LOCUS       sequence-213018-        3569 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression vector for LSS-EGFP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3569)
  TITLE     Gadella lab plasmids
REFERENCE   2  (bases 1 to 3569)
  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: pRSETb-LSS-mEGFP
FEATURES             Location/Qualifiers
     source          1..3569
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     20..39
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        20..38
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             70..92
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             112..129
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             142..162
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     regulatory      178..187
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(184..186,187..189,190..903)
                     /codon_start=1
                     /product="enhanced GFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mEGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLSYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSIQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(229..250)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(490..509)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     837..858
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(970..988)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      984..1031
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1128..1583
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1215..1234)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1425..1446
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        1609..1713
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1714..1782,1783..2574)
                     /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     complement(1932..1951)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2745..3333
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3234..3253
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3487..3504
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 gatctcgatc ccgcgaaatt aatacgactc actataggga gaccacaacg gtttccctct
       61 agaaataatt ttgtttaact ttaagaagga gatatacata tgcggggttc tcatcatcat
      121 catcatcatg gtatggctag tgaaaacctg tattttcagg ggctagcgct accggtcgcc
      181 accatggtga gcaagggcga ggagctgttc accggggtgg tgcccatcct ggtcgagctg
      241 gacggcgacg taaacggcca caagttcagc gtgtccggcg agggcgaggg cgatgccacc
      301 tacggcaagc tgaccctgaa gttcatctgc accaccggca agctgcccgt gccctggccc
      361 accctcgtga ccaccctgag ctacggcgtg cagtgcttca gccgctaccc cgaccacatg
      421 aagcagcacg acttcttcaa gtccgccatg cccgaaggct acgtccagga gcgcaccatc
      481 ttcttcaagg acgacggcaa ctacaagacc cgcgccgagg tgaagttcga gggcgacacc
      541 ctggtgaacc gcatcgagct gaagggcatc gacttcaagg aggacggcaa catcctgggg
      601 cacaagctgg agtacaacta caacagccac aacgtctata tcatggccga caagcagaag
      661 aacggcatca aggtgaactt caagatccgc cacaacatcg aggacggcag cgtgcagctc
      721 gccgaccact accagcagaa cacccccatc ggcgacggcc ccgtgctgct gcccgacaac
      781 cactacctga gcatccagtc caagctgagc aaagacccca acgagaagcg cgatcacatg
      841 gtcctgctgg agttcgtgac cgccgccggg atcactctcg gcatggacga gctgtacaag
      901 taaagcggcc gcgcaagctt gatccggctg ctaacaaagc ccgaaaggaa gctgagttgg
      961 ctgctgccac cgctgagcaa taactagcat aaccccttgg ggcctctaaa cgggtcttga
     1021 ggggtttttt gctgaaagga ggaactatat ccggatctgg cgtaatagcg aagaggcccg
     1081 caccgatcgc ccttcccaac agttgcgcag cctgaatggc gaatgggacg cgccctgtag
     1141 cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag
     1201 cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt
     1261 tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg ctttacggca
     1321 cctcgacccc aaaaaacttg attagggtga tggttcacgt agtgggccat cgccctgata
     1381 gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac tcttgttcca
     1441 aactggaaca acactcaacc ctatctcggt ctattctttt gatttataag ggattttgcc
     1501 gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg cgaattttaa
     1561 caaaatatta acgcttacaa tttaggtggc acttttcggg gaaatgtgcg cggaacccct
     1621 atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgagaca ataaccctga
     1681 taaatgcttc aataatattg aaaaaggaag agtatgagta ttcaacattt ccgtgtcgcc
     1741 cttattccct tttttgcggc attttgcctt cctgtttttg ctcacccaga aacgctggtg
     1801 aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg gttacatcga actggatctc
     1861 aacagcggta agatccttga gagttttcgc cccgaagaac gttttccaat gatgagcact
     1921 tttaaagttc tgctatgtgg cgcggtatta tcccgtattg acgccgggca agagcaactc
     1981 ggtcgccgca tacactattc tcagaatgac ttggttgagt actcaccagt cacagaaaag
     2041 catcttacgg atggcatgac agtaagagaa ttatgcagtg ctgccataac catgagtgat
     2101 aacactgcgg ccaacttact tctgacaacg atcggaggac cgaaggagct aaccgctttt
     2161 ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa
     2221 gccataccaa acgacgagcg tgacaccacg atgcctgtag caatggcaac aacgttgcgc
     2281 aaactattaa ctggcgaact acttactcta gcttcccggc aacaattaat agactggatg
     2341 gaggcggata aagttgcagg accacttctg cgctcggccc ttccggctgg ctggtttatt
     2401 gctgataaat ctggagccgg tgagcgtggg tctcgcggta tcattgcagc actggggcca
     2461 gatggtaagc cctcccgtat cgtagttatc tacacgacgg ggagtcaggc aactatggat
     2521 gaacgaaata gacagatcgc tgagataggt gcctcactga ttaagcattg gtaactgtca
     2581 gaccaagttt actcatatat actttagatt gatttaaaac ttcattttta atttaaaagg
     2641 atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg tgagttttcg
     2701 ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga tccttttttt
     2761 ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg
     2821 ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag agcgcagata
     2881 ccaaatactg ttcttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca
     2941 ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag
     3001 tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc
     3061 tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga
     3121 tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg
     3181 tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc agggggaaac
     3241 gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg tcgatttttg
     3301 tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg
     3361 ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc ccctgattct
     3421 gtggataacc gtattaccgc ctttgagtga gctgataccg ctcgccgcag ccgaacgacc
     3481 gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc caatacgcaa accgcctctc
     3541 cccgcgcgtt ggccgattca ttaatgcag
//
