LOCUS       Exported                7052 bp ds-DNA     circular SYN 21-DEC-2021
DEFINITION  For expression of METTL3 (methyltransferase-like 3) in human cells.
ACCESSION   .
VERSION     .
KEYWORDS    pcDNA3/Flag-METTL3
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7052)
  AUTHORS   Liu J, Yue Y, Han D, Wang X, Fu Y, Zhang L, Jia G, Yu M, Lu Z, Deng 
            X, Dai Q, Chen W, He C
  TITLE     A METTL3-METTL14 complex mediates mammalian nuclear RNA N6-adenosine
            methylation.
  JOURNAL   Nat Chem Biol. 2014 Feb;10(2):93-5. doi: 10.1038/nchembio.1432. Epub
            2013 Dec 6.
  PUBMED    24316715
REFERENCE   2  (bases 1 to 7052)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1038/nchembio.1432"; journalName: "Nat Chem Biol"; date: 
            "2014-02"; volume: "10"; issue: "2"; pages: "93-5"
FEATURES             Location/Qualifiers
     source          1..7052
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     primer_bind     23..44
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(163..183)
                     /label=pBABE 3
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        207..417
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     promoter        229..425
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      276..411
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     338..357
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             492..1286
                     /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(546..565)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     1156..1175
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    1460..1581
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1497..1516)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1551..1570
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(1630..1646)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1630..1646)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1643..1665)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1654..1670
                     /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(1678..1708)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1723..1744
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1861..1878)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2032..2620)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2112..2131)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(2791..3651)
                     /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     3414..3433
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3652..3756)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(3831..3850)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        4022..4401
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        4402..4605
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     4556..4576
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     4650..4669
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        4650..4668
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             4685..4708
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     promoter        complement(6464..6482)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(6465..6482)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(6502..6519)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6508..6732
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      join(6778..7052,1..154)
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6865..6884)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"