LOCUS       Exported                6181 bp ds-DNA     circular SYN 13-MAY-2021
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    pcDNA3.1 EYFP H148Q/I152L
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6181)
  AUTHORS   Galietta LJ, Haggie PM, Verkman AS
  TITLE     Green fluorescent protein-based halide indicators with improved 
            chloride and iodide affinities.
  JOURNAL   FEBS Lett. 2001 Jun 22. 499(3):220-4.
  PUBMED    11423120
REFERENCE   2  (bases 1 to 6181)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "FEBS Lett. 
            2001 Jun 22. 499(3):220-4."
FEATURES             Location/Qualifiers
     source          1..6181
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..584
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     629..648
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        629..647
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      726..735
                     /regulatory_class="other"
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             732..1451
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSQNVYLMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(777..798)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1038..1057)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1385..1406
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(1538..1555)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1544..1768
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1814..2242
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1901..1920)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2111..2132
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2251..2271)
                     /label=pBABE 3
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2256..2585
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2436..2571
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2498..2517
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2652..3446
                     /codon_start=1
                     /gene="aph(3)-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(2706..2725)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3316..3335
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3620..3741
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3657..3676)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3711..3730
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3790..3806)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3790..3806)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3803..3825)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3814..3830
                     /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(3838..3868)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3883..3904
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4021..4038)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4192..4780)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4272..4291)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(4951..5811)
                     /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     5574..5593
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5812..5916)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(5991..6010)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"