LOCUS       sequence-430164-        5985 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Encodes the enhanced green fluorescent protein.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5985)
  TITLE     Shanbhag lab plasmids
REFERENCE   2  (bases 1 to 5985)
  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: WT-eGFP
FEATURES             Location/Qualifiers
     source          1..5985
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     429..448
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(468..545)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     751..770
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     854..873
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        854..872
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    873..897
                     /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             912..934
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             947..964
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1703..1720
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1773..1791)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1787..1834
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1871..2326
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1958..1977)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2168..2189
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(2418..3233)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(2418..3233)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     3141..3160
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     3225..3244
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      3355..3943
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3844..3863
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4097..4114
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4129..4268
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4194..4213)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4354..4376
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4370..4561)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(join(5370..5985,1..467))
                     /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"
ORIGIN
        1 atgggtgtct ggtcagagac atcaagaaat aacgccggaa cattagtgca ggcagcttcc
       61 acagcaatgg catcctggtc atccagcgga tagttaatga tcagcccact gacgcgttgc
      121 gcgagaagat tgtgcaccgc cgctttacag gcttcgacgc cgcttcgttc taccatcgac
      181 accaccacgc tggcacccag ttgatcggcg cgagatttaa tcgccgcgac aatttgcgac
      241 ggcgcgtgca gggccagact ggaggtggca acgccaatca gcaacgactg tttgcccgcc
      301 agttgttgtg ccacgcggtt gggaatgtaa ttcagctccg ccatcgccgc ttccactttt
      361 tcccgcgttt tcgcagaaac gtggctggcc tggttcacca cgcgggaaac ggtctgataa
      421 gagacaccgg catactctgc gacatcgtat aacgttactg gtttcacatt caccaccctg
      481 aattgactct cttccgggcg ctatcatgcc ataccgcgaa aggttttgcg ccattcgatg
      541 gtgtccggga tctcgacgct ctcccttatg cgactcctgc attaggaagc agcccagtag
      601 taggttgagg ccgttgagca ccgccgccgc aaggaatggt gcatgcaagg agatggcgcc
      661 caacagtccc ccggccacgg ggcctgccac catacccacg ccgaaacaag cgctcatgag
      721 cccgaagtgg cgagcccgat cttccccatc ggtgatgtcg gcgatatagg cgccagcaac
      781 cgcacctgtg gcgccggtga tgccggccac gatgcgtccg gcgtagagga tcgagatctc
      841 gatcccgcga aattaatacg actcactata ggggaattgt gagcggataa caattcccct
      901 ctagaaataa ttttgtttaa ctttaagaag gagatatacc atgggccatc accatcacca
      961 tcacaccgat ccgatggtct ctaaaggtga agaactgttt acgggcgtgg ttccgattct
     1021 ggttgaactg gatggtgacg tcaatggcca taaattcagc gtttctggcg aaggtgaagg
     1081 cgacgcaacc tatggtaaac tgacgctgaa atttatctgc accaccggta aactgccggt
     1141 gccgtggccg accctggtta ccaccctggc gtatggcgtg ctgtgttttt cccgctaccc
     1201 ggatcacatg aaacagcacg actttttcaa aagcgctatg ccggaaggtt acgttcaaga
     1261 acgtaccatt ttctttaaag atgacggcaa ctacaaaacc cgcgccgaag tcaaatttga
     1321 aggtgatacg ctggtgaacc gtattgaact gaaaggcatc gatttcaaag aagacggtaa
     1381 tatcctgggc cataaactgg aatataacta caatagccac aacgtgtata ttatggcgga
     1441 taaacagaaa aacggtatca aatctaactt caaaatccgt cataatatcg aagatggcag
     1501 tgttcagctg gccgaccact atcagcaaaa caccccgatt ggtgatggcc cggtcctgct
     1561 gccggacaat cattacctga gtacgcagtc caaactgtca aaagatccga acgaaaaacg
     1621 cgaccacatg gtcctgctgg aatttgtgac cgcggccggt atcacgctgg gcatggatga
     1681 actgtacaaa taataactcg agcaccacca ccaccaccac tgagatccgg ctgctaacaa
     1741 agcccgaaag gaagctgagt tggctgctgc caccgctgag caataactag cataacccct
     1801 tggggcctct aaacgggtct tgaggggttt tttgctgaaa ggaggaacta tatccggatt
     1861 ggcgaatggg acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc
     1921 agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc
     1981 tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg
     2041 ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgattaggg tgatggttca
     2101 cgtagtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc
     2161 tttaatagtg gactcttgtt ccaaactgga acaacactca accctatctc ggtctattct
     2221 tttgatttat aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa
     2281 caaaaattta acgcgaattt taacaaaata ttaacgctta caatttaggt ggcacttttc
     2341 ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc
     2401 cgctcatgaa ttaattctta gaaaaactca tcgagcatca aatgaaactg caatttattc
     2461 atatcaggat tatcaatacc atatttttga aaaagccgtt tctgtaatga aggagaaaac
     2521 tcaccgaggc agttccatag gatggcaaga tcctggtatc ggtctgcgat tccgactcgt
     2581 ccaacatcaa tacaacctat taatttcccc tcgtcaaaaa taaggttatc aagtgagaaa
     2641 tcaccatgag tgacgactga atccggtgag aatggcaaaa gtttatgcat ttctttccag
     2701 acttgttcaa caggccagcc attacgctcg tcatcaaaat cactcgcatc aaccaaaccg
     2761 ttattcattc gtgattgcgc ctgagcgaga cgaaatacgc gatcgctgtt aaaaggacaa
     2821 ttacaaacag gaatcgaatg caaccggcgc aggaacactg ccagcgcatc aacaatattt
     2881 tcacctgaat caggatattc ttctaatacc tggaatgctg ttttcccggg gatcgcagtg
     2941 gtgagtaacc atgcatcatc aggagtacgg ataaaatgct tgatggtcgg aagaggcata
     3001 aattccgtca gccagtttag tctgaccatc tcatctgtaa catcattggc aacgctacct
     3061 ttgccatgtt tcagaaacaa ctctggcgca tcgggcttcc catacaatcg atagattgtc
     3121 gcacctgatt gcccgacatt atcgcgagcc catttatacc catataaatc agcatccatg
     3181 ttggaattta atcgcggcct agagcaagac gtttcccgtt gaatatggct cataacaccc
     3241 cttgtattac tgtttatgta agcagacagt tttattgttc atgaccaaaa tcccttaacg
     3301 tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga
     3361 tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt
     3421 ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag
     3481 agcgcagata ccaaatactg tccttctagt gtagccgtag ttaggccacc acttcaagaa
     3541 ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag
     3601 tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca
     3661 gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac
     3721 cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa
     3781 ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc
     3841 agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg
     3901 tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc
     3961 ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc
     4021 ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg ctcgccgcag
     4081 ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc tgatgcggta
     4141 ttttctcctt acgcatctgt gcggtatttc acaccgcaat ggtgcactct cagtacaatc
     4201 tgctctgatg ccgcatagtt aagccagtat acactccgct atcgctacgt gactgggtca
     4261 tggctgcgcc ccgacacccg ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc
     4321 cggcatccgc ttacagacaa gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt
     4381 caccgtcatc accgaaacgc gcgaggcagc tgcggtaaag ctcatcagcg tggtcgtgaa
     4441 gcgattcaca gatgtctgcc tgttcatccg cgtccagctc gttgagtttc tccagaagcg
     4501 ttaatgtctg gcttctgata aagcgggcca tgttaagggc ggttttttcc tgtttggtca
     4561 ctgatgcctc cgtgtaaggg ggatttctgt tcatgggggt aatgataccg atgaaacgag
     4621 agaggatgct cacgatacgg gttactgatg atgaacatgc ccggttactg gaacgttgtg
     4681 agggtaaaca actggcggta tggatgcggc gggaccagag aaaaatcact cagggtcaat
     4741 gccagcgctt cgttaataca gatgtaggtg ttccacaggg tagccagcag catcctgcga
     4801 tgcagatccg gaacataatg gtgcagggcg ctgacttccg cgtttccaga ctttacgaaa
     4861 cacggaaacc gaagaccatt catgttgttg ctcaggtcgc agacgttttg cagcagcagt
     4921 cgcttcacgt tcgctcgcgt atcggtgatt cattctgcta accagtaagg caaccccgcc
     4981 agcctagccg ggtcctcaac gacaggagca cgatcatgcg cacccgtggg gccgccatgc
     5041 cggcgataat ggcctgcttc tcgccgaaac gtttggtggc gggaccagtg acgaaggctt
     5101 gagcgagggc gtgcaagatt ccgaataccg caagcgacag gccgatcatc gtcgcgctcc
     5161 agcgaaagcg gtcctcgccg aaaatgaccc agagcgctgc cggcacctgt cctacgagtt
     5221 gcatgataaa gaagacagtc ataagtgcgg cgacgatagt catgccccgc gcccaccgga
     5281 aggagctgac tgggttgaag gctctcaagg gcatcggtcg agatcccggt gcctaatgag
     5341 tgagctaact tacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt
     5401 cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc
     5461 gccagggtgg tttttctttt caccagtgag acgggcaaca gctgattgcc cttcaccgcc
     5521 tggccctgag agagttgcag caagcggtcc acgctggttt gccccagcag gcgaaaatcc
     5581 tgtttgatgg tggttaacgg cgggatataa catgagctgt cttcggtatc gtcgtatccc
     5641 actaccgaga tatccgcacc aacgcgcagc ccggactcgg taatggcgcg cattgcgccc
     5701 agcgccatct gatcgttggc aaccagcatc gcagtgggaa cgatgccctc attcagcatt
     5761 tgcatggttt gttgaaaacc ggacatggca ctccagtcgc cttcccgttc cgctatcggc
     5821 tgaatttgat tgcgagtgag atatttatgc cagccagcca gacgcagacg cgccgagaca
     5881 gaacttaatg ggcccgctaa cagcgcgatt tgctggtgac ccaatgcgac cagatgctcc
     5941 acgcccagtc gcgtaccgtc ttcatgggag aaaataatac tgttg
//
