LOCUS       sequence-420494-        5737 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Vector backbone for random polymerase chain reaction screen.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5737)
  TITLE     Pospiech Lab Plasmids 2023
REFERENCE   2  (bases 1 to 5737)
  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: pEGGsH6-MCS_SHORT_inFrame
FEATURES             Location/Qualifiers
     source          1..5737
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(179..184,185..200,201..207)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    215..231
                     /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             254..907
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     865..887
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             914..931
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             968..988
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             join(995..997,998..1000,1001..1513)
                     /codon_start=1
                     /product="N-terminal fragment of mVenus for use in 
                     bimolecular fluorescence complementation (BiFC) (Kodama and
                     Hu, 2010)"
                     /label=VN173
                     /translation="MVSKGEELFTGVVPILVELDGVVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIE"
     primer_bind     complement(1301..1320)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1648..1669
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             1694..1714
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=7xHis
                     /translation="HHHHHHH"
     primer_bind     complement(1783..1805)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(1950..1968)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2036..2140
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2141..2209,2210..3001)
                     /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     complement(2359..2378)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3172..3760
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3661..3680
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3914..3931
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        4004..4081
                     /gene="lacI"
                     /label=lacI promoter
     CDS             4082..5164
                     /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(4101..4120)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(5213..5218,5219..5236,5237..5243)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5251..5267
                     /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     5275..5291
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5275..5291
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             join(5287..5737,1..77)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(5307..5324)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5307..5323)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5316..5338)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
ORIGIN
        1 tatcgactgc acggtgcacc aatgcttctg gcgtcaggca gccatcggaa gctgtggtat
       61 ggctgtgcag gtcgtaaatc actgcataat tcgtgtcgct caaggcgcac tcccgttctg
      121 gataatgttt tttgcgccga catcataacg gttctggcaa atattctgaa atgagctgtt
      181 gacaattaat catcggctcg tataatgtgt ggaattgtga gcggataaca atttcacaca
      241 ggaaacagta ttcatgtccc ctatactagg ttattggaaa attaagggcc ttgtgcaacc
      301 cactcgactt cttttggaat atcttgaaga aaaatatgaa gagcatttgt atgagcgcga
      361 tgaaggtgat aaatggcgaa acaaaaagtt tgaattgggt ttggagtttc ccaatcttcc
      421 ttattatatt gatggtgatg ttaaattaac acagtctatg gccatcatac gttatatagc
      481 tgacaagcac aacatgttgg gtggttgtcc aaaagagcgt gcagagattt caatgcttga
      541 aggagcggtt ttggatatta gatacggtgt ttcgagaatt gcatatagta aagactttga
      601 aactctcaaa gttgattttc ttagcaagct acctgaaatg ctgaaaatgt tcgaagatcg
      661 tttatgtcat aaaacatatt taaatggtga tcatgtaacc catcctgact tcatgttgta
      721 tgacgctctt gatgttgttt tatacatgga cccaatgtgc ctggatgcgt tcccaaaatt
      781 agtttgtttt aaaaaacgta ttgaagctat cccacaaatt gataagtact tgaaatccag
      841 caagtatata gcatggcctt tgcagggctg gcaagccacg tttggtggtg gcgaccatcc
      901 tccaaaatcg gatctggttc cgcgtggatc cgtcgacata tgcgcggccg cagatctgct
      961 agccggcgaa aacctgtatt tccaatcgcg acccatggtg agcaagggcg aggagctgtt
     1021 caccggggtg gtgcccatcc tggtcgagct ggacggcgtc gtaaacggcc acaagttcag
     1081 cgtgtccggc gagggcgagg gcgatgccac ctacggcaag ctgaccctga agttcatctg
     1141 caccaccggc aagctgcccg tgccctggcc caccctcgtg accaccctga cctacggcgt
     1201 gcagtgcttc agccgctacc ccgaccacat gaagcagcac gacttcttca agtccgccat
     1261 gcccgaaggc tacgtccagg agcgcaccat cttcttcaag gacgacggca actacaagac
     1321 ccgcgccgag gtgaagttcg agggcgacac cctggtgaac cgcatcgagc tgaagggcat
     1381 cgacttcaag gaggacggca acatcctggg gcacaagctg gagtacaact acaacagcca
     1441 caacgtctat atcatggccg acaagcagaa gaacggcatc aaggtgaact tcaagatccg
     1501 ccacaacatc gaggacggca gcgtgcagct cgccgaccac taccagcaga acacccccat
     1561 cggcgacggc cccgtgctgc tgcccgacaa ccactacctg agcacccagt ccgccctgag
     1621 caaagacccc aacgagaagc gcgatcacat ggtcctgctg gagttcgtga ccgccgccgg
     1681 gatcaccccc gggcaccatc atcaccacca tcatagttaa ttaagaattc atcgtgactg
     1741 actgacgatc tgcctcgcgc gtttcggtga tgacggtgaa aacctctgac acatgcagct
     1801 cccggagacg gtcacagctt gtctgtaagc ggatgccggg agcagacaag cccgtcaggg
     1861 cgcgtcagcg ggtgttggcg ggtgtcgggg cgcagccatg acccagtcac gtagcgatag
     1921 cggagtgtat aattcttgaa gacgaaaggg cctcgtgata cgcctatttt tataggttaa
     1981 tgtcatgata ataatggttt cttagacgtc aggtggcact tttcggggaa atgtgcgcgg
     2041 aacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca tgagacaata
     2101 accctgataa atgcttcaat aatattgaaa aaggaagagt atgagtattc aacatttccg
     2161 tgtcgccctt attccctttt ttgcggcatt ttgccttcct gtttttgctc acccagaaac
     2221 gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca cgagtgggtt acatcgaact
     2281 ggatctcaac agcggtaaga tccttgagag ttttcgcccc gaagaacgtt ttccaatgat
     2341 gagcactttt aaagttctgc tatgtggcgc ggtattatcc cgtgttgacg ccgggcaaga
     2401 gcaactcggt cgccgcatac actattctca gaatgacttg gttgagtact caccagtcac
     2461 agaaaagcat cttacggatg gcatgacagt aagagaatta tgcagtgctg ccataaccat
     2521 gagtgataac actgcggcca acttacttct gacaacgatc ggaggaccga aggagctaac
     2581 cgcttttttg cacaacatgg gggatcatgt aactcgcctt gatcgttggg aaccggagct
     2641 gaatgaagcc ataccaaacg acgagcgtga caccacgatg cctgcagcaa tggcaacaac
     2701 gttgcgcaaa ctattaactg gcgaactact tactctagct tcccggcaac aattaataga
     2761 ctggatggag gcggataaag ttgcaggacc acttctgcgc tcggcccttc cggctggctg
     2821 gtttattgct gataaatctg gagccggtga gcgtgggtct cgcggtatca ttgcagcact
     2881 ggggccagat ggtaagccct cccgtatcgt agttatctac acgacgggga gtcaggcaac
     2941 tatggatgaa cgaaatagac agatcgctga gataggtgcc tcactgatta agcattggta
     3001 actgtcagac caagtttact catatatact ttagattgat ttaaaacttc atttttaatt
     3061 taaaaggatc taggtgaaga tcctttttga taatctcatg accaaaatcc cttaacgtga
     3121 gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc
     3181 tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt
     3241 ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc
     3301 gcagatacca aatactgtcc ttctagtgta gccgtagtta ggccaccact tcaagaactc
     3361 tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg
     3421 cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg
     3481 gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga
     3541 actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc
     3601 ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg
     3661 gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg
     3721 atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt
     3781 tttacggttc ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc
     3841 tgattctgtg gataaccgta ttaccgcctt tgagtgagct gataccgctc gccgcagccg
     3901 aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa gagcgcctga tgcggtattt
     3961 tctccttacg catctgtgcg gtatttcaca ccgcataaat tccgacacca tcgaatggcg
     4021 caaaaccttt cgcggtatgg catgatagcg cccggaagag agtcaattca gggtggtgaa
     4081 tgtgaaacca gtaacgttat acgatgtcgc agagtatgcc ggtgtctctt atcagaccgt
     4141 ttcccgcgtg gtgaaccagg ccagccacgt ttctgcgaaa acgcgggaaa aagtggaagc
     4201 ggcgatggcg gagctgaatt acattcccaa ccgcgtggca caacaactgg cgggcaaaca
     4261 gtcgttgctg attggcgttg ccacctccag tctggccctg cacgcgccgt cgcaaattgt
     4321 cgcggcgatt aaatctcgcg ccgatcaact gggtgccagc gtggtggtgt cgatggtaga
     4381 acgaagcggc gtcgaagcct gtaaagcggc ggtgcacaat cttctcgcgc aacgcgtcag
     4441 tgggctgatc attaactatc cgctggatga ccaggatgcc attgctgtgg aagctgcctg
     4501 cactaatgtt ccggcgttat ttcttgatgt ctctgaccag acacccatca acagtattat
     4561 tttctcccat gaagacggta cgcgactggg cgtggagcat ctggtcgcat tgggtcacca
     4621 gcaaatcgcg ctgttagcgg gcccattaag ttctgtctcg gcgcgtctgc gtctggctgg
     4681 ctggcataaa tatctcactc gcaatcaaat tcagccgata gcggaacggg aaggcgactg
     4741 gagtgccatg tccggttttc aacaaaccat gcaaatgctg aatgagggca tcgttcccac
     4801 tgcgatgctg gttgccaacg atcagatggc gctgggcgca atgcgcgcca ttaccgagtc
     4861 cgggctgcgc gttggtgcgg atatctcggt agtgggatac gacgataccg aagacagctc
     4921 atgttatatc ccgccgttaa ccaccatcaa acaggatttt cgcctgctgg ggcaaaccag
     4981 cgtggaccgc ttgctgcaac tctctcaggg ccaggcggtg aagggcaatc agctgttgcc
     5041 cgtctcactg gtgaaaagaa aaaccaccct ggcgcccaat acgcaaaccg cctctccccg
     5101 cgcgttggcc gattcattaa tgcagctggc acgacaggtt tcccgactgg aaagcgggca
     5161 gtgagcgcaa cgcaattaat gtaagttagc tcactcatta ggcaccccag gctttacact
     5221 ttatgcttcc ggctcgtatg ttgtgtggaa ttgtgagcgg ataacaattt cacacaggaa
     5281 acagctatga ccatgattac ggattcactg gccgtcgttt tacaacgtcg tgactgggaa
     5341 aaccctggcg ttacccaact taatcgcctt gcagcacatc cccctttcgc cagctggcgt
     5401 aatagcgaag aggcccgcac cgatcgccct tcccaacagt tgcgcagcct gaatggcgaa
     5461 tggcgctttg cctggtttcc ggcaccagaa gcggtgccgg aaagctggct ggagtgcgat
     5521 cttcctgagg ccgatactgt cgtcgtcccc tcaaactggc agatgcacgg ttacgatgcg
     5581 cccatctaca ccaacgtgac ctatcccatt acggtcaatc cgccgtttgt tcccacggag
     5641 aatccgacgg gttgttactc gctcacattt aatgttgatg aaagctggct acaggaaggc
     5701 cagacgcgaa ttatttttga tggcgttgga attagct
//
