagttaat gatcagccca ctgacgcgtt gcgcgagaag attgtgcacc
     7141 gccgctttac aggcttcgac gccgcttcgt tctaccatcg acaccaccac gctggcaccc
     7201 agttgatcgg cgcgagattt aatcgccgcg acaatttgcg acggcgcgtg cagggccaga
     7261 ctggaggtgg caacgccaat cagcaacgac tgtttgcccg ccagttgttg tgccacgcgg
     7321 ttgggaatgt aattcagctc cgccatcgcc gcttccactt tttcccgcgt tttcgcagaa
     7381 acgtggctgg cctggttcac cacgcgggaa acggtctgat aagagacacc ggcatactct
     7441 gcgacatcgt ataacgttac tggtttcaca ttcaccaccc tgaattgact ctcttccggg
     7501 cgctatcatg ccataccgcg aaaggttttg cgccattcga tggtgtccgg gatctcgacg
     7561 ctctccctta tgcgactcct gcattaggaa gcagcccagt agtaggttga ggccgttgag
     7621 caccgccgcc gcaaggaatg gtgcatgcaa ggagatggcg cccaacagtc ccccggccac
     7681 ggggcctgcc accataccca cgccgaaaca agcgctcatg agcccgaagt ggcgagcccg
     7741 atcttcccca tcggtgatgt cggcgatata ggcgccagca accgcacctg tggcgccggt
     7801 gatgccggcc acgatgcgtc cggcgtagag gatcgagatc gatctcgatc ccgcgaaatt
     7861 aatacgactc actata
//
