LOCUS       sequence-467321-        5484 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Inducible protein expression in insect cell culture. Stable cell 
            line generation..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5484)
  TITLE     Vascular Endothelial Growth Factor C-Loaded Lignin Nanoparticles
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 5484)
  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"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: pMT-Ex
FEATURES             Location/Qualifiers
     source          1..5484
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(29..51)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     151..170
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     364..386
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     379..395
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        401..827
                     /label=MT promoter
                     /note="Drosophila metallothionein promoter"
     sig_peptide     845..898
                     /codon_start=1
                     /product="Drosophila BiP signal sequence"
                     /label=BiP signal sequence
                     /translation="MKLCILLAVVAFVGLSLG"
     CDS             992..1033
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             1043..1060
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1083..1100)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1133..1267
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     promoter        1459..1738
                     /label=copia promoter
                     /note="strong promoter from the Drosophila transposable 
                     element copia (Sinclair et al., 1986)"
     regulatory      1757..1766
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1763..2788
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     CDS             1763..2788
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     polyA_signal    3101..3235
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3263..3279)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3263..3279)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3276..3298)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3287..3303
                     /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(3311..3317,3318..3335,3336..3341))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3356..3377
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3494..3511)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3665..4253)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3745..3764)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4424..5215,5216..5284))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     5047..5066
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5285..5389)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5457..5475
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca
       61 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
      121 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
      181 accatatgcg gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc atcaggcgcc
      241 attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat
      301 tacgccagct ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta acgccagggt
      361 tttcccagtc acgacgttgt aaaacgacgg ccagtgaatt aattcgttgc aggacaggat
      421 gtggtgcccg atgtgactag ctctttgctg caggccgtcc tatcctctgg ttccgataag
      481 agacccagaa ctccggcccc ccaccgccca ccgccacccc catacatatg tggtacgcaa
      541 gtaagagtgc ctgcgcatgc cccatgtgcc ccaccaagag ctttgcatcc catacaagtc
      601 cccaaagtgg agaaccgaac caattcttcg cgggcagaac aaaagcttct gcacacgtct
      661 ccactcgaat ttggagccgg ccggcgtgtg caaaagaggt gaatcgaacg aaagacccgt
      721 gtgtaaagcc gcgtttccaa aatgtataaa accgagagca tctggccaat gtgcatcagt
      781 tgtggtcagc agcaaaatca agtgaatcat ctcagtgcaa ctaaaggggg gatccgatct
      841 caatatgaag ttatgcatat tactggccgt cgtggccttt gttggcctct cgctcggaga
      901 tctccatggc ccggggtacc tactagtcca gtgtggtgga attctgcaga tatccagcac
      961 agtggcggcc gctcgaggtc acccattcga aggtaagcct atccctaacc ctctcctcgg
     1021 tctcgattct acgcgtaccg gtcatcatca ccatcaccat tgagtttaaa cccgctgatc
     1081 agcctcgact gtgccttcta aggcctgagc tcgctgatca gcctcgatcg aggatccaga
     1141 catgataaga tacattgatg agtttggaca aaccacaact agaatgcagt gaaaaaaatg
     1201 ctttatttgt gaaatttgtg atgctattgc tttatttgta accattataa gctgcaataa
     1261 acaagttaac aacaacaatt gcattcattt tatgtttcag gttcaggggg aggtgtggga
     1321 ggttttttaa agcaagtaaa acctctacaa atgtggtatg gctgattatg atcagtcgac
     1381 tctagaggat ccggtgcgtg gtggtttcat gcttctggga acggcaaatg ggtttaggat
     1441 tgggaacccc tcatcatctg ttggaatata ctattcaacc tacaaaagta acgttaaaca
     1501 acactacttt atatttgata tgaatggcca caccttttat gccataaaac atattgtaag
     1561 agaataccac tctttttatt ccttctttcc ttcttgtacg ttttttgctg taagtaggtc
     1621 gtggtgctgg tgttgcagtt gaaataactt aaaatataaa tcataaaact caaacataaa
     1681 cttgactatt tatttattta ttaagaaagg aaatataaat tataaattac aacaggttat
     1741 ggggatcccc agcttggcca ccatgggaaa gcctgagctg accgcgacgt ctgtcgagaa
     1801 gtttctgatc gaaaagttcg acagcgtctc cgacctgatg cagctctcgg agggcgaaga
     1861 atctcgtgct ttcagcttcg atgtaggagg gcgtggatat gtcctgcggg taaatagctg
     1921 cgccgatggt ttctacaaag atcgttatgt ttatcggcac tttgcatcgg ccgcgctccc
     1981 gattccggaa gtgcttgaca ttggggaatt cagcgagagc ctgacctatt gcatctcccg
     2041 ccgtgcacag ggtgtcacgt tgcaagacct gcctgaaacc gaactgcccg ctgttctgca
     2101 gccggtcgcg gaggccatgg atgcgatcgc tgcggccgat cttagccaga cgagcgggtt
     2161 cggcccattc ggaccgcaag gaatcggtca atacactaca tggcgtgatt tcatatgcgc
     2221 gattgctgat ccccatgtgt atcactggca aactgtgatg gacgacaccg tcagtgcgtc
     2281 cgtcgcgcag gctctcgatg agctgatgct ttgggccgag gactgccccg aagtccggca
     2341 cctcgtgcac gcggatttcg gctccaacaa tgtcctgacg gacaatggcc gcataacagc
     2401 ggtcattgac tggagcgagg cgatgttcgg ggattcccaa tacgaggtcg ccaacatctt
     2461 cttctggagg ccgtggttgg cttgtatgga gcagcagacg cgctacttcg agcggaggca
     2521 tccggagctt gcaggatcgc cgcggctccg ggcgtatatg ctccgcattg gtcttgacca
     2581 actctatcag agcttggttg acggcaattt cgatgatgca gcttgggcgc agggtcgatg
     2641 cgacgcaatc gtccgatccg gagccgggac tgtcgggcgt acacaaatcg cccgcagaag
     2701 cgcggccgtc tggaccgatg gctgtgtaga agtactcgcc gatagtggaa accgacgccc
     2761 cagcactcgt ccgagggcaa aggaatagag tagatgccga ccgaacaaga gctgatttcg
     2821 agaacgcctc agccagcaac tcgcgcgagc ctagcaaggc aaatgcgaga gaacggcctt
     2881 acgcttggtg gcacagttct cgtccacagt tcgctaagct cgctcggctg ggtcgcggga
     2941 gggccggtcg cagtgattca ggcccttctg gattgtgttg gtccccaggg cacgattgtc
     3001 atgcccacgc actcgggtga tctgactgat cccgcagatt ggagatcgcc gcccgtgcct
     3061 gccgattggg tgcagatcag cctcgaggcc agctagcttg aacttgttta ttgcagctta
     3121 taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat ttttttcact
     3181 gcattctagt tgtggtttgt ccaaactcat caatgtatct tatcatgtct ggatcgggta
     3241 ccgagctcga attcgtaatc atgtcatagc tgtttcctgt gtgaaattgt tatccgctca
     3301 caattccaca caacatacga gccggaagca taaagtgtaa agcctggggt gcctaatgag
     3361 tgagctaact cacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt
     3421 cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc
     3481 gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg
     3541 tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa
     3601 agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg
     3661 cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga
     3721 ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg
     3781 tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg
     3841 gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc
     3901 gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg
     3961 gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca
     4021 ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt
     4081 ggcctaacta cggctacact agaagaacag tatttggtat ctgcgctctg ctgaagccag
     4141 ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg
     4201 gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc
     4261 ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt
     4321 tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt
     4381 ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca
     4441 gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg
     4501 tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac
     4561 cgcgagaccc acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg
     4621 ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc
     4681 gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta
     4741 caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac
     4801 gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc
     4861 ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac
     4921 tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact
     4981 caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa
     5041 tacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt
     5101 cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca
     5161 ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa
     5221 aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac
     5281 tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg
     5341 gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc
     5401 gaaaagtgcc acctgacgtc taagaaacca ttattatcat gacattaacc tataaaaata
     5461 ggcgtatcac gaggcccttt cgtc
//
