LOCUS       sequence-213480-        7788 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Protein expression and purification of GNAI2_G-alpha.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7788)
  AUTHORS   Teyra J, Kelil A, Jain S, Helmy M, Jajodia R, Hooda Y, Gu J, D'Cruz 
            AA, Nicholson SE, Min J, Sudol M, Kim PM, Bader GD, Sidhu SS
  TITLE     Large-scale survey and database of high affinity ligands for peptide
            recognition modules.
  JOURNAL   Mol Syst Biol. 2020 Dec;16(12):e9310. doi: 10.15252/msb.20199310.
  PUBMED    33438817
REFERENCE   2  (bases 1 to 7788)
  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: "Mol Syst 
            Biol."; date: "2020-12"; volume: "16(12):e9310. doi"; pages: " 
            10.15252/msb.20199310"
COMMENT     Sequence Label: pHH0103_GNAI2_G-alpha
FEATURES             Location/Qualifiers
     source          1..7788
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(1..6,7..22,23..29)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    37..53
                     /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     42..64
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     CDS             138..155
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             171..824
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     782..804
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             831..854
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             867..884
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     terminator      1983..2030
                     /label=fd terminator
                     /note="central terminator from bacteriophage fd (Otsuka and
                     Kunisawa, 1982)"
     rep_origin      complement(3623..4078)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3760..3781)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3972..3991
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     complement(4120..4142)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4228..4368
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4283..4302
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4383..4400)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4554..5141)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(join(5312..6103,6104..6172))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     5935..5954
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6173..6277)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6345..6363
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        6386..6463
                     /gene="lacI (mutant)"
                     /label=lacIq promoter
                     /note="In the lacIq allele, a single base change in the 
                     promoter boosts expression of the lacI gene about 10-fold."
     CDS             6464..7546
                     /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(6483..6502)
                     /label=LacI-R
                     /note="LacI, reverse primer"
ORIGIN
        1 ttgacaatta atcatcggct cgtataatgt gtggaattgt gagcggataa caatttcaca
       61 caggaaacag ccagtccgtt taggtgtttt cacgagcact tcaccaacaa ggaccataga
      121 ttatgaaaat cgaagaacac catcaccatc accattccag cggtaagctt atgtccccta
      181 tactaggtta ttggaaaatt aagggccttg tgcaacccac tcgacttctt ttggaatatc
      241 ttgaagaaaa atatgaagag catttgtatg agcgcgatga aggtgataaa tggcgaaaca
      301 aaaagtttga attgggtttg gagtttccca atcttcctta ttatattgat ggtgatgtta
      361 aattaacaca gtctatggcc atcatacgtt atatagctga caagcacaac atgttgggtg
      421 gttgtccaaa agagcgtgca gagatttcaa tgcttgaagg agcggttttg gatattagat
      481 acggtgtttc gagaattgca tatagtaaag actttgaaac tctcaaagtt gattttctta
      541 gcaagctacc tgaaatgctg aaaatgttcg aagatcgttt atgtcataaa acatatttaa
      601 atggtgatca tgtaacccat cctgacttca tgttgtatga cgctcttgat gttgttttat
      661 acatggaccc aatgtgcctg gatgcgttcc caaaattagt ttgttttaaa aaacgtattg
      721 aagctatccc acaaattgat aagtacttga aatccagcaa gtatatagca tggcctttgc
      781 agggctggca agccacgttt ggtggtggcg accatcctcc aaaatcggat ctggaagttc
      841 tgttccaggg gcccctgtcc agcggtctgg ttccgcgtgg ttccggtacc gcggcccagc
      901 cggccggttg tactgtttct gccgaagaca aagccgcagc tgaacgttct aagatgattg
      961 ataaaaacct gcgcgaggat ggcgaaaaag cggcacgtga agtcaaactg ctgctgctgg
     1021 gcgcaggcga atccggtaaa tctaccatcg taaaacagat gaaaatcatc cacgaagacg
     1081 gttacagcga agaagagtgc cgtcagtatc gtgcggttgt atattccaac accattcagt
     1141 ccatcatggc tatcgtcaaa gcaatgggta acctgcagat cgattttgct gatcctagcc
     1201 gtgcggatga tgcacgtcag ctgttcgctc tgagctgcac tgcagaggaa cagggcgtac
     1261 tgccggatga tctgtctggt gttattcgtc gtctgtgggc tgaccacggt gtacaagcct
     1321 gctttggtcg cagccgcgaa taccagctga acgatagcgc ggcgtactac ctgaatgatc
     1381 tggaacgcat tgcgcagagc gactacattc caacccagca ggacgtgctg cgtacgcgcg
     1441 tgaagacgac cggcatcgtg gaaactcatt tcactttcaa agacctgcac ttcaaaatgt
     1501 tcgacgtggg tggccaacgt tccgaacgta aaaagtggat tcactgtttt gaaggcgtta
     1561 ccgctatcat cttctgcgtt gcgctgtccg cttatgacct ggttctggcc gaggacgaag
     1621 agatgaatcg catgcatgag tccatgaaac tgtttgactc tatctgcaac aacaaatggt
     1681 tcaccgacac ctccatcatc ctgtttctga acaaaaagga cctgttcgaa gaaaagatta
     1741 cccactctcc gctgactatt tgtttcccgg agtacaccgg cgcgaacaaa tacgacgaag
     1801 cggcttctta catccagtct aaattcgaag acctgaacaa acgtaaagac accaaagaaa
     1861 tctatactca ctttacgtgt gccaccgata ccaaaaacgt tcagttcgtg ttcgatgcag
     1921 tcaccgacgt tatcatcaag aacaacctga aagattgcgg tctgttcgcg gccgcataat
     1981 aaaccgatac aattaaaggc tccttttgga gccttttttt tggagatttt caacgtgaaa
     2041 aaattattat tcgcaattcc tttagttgtt cctttctatt ctcactccgc tgaaactgtt
     2101 gaaagttgtt tagcaaaacc ccatacagaa aattcattta ctaacgtctg gaaagacgac
     2161 aaaactttag atcgttacgc taactatgag ggctgtctgt ggaatgctac aggcgttgta
     2221 gtttgtactg gtgacgaaac tcagtgtcta gctagagtgg cggtggctct ggttccggtg
     2281 attttgatta tgaaaagatg gcaaacgcta ataagggggc tatgaccgaa aatgccgatg
     2341 aaaacgcgct acagtctgac gctaaaggca aacttgattc tgtcgctact gattacggtg
     2401 ctgctatcga tggtttcatt ggtgacgttt ccggccttgc taatggtaat ggtgctactg
     2461 gtgattttgc tggctctaat tcccaaatgg ctcaagtcgg tgacggtgat aattcacctt
     2521 taatgaataa tttccgtcaa tatttacctt ccctccctca atcggttgaa tgtcgccctt
     2581 ttgtctttgg cgctggtaaa ccatatgaat tttctattga ttgtgacaaa ataaacttat
     2641 tccgtggtgt ctttgcgttt cttttatatg ttgccacctt tatgtatgta ttttctacgt
     2701 ttgctaacat actgcgtaat aaggagtctt aatcatgcca gttcttttgg ctagcgccgc
     2761 cctatacctt gtctgcctcc ccgcgttgcg tcgcggtgca tggagccggg ccacctcgac
     2821 ctgaatggaa gccggcggca cctcgctaac ggattcacca ctccaagaat tggagccaat
     2881 caattcttgc ggagaactgt gaatgcgcaa accaaccctt ggcagaacat atccatcgcg
     2941 tccgccatct ccagcagccg cacgcggcgc atctcgggca gcgttgggtc ctggccacgg
     3001 gtgcgcatga tcgtgctcct gtcgttgagg acccggctag gctggcgggg ttgccttact
     3061 ggttagcaga atgaatcacc gatacgcgag cgaacgtgaa gcgactgctg ctgcaaaacg
     3121 tctgcgacct gagcaacaac atgaatggtc ttcggtttcc gtgtttcgta aagtctggaa
     3181 acgcggaagt cagcgccctg caccattatg ttccggatct gcatcgcagg atgctgctgg
     3241 ctaccctgtg gaacacctac atctgtatta acgaagcgct ggcattgacc ctgagtgatt
     3301 tttctctggt cccgccgcat ccataccgcc agttgtttac cctcacaacg ttccagtaac
     3361 cgggcatgtt catcatcagt aacccgtatc gtgagcatcc tctctcgttt catcggtatc
     3421 attaccccca tgaacagaaa tcccccttac acggaggcat cagtgaccaa acaggaaaaa
     3481 accgccctta acatggcccg ctttatcaga agccagacat taacgcttct ggagaaactc
     3541 aacgagctgg acgcggatga acaggcagac atctgtgaat cgcttcacga ccacgctgat
     3601 gagctttacc gcaggatccg cgaaattgta agcgttaata ttttgttaaa attcgcgtta
     3661 aatttttgtt aaatcagctc attttttaac caataggccg aaatcggcaa aatcccttat
     3721 aaatcaaaag aatagaccga gatagggttg agtgttgttc cagtttggaa caagagtcca
     3781 ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc
     3841 ccactacgtg aaccatcacc ctaatcaagt tttttggggt cgaggtgccg taaagcacta
     3901 aatcggaacc ctaaagggag cccccgattt agagcttgac ggggaaagcc ggcgaacgtg
     3961 gcgagaaagg aagggaagaa agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg
     4021 gtcacgctgc gcgtaaccac cacacccgcc gcgcttaatg cgccgctaca gggcgcgtcg
     4081 cggatcctgc ctcgcgcgtt tcggtgatga cggtgaaaac ctctgacaca tgcagctccc
     4141 ggagacggtc acagcttgtc tgtaagcgga tgccgggagc agacaagccc gtcagggcgc
     4201 gtcagcgggt gttggcgggt gtcggggcgc agccatgacc cagtcacgta gcgatagcgg
     4261 agtgtatact ggcttaacta tgcggcatca gagcagattg tactgagagt gcaccatatg
     4321 cggtgtgaaa taccgcacag atgcgtaagg agaaaatacc gcatcaggcg ctcttccgct
     4381 tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac
     4441 tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga
     4501 gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat
     4561 aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac
     4621 ccgacaggac tataaagata ccagggtttc cccctggaag ctccctcgtg cgctctcctg
     4681 ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc
     4741 tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg
     4801 gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc
     4861 ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga
     4921 ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg
     4981 gctacactag aaggacagta tttggtatct gcgctctgct gaagccagtt accttcggaa
     5041 aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg
     5101 tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt
     5161 ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat
     5221 tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct
     5281 aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta
     5341 tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa
     5401 ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac
     5461 gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa
     5521 gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag
     5581 taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctgca ggcatcgtgg
     5641 tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag
     5701 ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg
     5761 tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc
     5821 ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat
     5881 tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaaca cgggataata
     5941 ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa
     6001 aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca
     6061 actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc
     6121 aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc
     6181 tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg
     6241 aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac
     6301 ctgacgtcta agaaaccatt attatcatga cattaaccta taaaaatagg cgtatcacga
     6361 ggccctttcg tcttcaagaa ttcccgacac catcgaatgg tgcaaaacct ttcgcggtat
     6421 ggcatgatag cgcccggaag agagtcaatt cagggtggtg aatgtgaaac cagtaacgtt
     6481 atacgatgtc gcagagtatg ccggtgtctc ttatcagacc gtttcccgcg tggtgaacca
     6541 ggccagccac gtttctgcga aaacgcggga aaaagtggaa gcggcgatgg cggagctgaa
     6601 ttacattccc aaccgcgtgg cacaacaact ggcgggcaaa cagtcgttgc tgattggcgt
     6661 tgccacctcc agtctggccc tgcacgcgcc gtcgcaaatt gtcgcggcga ttaaatctcg
     6721 cgccgatcaa ctgggtgcca gcgtggtggt gtcgatggta gaacgaagcg gcgtcgaagc
     6781 ctgtaaagcg gcggtgcaca atcttctcgc gcaacgcgtc agtgggctga tcattaacta
     6841 tccgctggat gaccaggatg ccattgctgt ggaagctgcc tgcactaatg ttccggcgtt
     6901 atttcttgat gtctctgacc agacacccat caacagtatt attttctccc atgaagacgg
     6961 tacgcgactg ggcgtggagc atctggtcgc attgggtcac cagcaaatcg cgctgttagc
     7021 gggcccatta agttctgtct cggcgcgtct gcgtctggct ggctggcata aatatctcac
     7081 tcgcaatcaa attcagccga tagcggaacg ggaaggcgac tggagtgcca tgtccggttt
     7141 tcaacaaacc atgcaaatgc tgaatgaggg catcgttccc actgcgatgc tggttgccaa
     7201 cgatcagatg gcgctgggcg caatgcgcgc cattaccgag tccgggctgc gcgttggtgc
     7261 ggatatctcg gtagtgggat acgacgatac cgaagacagc tcatgttata tcccgccgtt
     7321 aaccaccatc aaacaggatt ttcgcctgct ggggcaaacc agcgtggacc gcttgctgca
     7381 actctctcag ggccaggcgg tgaagggcaa tcagctgttg cccgtctcac tggtgaaaag
     7441 aaaaaccacc ctggcgccca atacgcaaac cgcctctccc cgcgcgttgg ccgattcatt
     7501 aatgcagctg gcacgacagg tttcccgact ggaaagcggg cagtgagcgc aacgcaatta
     7561 atgtaagtta gctcactcat taggcacaat tctcatgttt gacagcttat catcgactgc
     7621 acggtgcacc aatgcttctg gcgtcaggca gccatcggaa gctgtggtat ggctgtgcag
     7681 gtcgtaaatc actgcataat tcgtgtcgct caaggcgcac tcccgttctg gataatgttt
     7741 tttgcgccga catcataacg gttctggcaa atattctgaa atgagctg
//
