LOCUS       sequence-456249-        6065 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Recombinant expression of UNK Swap with dual-affinity tag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6065)
  TITLE     Dissecting RNA Selectivity Mediated by Tandem RNA-Binding Domains
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6065)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: UNK_Swap_pGEX
FEATURES             Location/Qualifiers
     source          1..6065
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(754..867)
                     /codon_start=1
                     /product="streptavidin-binding peptide"
                     /label=SBP
                     /note="selected from a peptide library; binds streptavidin 
                     with nanomolar affinity (Keefe et al., 2001)"
                     /translation="MDEKTTGWRGGHVVEGLAGELEQLRARLEHHPQGQREP"
     CDS             complement(877..900)
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             complement(907..1560)
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     complement(927..949)
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     protein_bind    1583..1599
                     /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(join(1607..1613,1614..1629,1630..1635))
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     CDS             complement(1737..2264)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     2213..2235
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     2227..2244
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     2228..2244
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2260..2276)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2260..2276)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     protein_bind    2284..2300
                     /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(join(2308..2314,2315..2332,2333..2338))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     CDS             complement(2387..3469)
                     /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     3431..3450
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(3470..3547)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     complement(3620..3637)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3791..4379)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3871..3890)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4550..5341,5342..5410))
                     /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     5173..5192
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5411..5515)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5583..5601
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     5746..5768
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
ORIGIN
        1 gcgctcagtg tcccctgtgg ttctgtgcag gaatgggcac tcgtcgccct ccgggcagag
       61 gcctgtagcc tcgtcgtact tggtgcagta gacgtcaggg ctgtaattga aggtgccgtc
      121 ccgacggcgg atggaccggc ggcgccgctg gttcacgaag tgccagtgga agcaggtgta
      181 gggccgatgc tgcgtgcatt tgtgttgcac aaagagtggg cactgctctg tgcggaattc
      241 tttcaggtac gtgtagtgct gcggctcctc gctgaggatc ttttctatca tggcatggct
      301 cgcagcccca gccgactggc cctctatgct cccctctacc gtggtctggc cggacacagc
      361 tgaggaaggc tgcaggtcgt cactcagggg tggctgttct acgtgggcaa aggcgcagaa
      421 gggtcctcgg ggacagctgc ccgactgctg catgtcgttg cacttggtgg acttgtagat
      481 ctcggggtgg aactgctgct cggtgcgggt gtggcagtac tggcaggcgt ctccgttctc
      541 acacttgcca gggtctcccc actcatcccc gtgcttgacg tttggacatg gagacgacct
      601 gtatttgtgc ttccgggggc tccgccgccg gtccttgctg ttgtggtagt agggacaggc
      661 atagccttgg cggcacagcc gcgggggctt cttgcaaggc tccgtcttat agttccccag
      721 cacataagca gtctcttgcc accgagggga tcccggttca cgctgaccct gcgggtggtg
      781 ttccagacga gcacgcagct gttccagttc accagccaga ccttcaacaa cgtgaccacc
      841 acgccaaccg gtggtttttt cgtccattaa tcccaggggc ccctggaaca gaacttccag
      901 atccgatttt ggaggatggt cgccaccacc aaacgtggct tgccagccct gcaaaggcca
      961 tgctatatac ttgctggatt tcaagtactt atcaatttgt gggatagctt caatacgttt
     1021 tttaaaacaa actaattttg ggaacgcatc caggcacatt gggtccatgt ataaaacaac
     1081 atcaagagcg tcatacaaca tgaagtcagg atgggttaca tgatcaccat ttaaatatgt
     1141 tttatgacat aaacgatctt cgaacatttt cagcatttca ggtagcttgc taagaaaatc
     1201 aactttgaga gtttcaaagt ctttactata tgcaattctc gaaacaccgt atctaatatc
     1261 caaaaccgct ccttcaagca ttgaaatctc tgcacgctct tttggacaac cacccaacat
     1321 gttgtgcttg tcagctatat aacgtatgat ggccatagac tgtgttaatt taacatcacc
     1381 atcaatataa taaggaagat tgggaaactc caaacccaat tcaaactttt tgtttcgcca
     1441 tttatcacct tcatcgcgct catacaaatg ctcttcatat ttttcttcaa gatattccaa
     1501 aagaagtcga gtgggttgca caaggccctt aattttccaa taacctagta taggggacat
     1561 gaatactgtt tcctgtgtga aattgttatc cgctcacaat tccacacatt atacgagccg
     1621 atgattaatt gtcaacagct catttcagaa tatttgccag aaccgttatg atgtcggcgc
     1681 aaaaaacatt atccagaacg ggagtgcgcc ttgagcgaca cgaattatgc agtgatttac
     1741 gacctgcaca gccataccac agcttccgat ggctgcctga cgccagaagc attggtgcac
     1801 cgtgcagtcg ataagctaat tccaacgcca tcaaaaataa ttcgcgtctg gccttcctgt
     1861 agccagcttt catcaacatt aaatgtgagc gagtaacaac ccgtcggatt ctccgtggga
     1921 acaaacggcg gattgaccgt aatgggatag gtcacgttgg tgtagatggg cgcatcgtaa
     1981 ccgtgcatct gccagtttga ggggacgacg acagtatcgg cctcaggaag atcgcactcc
     2041 agccagcttt ccggcaccgc ttctggtgcc ggaaaccagg caaagcgcca ttcgccattc
     2101 aggctgcgca actgttggga agggcgatcg gtgcgggcct cttcgctatt acgccagctg
     2161 gcgaaagggg gatgtgctgc aaggcgatta agttgggtaa cgccagggtt ttcccagtca
     2221 cgacgttgta aaacgacggc cagtgaatcc gtaatcatgg tcatagctgt ttcctgtgtg
     2281 aaattgttat ccgctcacaa ttccacacaa catacgagcc ggaagcataa agtgtaaagc
     2341 ctggggtgcc taatgagtga gctaacttac attaattgcg ttgcgctcac tgcccgcttt
     2401 ccagtcggga aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg
     2461 cggtttgcgt attgggcgct agcgtggttt ttcttttcac cagtgagacg ggcaacagct
     2521 gattgccctt caccgcctgg ccctgagaga gttgcagcaa gcggtccacg ctggtttgcc
     2581 ccagcaggcg aaaatcctgt ttgatggtgg ttaacggcgg gatataacat gagctgtctt
     2641 cggtatcgtc gtatcccact accgagatat ccgcaccaac gcgcagcccg gactcggtaa
     2701 tggcgcgcat tgcgcccagc gccatctgat cgttggcaac cagcatcgca gtgggaacga
     2761 tgccctcatt cagcatttgc atggtttgtt gaaaaccgga catggcactc cagtcgcctt
     2821 cccgttccgc tatcggctga atttgattgc gagtgagata tttatgccag ccagccagac
     2881 gcagacgcgc cgagacagaa cttaatgggc ccgctaacag cgcgatttgc tggtgaccca
     2941 atgcgaccag atgctccacg cccagtcgcg taccgtcttc atgggagaaa ataatactgt
     3001 tgatgggtgt ctggtcagag acatcaagaa ataacgccgg aacattagtg caggcagctt
     3061 ccacagcaat ggcatcctgg tcatccagcg gatagttaat gatcagccca ctgacgcgtt
     3121 gcgcgagaag attgtgcacc gccgctttac aggcttcgac gccgcttcgt tctaccatcg
     3181 acaccaccac gctggcaccc agttgatcgg cgcgagattt aatcgccgcg acaatttgcg
     3241 acggcgcgtg cagggccaga ctggaggtgg caacgccaat cagcaacgac tgtttgcccg
     3301 ccagttgttg tgccacgcgg ttgggaatgt aattcagctc cgccatcgcc gcttccactt
     3361 tttcccgcgt tttcgcagaa acgtggctgg cctggttcac cacgcgggaa acggtctgat
     3421 aagagacacc ggcatactct gcgacatcgt ataacgttac tggtttcaca ttcaccaccc
     3481 tgaattgact ctcttccggg cgctatcatg ccataccgcg aaaggttttg cgccattcga
     3541 tggtgtcgga atttatgcgg tgtgaaatac cgcacagatg cgtaaggaga aaataccgca
     3601 tcaggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc
     3661 gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg
     3721 caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt
     3781 tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa
     3841 gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct
     3901 ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc
     3961 cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg
     4021 tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct
     4081 tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag
     4141 cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga
     4201 agtggtggcc taactacggc tacactagaa ggacagtatt tggtatctgc gctctgctga
     4261 agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg
     4321 gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag
     4381 aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag
     4441 ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat
     4501 gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct
     4561 taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac
     4621 tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa
     4681 tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg
     4741 gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt
     4801 gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca
     4861 ttgctgcagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt
     4921 cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct
     4981 tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg
     5041 cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg
     5101 agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg
     5161 cgtcaacacg ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa
     5221 aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt
     5281 aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt
     5341 gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt
     5401 gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca
     5461 tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat
     5521 ttccccgaaa agtgccacct gacgtctaag aaaccattat tatcatgaca ttaacctata
     5581 aaaataggcg tatcacgagg ccctttcgtc ttcaagaatt atacactccg ctatcgctac
     5641 gtgactgggt catggctgcg ccccgacacc cgccaacacc cgctgacgcg ccctgacggg
     5701 cttgtctgct cccggcatcc gcttacagac aagctgtgac cgtctccggg agctgcatgt
     5761 gtcagaggtt ttcaccgtca tcaccgaaac gcgcgaggca gatcgtcagt cagtcacgat
     5821 gcggccgctc agctcacagg caccgagtct ccggcggcag atggcatgta caggacagga
     5881 cccggctggg tggggctatt ctgcaaggcc tccatggcct gaagctccct gatgtcgtag
     5941 acaggggagc ggaggtcatg gggcccgtgg gcaaaagcgc agtgcaggcc gtttttggtg
     6001 cagttgcctt tcgagtctgt ctcgtggatg cagattccag ttttgtagta acgaaggtgg
     6061 tacct
//
