LOCUS       pCMV_FRB_tTA_Myc        6315 bp ds-DNA     circular     13-OCT-2021
DEFINITION  Expression of a Myc tagged VSV-G protein.
ACCESSION   .
VERSION     .
KEYWORDS    pCMV-VSV-G-Myc
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6537)
  AUTHORS   Votteler J, Ogohara C, Yi S, Hsia Y, Nattermann U, Belnap DM, King
            NP, Sundquist WI
  TITLE     Designed proteins induce the formation of nanocage-containing
            extracellular vesicles.
  JOURNAL   Nature. 2016 Dec 8;540(7632):292-295. doi: 10.1038/nature20607.
            Epub 2016 Nov 30.
  PUBMED    27919066
REFERENCE   2  (bases 1 to 6537)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jan 10, 2018 from SnapGene Server 1.1.58
            http://www.snapgene.com
COMMENT     
COMMENT     ApEinfo:methylated:1
FEATURES             Location/Qualifiers
     misc_feature    1777..1806
                     /locus_tag="Confirmed sequence (180511, #40)"
                     /label="Confirmed sequence (180511, #40)"
                     /ApEinfo_label="Confirmed sequence (180511, #40)"
                     /ApEinfo_fwdcolor="cyan"
                     /ApEinfo_revcolor="green"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
                     /ApEinfo_hidden
     source          1498..1776
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
                     /locus_tag="source:synthetic DNA construct"
                     /label="source:synthetic DNA construct"
                     /ApEinfo_label="source:synthetic DNA construct"
                     /ApEinfo_fwdcolor="pink"
                     /ApEinfo_revcolor="pink"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
                     /ApEinfo_hidden
     misc_feature    1495..1497
                     /locus_tag="confirmed sequence (180511, #40)"
                     /label="confirmed sequence (180511, #40)"
                     /ApEinfo_label="confirmed sequence (180511, #40)"
                     /ApEinfo_fwdcolor="cyan"
                     /ApEinfo_revcolor="green"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
                     /ApEinfo_hidden
     misc_feature    1495..1497
                     /locus_tag="confirmed sequence (180508, #31)"
                     /label="confirmed sequence (180508, #31)"
                     /ApEinfo_label="confirmed sequence (180508, #31)"
                     /ApEinfo_fwdcolor="cyan"
                     /ApEinfo_revcolor="green"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
                     /ApEinfo_hidden
     misc_feature    2806..3057
                     /locus_tag="confirmed sequence(#31, 180420)"
                     /label="confirmed sequence(#31, 180420)"
                     /ApEinfo_label="confirmed sequence(#31, 180420)"
                     /ApEinfo_fwdcolor="cyan"
                     /ApEinfo_revcolor="green"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
                     /ApEinfo_hidden
     source          1495..1497
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
                     /locus_tag="source:synthetic DNA construct(1)"
                     /label="source:synthetic DNA construct(1)"
                     /ApEinfo_label="source:synthetic DNA construct"
                     /ApEinfo_fwdcolor="pink"
                     /ApEinfo_revcolor="pink"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
                     /ApEinfo_hidden
     CDS             1807..2805
                     /codon_start=1
                     /product="tetracycline-controlled transactivator,
                     comprising a fusion of the tetracycline repressor TetR
                     with the C-terminal activation domain of herpes simplex
                     virus  VP16"
                     /note="In the Tet-Off(R) system, tTA binds to the
                     Tet-responsive element and stimulates transcription only
                     in the absence of tetracycline or doxycycline."
                     /translation="RLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWHVK
                     N
                     KRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTRP
                     T
                     EKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPTTD
                     S
                     MPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGSAYSRARTKNNYGSTIE
                     G
                     LLDLPDDDAPEEAGLAAPRLSFLPAGHTRRLSTAPPTDVSLGDELHLDGEDVAMAHAD
                     A LDDFDLDMLGDGDSPGPGFTPHDSAPYGALDMADFEFEQMFTDALGIDEYGG"
                     /locus_tag="tTA"
                     /label="tTA"
                     /ApEinfo_label="tTA"
                     /ApEinfo_fwdcolor="#5518fe"
                     /ApEinfo_revcolor="#e9d024"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
     CDS             1495..1497
                     /codon_start=1
                     /product="vesicular stomatitis virus G glycoprotein"
                     /note="Indiana strain"
                     /translation="MKCLLYLAFLFIGVNCKFTIVFPHNQKGNWKNVPSNYHYCPSSS
                     D
                     LNWHNDLIGTALQVKMPKSHKAIQADGWMCHASKWVTTCDFRWYGPKYITHSIRSFTP
                     S
                     VEQCKESIEQTKQGTWLNPGFPPQSCGYATVTDAEAVIVQVTPHHVLVDEYTGEWVDS
                     Q
                     FINGKCSNYICPTVHNSTTWHSDYKVKGLCDSNLISMDITFFSEDGELSSLGKEGTGF
                     R
                     SNYFAYETGGKACKMQYCKHWGVRLPSGVWFEMADKDLFAAARFPECPEGSSISAPSQ
                     T
                     SVDVSLIQDVERILDYSLCQETWSKIRAGLPISPVDLSYLAPKNPGTGPAFTIINGTL
                     K
                     YFETRYIRVDIAAPILSRMVGMISGTTTERELWDDWAPYEDVEIGPNGVLRTSSGYKF
                     P
                     LYMIGHGMLDSDLHLSSKAQVFEHPHIQDAASQLPDDESLFFGDTGLSKNPIELVEGW
                     F SSWKSSIASFFFIIGLIIGLFLVLRVGIHLCIKLKHTKKRQIYTDIEMNRLGK"
                     /locus_tag="VSV-G"
                     /label="VSV-G"
                     /ApEinfo_label="VSV-G"
                     /ApEinfo_fwdcolor="#e9d024"
                     /ApEinfo_revcolor="#e9d024"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
     primer_bind     complement(1874..1893)
                     /note="Tetracycline resistance gene, reverse primer"
                     /locus_tag="Tet-R"
                     /label="Tet-R"
                     /ApEinfo_label="Tet-R"
                     /ApEinfo_fwdcolor="#14c0bd"
                     /ApEinfo_revcolor="#4ec02b"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
     CDS             2806..2835
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /translation="EQKLISEEDL"
                     /locus_tag="Myc"
                     /label="Myc"
                     /ApEinfo_label="Myc"
                     /ApEinfo_fwdcolor="#fb2a5b"
                     /ApEinfo_revcolor="#e9d024"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
     primer_bind     complement(3065..3084)
                     /note="Rabbit beta-globin polyA region, reverse primer"
                     /locus_tag="Bglob-pA-R"
                     /label="Bglob-pA-R"
                     /ApEinfo_label="Bglob-pA-R"
                     /ApEinfo_fwdcolor="#14c0bd"
                     /ApEinfo_revcolor="#4ec02b"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
     CDS             1498..1776
                     /codon_start=1
                     /product="FKBP-rapamycin binding domain of human FRAP"
                     /translation="ILWHEMWHEGLEEASRLYFGERNVKGMFEVLEPLHAMMERGPQT
                     L KETSFNQAYGRDLMEAQEWCRKYMKSGNVKDLTQAWDLYYHVFRRISK"
                     /locus_tag="FRB"
                     /label="FRB"
                     /ApEinfo_label="FRB"
                     /ApEinfo_fwdcolor="#e9d024"
                     /ApEinfo_revcolor="#e9d024"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
     polyA_site      3130..3185
                     /note="rabbit beta-globin polyadenylation signal"
                     /locus_tag="-beta-globin poly(A) signal"
                     /label="-beta-globin poly(A) signal"
                     /ApEinfo_label="-beta-globin poly(A) signal"
                     /ApEinfo_fwdcolor="cyan"
                     /ApEinfo_revcolor="cyan"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
     primer_bind     complement(3184..3203)
                     /note="Rabbit beta-globin polyA, reverse primer. Also
                     called rb-glob-pA-term-R"
                     /locus_tag="rbglobpA-R"
                     /label="rbglobpA-R"
                     /ApEinfo_label="rbglobpA-R"
                     /ApEinfo_fwdcolor="#14c0bd"
                     /ApEinfo_revcolor="#4ec02b"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
                     /ApEinfo_hidden
     primer_bind     complement(3543..3559)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /locus_tag="M13 rev"
                     /label="M13 rev"
                     /ApEinfo_label="M13 rev"
                     /ApEinfo_fwdcolor="#14c0bd"
                     /ApEinfo_revcolor="#4ec02b"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
                     /ApEinfo_hidden
     primer_bind     complement(3543..3559)
                     /note="In lacZ gene. Also called M13-rev"
                     /locus_tag="M13 Reverse"
                     /label="M13 Reverse"
                     /ApEinfo_label="M13 Reverse"
                     /ApEinfo_fwdcolor="#14c0bd"
                     /ApEinfo_revcolor="#4ec02b"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
                     /ApEinfo_hidden
     primer_bind     complement(3556..3578)
                     /note="In lacZ gene"
                     /locus_tag="M13/pUC Reverse"
                     /label="M13/pUC Reverse"
                     /ApEinfo_label="M13/pUC Reverse"
                     /ApEinfo_fwdcolor="#14c0bd"
                     /ApEinfo_revcolor="#4ec02b"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
                     /ApEinfo_hidden
     protein_bind    3567..3583
                     /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)."
                     /locus_tag="lac operator"
                     /label="lac operator"
                     /ApEinfo_label="lac operator"
                     /ApEinfo_fwdcolor="pink"
                     /ApEinfo_revcolor="pink"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
                     /ApEinfo_hidden
     promoter        complement(3591..3621)
                     /note="promoter for the E. coli lac operon"
                     /locus_tag="lac promoter"
                     /label="lac promoter"
                     /ApEinfo_label="lac promoter"
                     /ApEinfo_fwdcolor="#346ee0"
                     /ApEinfo_revcolor="#346ee0"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
                     /ApEinfo_hidden
     protein_bind    3636..3657
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /locus_tag="CAP binding site"
                     /label="CAP binding site"
                     /ApEinfo_label="CAP binding site"
                     /ApEinfo_fwdcolor="pink"
                     /ApEinfo_revcolor="pink"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
                     /ApEinfo_hidden
     primer_bind     complement(3774..3791)
                     /note="L4440 vector, forward primer"
                     /locus_tag="L4440"
                     /label="L4440"
                     /ApEinfo_label="L4440"
                     /ApEinfo_fwdcolor="#14c0bd"
                     /ApEinfo_revcolor="#4ec02b"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
                     /ApEinfo_hidden
     rep_origin      complement(3945..4533)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /locus_tag="ori"
                     /label="ori"
                     /ApEinfo_label="ori"
                     /ApEinfo_fwdcolor="#999999"
                     /ApEinfo_revcolor="#999999"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
                     /ApEinfo_hidden
     primer_bind     complement(4025..4044)
                     /note="pBR322 origin, forward primer"
                     /locus_tag="pBR322ori-F"
                     /label="pBR322ori-F"
                     /ApEinfo_label="pBR322ori-F"
                     /ApEinfo_fwdcolor="#14c0bd"
                     /ApEinfo_revcolor="#4ec02b"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
                     /ApEinfo_hidden
     CDS             complement(4704..5564)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /note="confers resistance to ampicillin, carbenicillin,
                     and related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGY
                     I
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEY
                     S
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDR
                     W
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRS
                     A
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGA
                     S LIKHW"
                     /locus_tag="AmpR"
                     /label="AmpR"
                     /ApEinfo_label="AmpR"
                     /ApEinfo_fwdcolor="#e9d024"
                     /ApEinfo_revcolor="#e9d024"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
                     /ApEinfo_hidden
     primer_bind     5327..5346
                     /note="Ampicillin resistance gene, reverse primer"
                     /locus_tag="Amp-R"
                     /label="Amp-R"
                     /ApEinfo_label="Amp-R"
                     /ApEinfo_fwdcolor="#14c0bd"
                     /ApEinfo_revcolor="#4ec02b"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
                     /ApEinfo_hidden
     promoter        complement(5565..5669)
                     /gene="bla"
                     /locus_tag="AmpR promoter"
                     /label="AmpR promoter"
                     /ApEinfo_label="AmpR promoter"
                     /ApEinfo_fwdcolor="#346ee0"
                     /ApEinfo_revcolor="#346ee0"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
                     /ApEinfo_hidden
     rep_origin      complement(5695..6150)
                     /direction=LEFT
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
                     /locus_tag="f1 ori"
                     /label="f1 ori"
                     /ApEinfo_label="f1 ori"
                     /ApEinfo_fwdcolor="#999999"
                     /ApEinfo_revcolor="#999999"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
                     /ApEinfo_hidden
     primer_bind     complement(5832..5853)
                     /note="F1 origin, forward primer"
                     /locus_tag="F1ori-F"
                     /label="F1ori-F"
                     /ApEinfo_label="F1ori-F"
                     /ApEinfo_fwdcolor="#14c0bd"
                     /ApEinfo_revcolor="#4ec02b"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
                     /ApEinfo_hidden
     primer_bind     6044..6063
                     /note="F1 origin, reverse primer"
                     /locus_tag="F1ori-R"
                     /label="F1ori-R"
                     /ApEinfo_label="F1ori-R"
                     /ApEinfo_fwdcolor="#14c0bd"
                     /ApEinfo_revcolor="#4ec02b"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
                     /ApEinfo_hidden
     primer_bind     6277..6299
                     /note="In lacZ gene"
                     /locus_tag="M13/pUC Forward"
                     /label="M13/pUC Forward"
                     /ApEinfo_label="M13/pUC Forward"
                     /ApEinfo_fwdcolor="#14c0bd"
                     /ApEinfo_revcolor="#4ec02b"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
                     /ApEinfo_hidden
     primer_bind     6291..6308
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21)
                     Forward"
                     /locus_tag="M13 Forward"
                     /label="M13 Forward"
                     /ApEinfo_label="M13 Forward"
                     /ApEinfo_fwdcolor="#14c0bd"
                     /ApEinfo_revcolor="#4ec02b"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
                     /ApEinfo_hidden
     primer_bind     6292..6308
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /locus_tag="M13 fwd"
                     /label="M13 fwd"
                     /ApEinfo_label="M13 fwd"
                     /ApEinfo_fwdcolor="#14c0bd"
                     /ApEinfo_revcolor="#4ec02b"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
                     /ApEinfo_hidden
     enhancer        515..521
                     /note="human cytomegalovirus immediate early enhancer"
                     /locus_tag="CMV enhancer"
                     /label="CMV enhancer"
                     /ApEinfo_label="CMV enhancer"
                     /ApEinfo_fwdcolor="#5ac3fa"
                     /ApEinfo_revcolor="#5ac3fa"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
                     /ApEinfo_hidden
     source          2806..6315
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
                     /locus_tag="source:synthetic DNA construct(2)"
                     /label="source:synthetic DNA construct(2)"
                     /ApEinfo_label="source:synthetic DNA construct"
                     /ApEinfo_fwdcolor="pink"
                     /ApEinfo_revcolor="pink"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
                     /ApEinfo_hidden
     source          1..514
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
                     /locus_tag="source:synthetic DNA construct(3)"
                     /label="source:synthetic DNA construct(3)"
                     /ApEinfo_label="source:synthetic DNA construct"
                     /ApEinfo_fwdcolor="pink"
                     /ApEinfo_revcolor="pink"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
                     /ApEinfo_hidden
     source          515..1494
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
                     /locus_tag="source:synthetic DNA construct(4)"
                     /label="source:synthetic DNA construct(4)"
                     /ApEinfo_label="source:synthetic DNA construct"
                     /ApEinfo_fwdcolor="pink"
                     /ApEinfo_revcolor="pink"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
                     /ApEinfo_hidden
     misc_signal     1777..1806
                     /locus_tag="GSx2 linker"
                     /label="GSx2 linker"
                     /ApEinfo_label="GSx2 linker"
                     /ApEinfo_fwdcolor="#008040"
                     /ApEinfo_revcolor="#008040"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
     promoter        522..725
                     /note="human cytomegalovirus (CMV) immediate early
                     promoter"
                     /locus_tag="CMV promoter"
                     /label="CMV promoter"
                     /ApEinfo_label="CMV promoter"
                     /ApEinfo_fwdcolor="#346ee0"
                     /ApEinfo_revcolor="#346ee0"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
     primer_bind     676..696
                     /note="Human CMV immediate early promoter, forward primer"
                     /locus_tag="CMV-F"
                     /label="CMV-F"
                     /ApEinfo_label="CMV-F"
                     /ApEinfo_fwdcolor="#14c0bd"
                     /ApEinfo_revcolor="#4ec02b"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
     primer_bind     722..746
                     /note="Human CMV promoter, forward primer"
                     /locus_tag="LNCX"
                     /label="LNCX"
                     /ApEinfo_label="LNCX"
                     /ApEinfo_fwdcolor="#14c0bd"
                     /ApEinfo_revcolor="#4ec02b"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
                     /ApEinfo_hidden
     intron          837..1409
                     /note="intron from rabbit beta-globin gene"
                     /locus_tag="-beta-globin intron"
                     /label="-beta-globin intron"
                     /ApEinfo_label="-beta-globin intron"
                     /ApEinfo_fwdcolor="#eb606c"
                     /ApEinfo_revcolor="#eb606c"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
     primer_bind     3..22
                     /note="T7 promoter, forward primer"
                     /locus_tag="T7"
                     /label="T7"
                     /ApEinfo_label="T7"
                     /ApEinfo_fwdcolor="#14c0bd"
                     /ApEinfo_revcolor="#4ec02b"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
                     /ApEinfo_hidden
     promoter        3..21
                     /note="promoter for bacteriophage T7 RNA polymerase"
                     /locus_tag="T7 promoter"
                     /label="T7 promoter"
                     /ApEinfo_label="T7 promoter"
                     /ApEinfo_fwdcolor="#346ee0"
                     /ApEinfo_revcolor="#346ee0"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
                     /ApEinfo_hidden
     enhancer        142..514
                     /note="human cytomegalovirus immediate early enhancer"
                     /locus_tag="CMV enhancer(1)"
                     /label="CMV enhancer(1)"
                     /ApEinfo_label="CMV enhancer"
                     /ApEinfo_fwdcolor="#5ac3fa"
                     /ApEinfo_revcolor="#5ac3fa"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
     primer_bind     6291..6308
                     /locus_tag="M13-fwd"
                     /label="M13-fwd"
                     /ApEinfo_label="M13-fwd"
                     /ApEinfo_fwdcolor="cyan"
                     /ApEinfo_revcolor="green"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
     primer_bind     2..22
                     /locus_tag="T7(1)"
                     /label="T7(1)"
                     /ApEinfo_label="T7"
                     /ApEinfo_fwdcolor="cyan"
                     /ApEinfo_revcolor="green"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
     rep_origin      complement(3927..4609)
                     /locus_tag="ColE1 origin"
                     /label="ColE1 origin"
                     /ApEinfo_label="ColE1 origin"
                     /ApEinfo_fwdcolor="gray50"
                     /ApEinfo_revcolor="gray50"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
     rep_origin      complement(5827..6133)
                     /locus_tag="F1 ori"
                     /label="F1 ori"
                     /ApEinfo_label="F1 ori"
                     /ApEinfo_fwdcolor="gray50"
                     /ApEinfo_revcolor="gray50"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
     CDS             complement(6152..6220)
                     /locus_tag="LacZ alpha"
                     /label="LacZ alpha"
                     /ApEinfo_label="LacZ alpha"
                     /ApEinfo_fwdcolor="#6495ed"
                     /ApEinfo_revcolor="#6495ed"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
     misc_binding    complement(3565..3587)
                     /locus_tag="LacO"
                     /label="LacO"
                     /ApEinfo_label="LacO"
                     /ApEinfo_fwdcolor="#6495ed"
                     /ApEinfo_revcolor="#6495ed"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
     CDS             complement(4707..5366)
                     /locus_tag="AmpR(1)"
                     /label="AmpR(1)"
                     /ApEinfo_label="AmpR"
                     /ApEinfo_fwdcolor="yellow"
                     /ApEinfo_revcolor="yellow"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
     promoter        218..744
                     /locus_tag="CMV promoter(1)"
                     /label="CMV promoter(1)"
                     /ApEinfo_label="CMV promoter"
                     /ApEinfo_fwdcolor="cornflower blue"
                     /ApEinfo_revcolor="cornflower blue"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
     primer_bind     complement(1874..1893)
                     /locus_tag="TetR-R"
                     /label="TetR-R"
                     /ApEinfo_label="TetR-R"
                     /ApEinfo_fwdcolor="#14c0bd"
                     /ApEinfo_revcolor="#4ec02b"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
     primer_bind     5327..5346
                     /locus_tag="Amp-R (& -comp-R)"
                     /label="Amp-R (& -comp-R)"
                     /ApEinfo_label="Amp-R (& -comp-R)"
                     /ApEinfo_fwdcolor="#14c0bd"
                     /ApEinfo_revcolor="#4ec02b"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
     primer_bind     1281..1300
                     /locus_tag="Bglob-intron-F"
                     /label="Bglob-intron-F"
                     /ApEinfo_label="Bglob-intron-F"
                     /ApEinfo_fwdcolor="#14c0bd"
                     /ApEinfo_revcolor="#4ec02b"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
ORIGIN
        1 cgtaatacga ctcactatag ggcgaattgg agctccaccg cggtggcggc cgctctagag
       61 agcttggccc attgcatacg ttgtatccat atcataatat gtacatttat attggctcat
      121 gtccaacatt accgccatgt tgacattgat tattgactag ttattaatag taatcaatta
      181 cggggtcatt agttcatagc ccatatatgg agttccgcgt tacataactt acggtaaatg
      241 gcccgcctgg ctgaccgccc aacgaccccc gcccattgac gtcaataatg acgtatgttc
      301 ccatagtaac gccaataggg actttccatt gacgtcaatg ggtggagtat ttacggtaaa
      361 ctgcccactt ggcagtacat caagtgtatc atatgccaag tacgccccct attgacgtca
      421 atgacggtaa atggcccgcc tggcattatg cccagtacat gaccttatgg gactttccta
      481 cttggcagta catctacgta ttagtcatcg ctattaccat ggtgatgcgg ttttggcagt
      541 acatcaatgg gcgtggatag cggtttgact cacggggatt tccaagtctc caccccattg
      601 acgtcaatgg gagtttgttt tggcaccaaa atcaacggga ctttccaaaa tgtcgtaaca
      661 actccgcccc attgacgcaa atgggcggta ggcgtgtacg gtgggaggtc tatataagca
      721 gagctcgttt agtgaaccgt cagatcgcct ggagacgcca tccacgctgt tttgacctcc
      781 atagaagaca ccgggaccga tccagcctcc ggtcgaccga tcctgagaac ttcagggtga
      841 gtttggggac ccttgattgt tctttctttt tcgctattgt aaaattcatg ttatatggag
      901 ggggcaaagt tttcagggtg ttgtttagaa tgggaagatg tcccttgtat caccatggac
      961 cctcatgata attttgtttc tttcactttc tactctgttg acaaccattg tctcctctta
     1021 ttttcttttc attttctgta actttttcgt taaactttag cttgcatttg taacgaattt
     1081 ttaaattcac ttttgtttat ttgtcagatt gtaagtactt tctctaatca cttttttttc
     1141 aaggcaatca gggtatatta tattgtactt cagcacagtt ttagagaaca attgttataa
     1201 ttaaatgata aggtagaata tttctgcata taaattctgg ctggcgtgga aatattctta
     1261 ttggtagaaa caactacacc ctggtcatca tcctgccttt ctctttatgg ttacaatgat
     1321 atacactgtt tgagatgagg ataaaatact ctgagtccaa accgggcccc tctgctaacc
     1381 atgttcatgc cttcttctct ttcctacagc tcctgggcaa cgtgctggtt gttgtgctgt
     1441 ctcatcattt tggcaaagaa ttcctcgacg gatccctcga ggaattctga cactatgatc
     1501 ctctggcatg agatgtggca tgaaggcctg gaagaggcat ctcgtttgta ctttggggaa
     1561 aggaacgtga aaggcatgtt tgaggtgctg gagcccttgc atgctatgat ggaacggggc
     1621 ccccagactc tgaaggaaac atcctttaat caggcctatg gtcgagattt aatggaggcc
     1681 caagagtggt gcaggaagta catgaaatca gggaatgtca aggacctcac ccaagcctgg
     1741 gacctctatt atcatgtgtt ccgacgaatc tcaaagGGTG GTGGAGGCAG TGGCGGTGGA
     1801 GGCAGCagat tagataaaag taaagtgatt aacagcgcat tagagctgct taatgaggtc
     1861 ggaatcgaag gtttaacaac ccgtaaactc gcccagaagc taggtgtaga gcagcctaca
     1921 ttgtattggc atgtaaaaaa taagcgggct ttgctcgacg ccttagccat tgagatgtta
     1981 gataggcacc atactcactt ttgcccttta gaaggggaaa gctggcaaga ttttttacgt
     2041 aataacgcta aaagttttag atgtgcttta ctaagtcatc gcgatggagc aaaagtacat
     2101 ttaggtacac ggcctacaga aaaacagtat gaaactctcg aaaatcaatt agccttttta
     2161 tgccaacaag gtttttcact agagaatgca ttatatgcac tcagcgctgt ggggcatttt
     2221 actttaggtt gcgtattgga agatcaagag catcaagtcg ctaaagaaga aagggaaaca
     2281 cctactactg atagtatgcc gccattatta cgacaagcta tcgaattatt tgatcaccaa
     2341 ggtgcagagc cagccttctt attcggcctt gaattgatca tatgcggatt agaaaaacaa
     2401 cttaaatgtg aaagtgggtc cgcgtacagc cgcgcgcgta cgaaaaacaa ttacgggtct
     2461 accatcgagg gcctgctcga tctcccggac gacgacgccc ccgaagaggc ggggctggcg
     2521 gctccgcgcc tgtcctttct ccccgcggga cacacgcgca gactgtcgac ggcccccccg
     2581 accgatgtca gcctggggga cgagctccac ttagacggcg aggacgtggc gatggcgcat
     2641 gccgacgcgc tagacgattt cgatctggac atgttggggg acggggattc cccgggtccg
     2701 ggatttaccc cccacgactc cgccccctac ggcgctctgg atatggccga cttcgagttt
     2761 gagcagatgt ttaccgatgc ccttggaatt gacgagtacg gtggggaaca gaaactgatt
     2821 agcgaagaag atctgtaact caaatcctgc acaacagatt cttcatgttt ggaccaaatc
     2881 aacttgtgat accatgctca aagaggcctc aattatattt gagtttttaa tttttatgaa
     2941 aaaaaaaaaa aaaaacggaa ttcctcgagg gatccgtcga ggaattcact cctcaggtgc
     3001 aggctgccta tcagaaggtg gtggctggtg tggccaatgc cctggctcac aaataccact
     3061 gagatctttt tccctctgcc aaaaattatg gggacatcat gaagcccctt gagcatctga
     3121 cttctggcta ataaaggaaa tttattttca ttgcaatagt gtgttggaat tttttgtgtc
     3181 tctcactcgg aaggacatat gggagggcaa atcatttaaa acatcagaat gagtatttgg
     3241 tttagagttt ggcaacatat gcccatatgc tggctgccat gaacaaaggt tggctataaa
     3301 gaggtcatca gtatatgaaa cagccccctg ctgtccattc cttattccat agaaaagcct
     3361 tgacttgagg ttagattttt tttatatttt gttttgtgtt atttttttct ttaacatccc
     3421 taaaattttc cttacatgtt ttactagcca gatttttcct cctctcctga ctactcccag
     3481 tcatagctgt ccctcttctc ttatggagat ccctcgacgg atcggccgca attcgtaatc
     3541 atgtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca caacatacga
     3601 gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact cacattaatt
     3661 gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct gcattaatga
     3721 atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc
     3781 actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg
     3841 gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc
     3901 cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc
     3961 ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga
     4021 ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc
     4081 ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat
     4141 agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg
     4201 cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc
     4261 aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga
     4321 gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact
     4381 agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt
     4441 ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag
     4501 cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg
     4561 tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa
     4621 aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata
     4681 tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg
     4741 atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata
     4801 cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg
     4861 gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct
     4921 gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt
     4981 tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc
     5041 tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga
     5101 tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt
     5161 aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc
     5221 atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa
     5281 tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca
     5341 catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca
     5401 aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct
     5461 tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc
     5521 gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa
     5581 tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt
     5641 tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctaaattg
     5701 taagcgttaa tattttgtta aaattcgcgt taaatttttg ttaaatcagc tcatttttta
     5761 accaataggc cgaaatcggc aaaatccctt ataaatcaaa agaatagacc gagatagggt
     5821 tgagtgttgt tccagtttgg aacaagagtc cactattaaa gaacgtggac tccaacgtca
     5881 aagggcgaaa aaccgtctat cagggcgatg gcccactacg tgaaccatca ccctaatcaa
     5941 gttttttggg gtcgaggtgc cgtaaagcac taaatcggaa ccctaaaggg agcccccgat
     6001 ttagagcttg acggggaaag ccggcgaacg tggcgagaaa ggaagggaag aaagcgaaag
     6061 gagcgggcgc tagggcgctg gcaagtgtag cggtcacgct gcgcgtaacc accacacccg
     6121 ccgcgcttaa tgcgccgcta cagggcgcgt cccattcgcc attcaggctg cgcaactgtt
     6181 gggaagggcg atcggtgcgg gcctcttcgc tattacgcca gctggcgaaa gggggatgtg
     6241 ctgcaaggcg attaagttgg gtaacgccag ggttttccca gtcacgacgt tgtaaaacga
     6301 cggccagtga gcgcg    
//
