LOCUS       sequence-77958-e        7697 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses a fibronectin fusion with gelonin targeting EGFR in 
            bacteria.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7697)
  AUTHORS   Pirie CM, Hackel BJ, Rosenblum MG, Wittrup KD
  TITLE     Convergent potency of internalized gelonin immunotoxins across 
            varied cell lines, antigens, and targeting moieties.
  JOURNAL   J Biol Chem. 2011 Feb 11;286(6):4165-72. doi: 
            10.1074/jbc.M110.186973. Epub 2010 Dec 7.
  PUBMED    21138845
REFERENCE   2  (bases 1 to 7697)
  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"; doi: 
            "10.1074/jbc.M110.186973"; journalName: "J Biol Chem"; date: 
            "2011-02-11- 11"; volume: "286"; issue: "6"; pages: "4165-72"
COMMENT     Sequence Label: pMal-E4rGel
FEATURES             Location/Qualifiers
     source          1..7697
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     44..61
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    76..218
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(144..163)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     304..326
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(320..511)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     promoter        1007..1084
                     /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             1085..2167
                     /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(1104..1123)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(2410..2415,2416..2431,2432..2438)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    2446..2462
                     /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     2451..2473
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     CDS             2532..3632
                     /codon_start=1
                     /gene="malE (mutated)"
                     /product="maltose binding protein from E. coli"
                     /label=MBP
                     /note="This version of the gene does not encode a signal 
                     sequence, so MBP will remain in the cytosol."
                     /translation="MKIEEGKLVIWINGDKGYNGLAEVGKKFEKDTGIKVTVEHPDKLE
                     EKFPQVAATGDGPDIIFWAHDRFGGYAQSGLLAEITPDKAFQDKLYPFTWDAVRYNGKL
                     IAYPIAVEALSLIYNKDLLPNPPKTWEEIPALDKELKAKGKSALMFNLQEPYFTWPLIA
                     ADGGYAFKYENGKYDIKDVGVDNAGAKAGLTFLVDLIKNKHMNADTDYSIAEAAFNKGE
                     TAMTINGPWAWSNIDTSKVNYGVTVLPTFKGQPSKPFVGVLSAGINAASPNKELAKEFL
                     ENYLLTDEGLEAVNKDKPLGAVALKSYEEELAKDPRIAATMENAQKGEIMPNIPQMSAF
                     WYAVRTAVINAASGRQTVDEALKDAQT"
     primer_bind     3606..3629
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     CDS             3681..3692
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(4791..4808)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4791..4807)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4800..4822)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(4986..5003)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      5156..5242
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      5334..5361
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        5381..5472
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5473..5541,5542..6333)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERSPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(5691..5710)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      complement(6375..6888)
                     /direction=LEFT
                     /label=M13 ori
                     /note="M13 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6570..6591)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6782..6801
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     rep_origin      6999..7587
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7488..7507
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tgagtgagct gataccgctc gccgcagccg aacgaccgag cgcagcgagt cagtgagcga
       61 ggaagcggaa gagcgcctga tgcggtattt tctccttacg catctgtgcg gtatttcaca
      121 ccgcatatat ggtgcactct cagtacaatc tgctctgatg ccgcatagtt aagccagtat
      181 acactccgct atcgctacgt gactgggtca tggctgcgcc ccgacacccg ccaacacccg
      241 ctgacgcgcc ctgacgggct tgtctgctcc cggcatccgc ttacagacaa gctgtgaccg
      301 tctccgggag ctgcatgtgt cagaggtttt caccgtcatc accgaaacgc gcgaggcagc
      361 tgcggtaaag ctcatcagcg tggtcgtgca gcgattcaca gatgtctgcc tgttcatccg
      421 cgtccagctc gttgagtttc tccagaagcg ttaatgtctg gcttctgata aagcgggcca
      481 tgttaagggc ggttttttcc tgtttggtca ctgatgcctc cgtgtaaggg ggatttctgt
      541 tcatgggggt aatgataccg atgaaacgag agaggatgct cacgatacgg gttactgatg
      601 atgaacatgc ccggttactg gaacgttgtg agggtaaaca actggcggta tggatgcggc
      661 gggaccagag aaaaatcact cagggtcaat gccagcgctt cgttaataca gatgtaggtg
      721 ttccacaggg tagccagcag catcctgcga tgcagatccg gaacataatg gtgcagggcg
      781 ctgacttccg cgtttccaga ctttacgaaa cacggaaacc gaagaccatt catgttgttg
      841 ctcaggtcgc agacgttttg cagcagcagt cgcttcacgt tcgctcgcgt atcggtgatt
      901 cattctgcta accagtaagg caaccccgcc agcctagccg ggtcctcaac gacaggagca
      961 cgatcatgcg cacccgtggc caggacccaa cgctgcccga aattccgaca ccatcgaatg
     1021 gtgcaaaacc tttcgcggta tggcatgata gcgcccggaa gagagtcaat tcagggtggt
     1081 gaatgtgaaa ccagtaacgt tatacgatgt cgcagagtat gccggtgtct cttatcagac
     1141 cgtttcccgc gtggtgaacc aggccagcca cgtttctgcg aaaacgcggg aaaaagtgga
     1201 agcggcgatg gcggagctga attacattcc caaccgcgtg gcacaacaac tggcgggcaa
     1261 acagtcgttg ctgattggcg ttgccacctc cagtctggcc ctgcacgcgc cgtcgcaaat
     1321 tgtcgcggcg attaaatctc gcgccgatca actgggtgcc agcgtggtgg tgtcgatggt
     1381 agaacgaagc ggcgtcgaag cctgtaaagc ggcggtgcac aatcttctcg cgcaacgcgt
     1441 cagtgggctg atcattaact atccgctgga tgaccaggat gccattgctg tggaagctgc
     1501 ctgcactaat gttccggcgt tatttcttga tgtctctgac cagacaccca tcaacagtat
     1561 tattttctcc catgaagacg gtacgcgact gggcgtggag catctggtcg cattgggtca
     1621 ccagcaaatc gcgctgttag cgggcccatt aagttctgtc tcggcgcgtc tgcgtctggc
     1681 tggctggcat aaatatctca ctcgcaatca aattcagccg atagcggaac gggaaggcga
     1741 ctggagtgcc atgtccggtt ttcaacaaac catgcaaatg ctgaatgagg gcatcgttcc
     1801 cactgcgatg ctggttgcca acgatcagat ggcgctgggc gcaatgcgcg ccattaccga
     1861 gtccgggctg cgcgttggtg cggatatctc ggtagtggga tacgacgata ccgaagacag
     1921 ctcatgttat atcccgccgt taaccaccat caaacaggat tttcgcctgc tggggcaaac
     1981 cagcgtggac cgcttgctgc aactctctca gggccaggcg gtgaagggca atcagctgtt
     2041 gcccgtctca ctggtgaaaa gaaaaaccac cctggcgccc aatacgcaaa ccgcctctcc
     2101 ccgcgcgttg gccgattcat taatgcagct ggcacgacag gtttcccgac tggaaagcgg
     2161 gcagtgagcg caacgcaatt aatgtaagtt agctcactca ttaggcacaa ttctcatgtt
     2221 tgacagctta tcatcgactg cacggtgcac caatgcttct ggcgtcaggc agccatcgga
     2281 agctgtggta tggctgtgca ggtcgtaaat cactgcataa ttcgtgtcgc tcaaggcgca
     2341 ctcccgttct ggataatgtt ttttgcgccg acatcataac ggttctggca aatattctga
     2401 aatgagctgt tgacaattaa tcatcggctc gtataatgtg tggaattgtg agcggataac
     2461 aatttcacac aggaaacagc cagtccgttt aggtgttttc acgagcactt caccaacaag
     2521 gaccatagca tatgaaaatc gaagaaggta aactggtaat ctggattaac ggcgataaag
     2581 gctataacgg tctcgctgaa gtcggtaaga aattcgagaa agataccgga attaaagtca
     2641 ccgttgagca tccggataaa ctggaagaga aattcccaca ggttgcggca actggcgatg
     2701 gccctgacat tatcttctgg gcacacgacc gctttggtgg ctacgctcaa tctggcctgt
     2761 tggctgaaat caccccggac aaagcgttcc aggacaagct gtatccgttt acctgggatg
     2821 ccgtacgtta caacggcaag ctgattgctt acccgatcgc tgttgaagcg ttatcgctga
     2881 tttataacaa agatctgctg ccgaacccgc caaaaacctg ggaagagatc ccggcgctgg
     2941 ataaagaact gaaagcgaaa ggtaagagcg cgctgatgtt caacctgcaa gaaccgtact
     3001 tcacctggcc gctgattgct gctgacgggg gttatgcgtt caagtatgaa aacggcaagt
     3061 acgacattaa agacgtgggc gtggataacg ctggcgcgaa agcgggtctg accttcctgg
     3121 ttgacctgat taaaaacaaa cacatgaatg cagacaccga ttactccatc gcagaagctg
     3181 cctttaataa aggcgaaaca gcgatgacca tcaacggccc gtgggcatgg tccaacatcg
     3241 acaccagcaa agtgaattat ggtgtaacgg tactgccgac cttcaagggt caaccatcca
     3301 aaccgttcgt tggcgtgctg agcgcaggta ttaacgccgc cagtccgaac aaagagctgg
     3361 caaaagagtt cctcgaaaac tatctgctga ctgatgaagg tctggaagcg gttaataaag
     3421 acaaaccgct gggtgccgta gcgctgaagt cttacgagga agagttggcg aaagatccac
     3481 gtattgccgc cactatggaa aacgcccaga aaggtgaaat catgccgaac atcccgcaga
     3541 tgtccgcttt ctggtatgcc gtgcgtactg cggtgatcaa cgccgccagc ggtcgtcaga
     3601 ctgtcgatga agccctgaaa gacgcgcaga ctaattcgag ctcgaacaac aacaacaata
     3661 acaataacaa caacctcggg atcgagggaa ggatttcaga attcgctagc gtttctgatg
     3721 ttccgaggga cctggaagtt gttgctgcga cccccaccag cctactgatc agctggtatc
     3781 atcctttcta ttatgtcgcg cattcttaca ggatcactta cggagaaaca ggaggaaata
     3841 gccctgtcca ggagttcact gtgcctcgtt cgccctggtt tgctaccatc agcggcctta
     3901 aacctggagt tgattatacc atcactgtgt atgctgtcac tgatagtaac ggttctcatc
     3961 caatttccat taattaccga acagaaattg acaaaccatc ccagggatcc ggaggtggag
     4021 gtagcggtct ggacacggtt tctttctcta ctaaaggtgc cacttacatt acctacgtta
     4081 acttcctgaa cgaactgcgc gtgaaactga aaccggaggg caacagccac ggtatcccgc
     4141 tgctgcgtaa aaagtgtgac gacccgggta aatgctttgt cctggtggcg ctgtctaatg
     4201 acaacggtca gctggctgaa attgccattg acgtgacgtc tgtttacgtt gtaggctacc
     4261 aggtgcgtaa tcgctcttac ttctttaagg atgcaccgga tgcagcgtat gagggcctgt
     4321 tcaaaaacac catcaagacc cgtctgcatt tcggcggctc ttacccgtcc ctggagggcg
     4381 agaaagcgta ccgcgaaacc accgaactgg gcattgagcc gctgcgtatc ggcatcaaaa
     4441 agctggacga gaatgcaatc gataactaca aaccgaccga aatcgcttcc tccctgctgg
     4501 ttgttatcca aatggtgtcc gaggccgccc gtttcacttt catcgaaaac cagattcgta
     4561 acaacttcca gcagcgtatt cgtccggcca acaacaccat ttccctggaa aacaaatggg
     4621 gtaagctgag ctttcagatt cgcacctctg gtgcgaacgg tatgttttct gaagcagtgg
     4681 aactggaacg tgcaaatggt aaaaagtact atgtgactgc cgtggatcaa gttaaaccga
     4741 aaatcgcact gctgaaattc gtggataagg acccgaaata gaagcttggc actggccgtc
     4801 gttttacaac gtcgtgactg ggaaaaccct ggcgttaccc aacttaatcg ccttgcagca
     4861 catccccctt tcgccagctg gcgtaatagc gaagaggccc gcaccgatcg cccttcccaa
     4921 cagttgcgca gcctgaatgg cgaatggcag cttggctgtt ttggcggatg agataagatt
     4981 ttcagcctga tacagattaa atcagaacgc agaagcggtc tgataaaaca gaatttgcct
     5041 ggcggcagta gcgcggtggt cccacctgac cccatgccga actcagaagt gaaacgccgt
     5101 agcgccgatg gtagtgtggg gtctccccat gcgagagtag ggaactgcca ggcatcaaat
     5161 aaaacgaaag gctcagtcga aagactgggc ctttcgtttt atctgttgtt tgtcggtgaa
     5221 cgctctcctg agtaggacaa atccgccggg agcggatttg aacgttgcga agcaacggcc
     5281 cggagggtgg cgggcaggac gcccgccata aactgccagg catcaaatta agcagaaggc
     5341 catcctgacg gatggccttt ttgcgtttct acaaactctt tttgtttatt tttctaaata
     5401 cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga
     5461 aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca
     5521 ttttgccttc ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat
     5581 cagttgggtg cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag
     5641 agttttcgcc ccgaagaacg ttctccaatg atgagcactt ttaaagttct gctatgtggc
     5701 gcggtattat cccgtgttga cgccgggcaa gagcaactcg gtcgccgcat acactattct
     5761 cagaatgact tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca
     5821 gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt
     5881 ctgacaacga tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat
     5941 gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt
     6001 gacaccacga tgcctgtagc aatggcaaca acgttgcgca aactattaac tggcgaacta
     6061 cttactctag cttcccggca acaattaata gactggatgg aggcggataa agttgcagga
     6121 ccacttctgc gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt
     6181 gagcgtgggt ctcgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc
     6241 gtagttatct acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct
     6301 gagataggtg cctcactgat taagcattgg taactgtcag accaagttta ctcatatata
     6361 ctttagattg atttaccccg gttgataatc agaaaagccc caaaaacagg aagattgtat
     6421 aagcaaatat ttaaattgta aacgttaata ttttgttaaa attcgcgtta aatttttgtt
     6481 aaatcagctc attttttaac caataggccg aaatcggcaa aatcccttat aaatcaaaag
     6541 aatagaccga gatagggttg agtgttgttc cagtttggaa caagagtcca ctattaaaga
     6601 acgtggactc caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg
     6661 aaccatcacc caaatcaagt tttttggggt cgaggtgccg taaagcacta aatcggaacc
     6721 ctaaagggag cccccgattt agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg
     6781 aagggaagaa agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc
     6841 gcgtaaccac cacacccgcc gcgcttaatg cgccgctaca gggcgcgtaa aaggatctag
     6901 gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac
     6961 tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc
     7021 gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat
     7081 caagagctac caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat
     7141 actgtccttc tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct
     7201 acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt
     7261 cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg
     7321 gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta
     7381 cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg
     7441 gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg
     7501 tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc
     7561 tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg
     7621 gccttttgct ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat
     7681 aaccgtatta ccgcctt
//
