LOCUS       sequence-369505-        6048 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Encodes enhanced green flourescent protein linked to Pep-H 
            self-assembling peptide via a GS-linker sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6048)
  AUTHORS   Shanbhag BK, Liu C, Pradeep GC, Younas T, Hu KKY, Fulcher AJ, Struwe
            WB, Steer D, Dumsday G, Harper IS, Kukura P, Haritos VS, He L
  TITLE     Custom Design of Protein Particles as Multifunctional Biomaterials
  JOURNAL   Adv. Funct. Mater. 2022, 32, 2108039
REFERENCE   2  (bases 1 to 6048)
  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: "Adv. Funct.
            Mater. 2022, 32, 2108039"
COMMENT     Sequence Label: pET28a-eGFP-Pep-H
FEATURES             Location/Qualifiers
     source          1..6048
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     971..990
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(1010..1087)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     1293..1312
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     1396..1415
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1396..1414
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1415..1439
                     /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             1454..1476
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             1489..1506
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             2308..2325
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2378..2396)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2392..2439
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      2476..2931
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2563..2582)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2773..2794
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(3023..3838)
                     /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(3023..3838)
                     /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     3746..3765
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     3830..3849
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      3960..4548
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4449..4468
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4702..4719
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4734..4873
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4799..4818)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4959..4981
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4975..5166)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(join(5975..6048,1..1009))
                     /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 cggtttgcgt attgggcgcc agggtggttt ttcttttcac cagtgagacg ggcaacagct
       61 gattgccctt caccgcctgg ccctgagaga gttgcagcaa gcggtccacg ctggtttgcc
      121 ccagcaggcg aaaatcctgt ttgatggtgg ttaacggcgg gatataacat gagctgtctt
      181 cggtatcgtc gtatcccact accgagatat ccgcaccaac gcgcagcccg gactcggtaa
      241 tggcgcgcat tgcgcccagc gccatctgat cgttggcaac cagcatcgca gtgggaacga
      301 tgccctcatt cagcatttgc atggtttgtt gaaaaccgga catggcactc cagtcgcctt
      361 cccgttccgc tatcggctga atttgattgc gagtgagata tttatgccag ccagccagac
      421 gcagacgcgc cgagacagaa cttaatgggc ccgctaacag cgcgatttgc tggtgaccca
      481 atgcgaccag atgctccacg cccagtcgcg taccgtcttc atgggagaaa ataatactgt
      541 tgatgggtgt ctggtcagag acatcaagaa ataacgccgg aacattagtg caggcagctt
      601 ccacagcaat ggcatcctgg tcatccagcg gatagttaat gatcagccca ctgacgcgtt
      661 gcgcgagaag attgtgcacc gccgctttac aggcttcgac gccgcttcgt tctaccatcg
      721 acaccaccac gctggcaccc agttgatcgg cgcgagattt aatcgccgcg acaatttgcg
      781 acggcgcgtg cagggccaga ctggaggtgg caacgccaat cagcaacgac tgtttgcccg
      841 ccagttgttg tgccacgcgg ttgggaatgt aattcagctc cgccatcgcc gcttccactt
      901 tttcccgcgt tttcgcagaa acgtggctgg cctggttcac cacgcgggaa acggtctgat
      961 aagagacacc ggcatactct gcgacatcgt ataacgttac tggtttcaca ttcaccaccc
     1021 tgaattgact ctcttccggg cgctatcatg ccataccgcg aaaggttttg cgccattcga
     1081 tggtgtccgg gatctcgacg ctctccctta tgcgactcct gcattaggaa gcagcccagt
     1141 agtaggttga ggccgttgag caccgccgcc gcaaggaatg gtgcatgcaa ggagatggcg
     1201 cccaacagtc ccccggccac ggggcctgcc accataccca cgccgaaaca agcgctcatg
     1261 agcccgaagt ggcgagcccg atcttcccca tcggtgatgt cggcgatata ggcgccagca
     1321 accgcacctg tggcgccggt gatgccggcc acgatgcgtc cggcgtagag gatcgagatc
     1381 tcgatcccgc gaaattaata cgactcacta taggggaatt gtgagcggat aacaattccc
     1441 ctctagaaat aattttgttt aactttaaga aggagatata ccatgggtca ccaccaccac
     1501 caccacaccg acccgatggt gagcaaaggc gaggaactgt tcaccggtgt ggttccgatc
     1561 ctggtggaac tggacggcga tgttaacggt cacaagttta gcgttagcgg cgagggcgaa
     1621 ggtgatgcga cctacggcaa gctgaccctg aaattcattt gcaccaccgg taaactgccg
     1681 gtgccgtggc cgaccctggt taccaccctg gcgtacggtg tgctgtgctt tagccgttat
     1741 ccggaccaca tgaagcagca cgatttcttt aaaagcgcga tgccggaagg ctacgttcaa
     1801 gagcgtacca tcttctttaa ggacgatggt aactataaaa cccgtgcgga agtgaagttc
     1861 gagggcgaca ccctggttaa ccgtatcgaa ctgaagggta ttgactttaa agaggatggc
     1921 aacattctgg gtcacaagct ggaatacaac tataacagcc acaacgtgta catcatggcg
     1981 gataagcaga aaaacggcat taagagcaac tttaaaatcc gtcacaacat tgaggacggt
     2041 agcgtgcaac tggcggatca ctaccagcaa aacaccccga ttggtgatgg tccggttctg
     2101 ctgccggata accactatct gagcacccag agcaagctga gcaaagaccc gaacgaaaaa
     2161 cgtgatcaca tggtgctgct ggagttcgtt accgcggcgg gcattaccct gggtatggac
     2221 gagctgtata agggtggcgg tggcagcggt ggcggtggca gctggttccg ttttgagtgg
     2281 gagttcgagt tttggtaata actcgagcac caccaccacc accactgaga tccggctgct
     2341 aacaaagccc gaaaggaagc tgagttggct gctgccaccg ctgagcaata actagcataa
     2401 ccccttgggg cctctaaacg ggtcttgagg ggttttttgc tgaaaggagg aactatatcc
     2461 ggattggcga atgggacgcg ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta
     2521 cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc cgctcctttc gctttcttcc
     2581 cttcctttct cgccacgttc gccggctttc cccgtcaagc tctaaatcgg gggctccctt
     2641 tagggttccg atttagtgct ttacggcacc tcgaccccaa aaaacttgat tagggtgatg
     2701 gttcacgtag tgggccatcg ccctgataga cggtttttcg ccctttgacg ttggagtcca
     2761 cgttctttaa tagtggactc ttgttccaaa ctggaacaac actcaaccct atctcggtct
     2821 attcttttga tttataaggg attttgccga tttcggccta ttggttaaaa aatgagctga
     2881 tttaacaaaa atttaacgcg aattttaaca aaatattaac gcttacaatt taggtggcac
     2941 ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac attcaaatat
     3001 gtatccgctc atgaattaat tcttagaaaa actcatcgag catcaaatga aactgcaatt
     3061 tattcatatc aggattatca ataccatatt tttgaaaaag ccgtttctgt aatgaaggag
     3121 aaaactcacc gaggcagttc cataggatgg caagatcctg gtatcggtct gcgattccga
     3181 ctcgtccaac atcaatacaa cctattaatt tcccctcgtc aaaaataagg ttatcaagtg
     3241 agaaatcacc atgagtgacg actgaatccg gtgagaatgg caaaagttta tgcatttctt
     3301 tccagacttg ttcaacaggc cagccattac gctcgtcatc aaaatcactc gcatcaacca
     3361 aaccgttatt cattcgtgat tgcgcctgag cgagacgaaa tacgcgatcg ctgttaaaag
     3421 gacaattaca aacaggaatc gaatgcaacc ggcgcaggaa cactgccagc gcatcaacaa
     3481 tattttcacc tgaatcagga tattcttcta atacctggaa tgctgttttc ccggggatcg
     3541 cagtggtgag taaccatgca tcatcaggag tacggataaa atgcttgatg gtcggaagag
     3601 gcataaattc cgtcagccag tttagtctga ccatctcatc tgtaacatca ttggcaacgc
     3661 tacctttgcc atgtttcaga aacaactctg gcgcatcggg cttcccatac aatcgataga
     3721 ttgtcgcacc tgattgcccg acattatcgc gagcccattt atacccatat aaatcagcat
     3781 ccatgttgga atttaatcgc ggcctagagc aagacgtttc ccgttgaata tggctcataa
     3841 caccccttgt attactgttt atgtaagcag acagttttat tgttcatgac caaaatccct
     3901 taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct
     3961 tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca
     4021 gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc
     4081 agcagagcgc agataccaaa tactgtcctt ctagtgtagc cgtagttagg ccaccacttc
     4141 aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct
     4201 gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag
     4261 gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc
     4321 tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg
     4381 agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag
     4441 cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt
     4501 gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac
     4561 gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgtt ctttcctgcg
     4621 ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga taccgctcgc
     4681 cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga gcgcctgatg
     4741 cggtattttc tccttacgca tctgtgcggt atttcacacc gcaatggtgc actctcagta
     4801 caatctgctc tgatgccgca tagttaagcc agtatacact ccgctatcgc tacgtgactg
     4861 ggtcatggct gcgccccgac acccgccaac acccgctgac gcgccctgac gggcttgtct
     4921 gctcccggca tccgcttaca gacaagctgt gaccgtctcc gggagctgca tgtgtcagag
     4981 gttttcaccg tcatcaccga aacgcgcgag gcagctgcgg taaagctcat cagcgtggtc
     5041 gtgaagcgat tcacagatgt ctgcctgttc atccgcgtcc agctcgttga gtttctccag
     5101 aagcgttaat gtctggcttc tgataaagcg ggccatgtta agggcggttt tttcctgttt
     5161 ggtcactgat gcctccgtgt aagggggatt tctgttcatg ggggtaatga taccgatgaa
     5221 acgagagagg atgctcacga tacgggttac tgatgatgaa catgcccggt tactggaacg
     5281 ttgtgagggt aaacaactgg cggtatggat gcggcgggac cagagaaaaa tcactcaggg
     5341 tcaatgccag cgcttcgtta atacagatgt aggtgttcca cagggtagcc agcagcatcc
     5401 tgcgatgcag atccggaaca taatggtgca gggcgctgac ttccgcgttt ccagacttta
     5461 cgaaacacgg aaaccgaaga ccattcatgt tgttgctcag gtcgcagacg ttttgcagca
     5521 gcagtcgctt cacgttcgct cgcgtatcgg tgattcattc tgctaaccag taaggcaacc
     5581 ccgccagcct agccgggtcc tcaacgacag gagcacgatc atgcgcaccc gtggggccgc
     5641 catgccggcg ataatggcct gcttctcgcc gaaacgtttg gtggcgggac cagtgacgaa
     5701 ggcttgagcg agggcgtgca agattccgaa taccgcaagc gacaggccga tcatcgtcgc
     5761 gctccagcga aagcggtcct cgccgaaaat gacccagagc gctgccggca cctgtcctac
     5821 gagttgcatg ataaagaaga cagtcataag tgcggcgacg atagtcatgc cccgcgccca
     5881 ccggaaggag ctgactgggt tgaaggctct caagggcatc ggtcgagatc ccggtgccta
     5941 atgagtgagc taacttacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa
     6001 cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagagg
//
