LOCUS       Exported                8104 bp ds-DNA     circular SYN 13-MAY-2021
DEFINITION  To introduce MS2 coat protein fused Tet1-CD.
ACCESSION   .
VERSION     .
KEYWORDS    pcDNA3.1-MS2-Tet1-CD
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8104)
  AUTHORS   Xu X, Tao Y, Gao X, Zhang L, Li X, Zou W, Ruan K, Wang F, Xu GL, Hu 
            R
  TITLE     A CRISPR-based approach for targeted DNA demethylation.
  JOURNAL   Cell Discov. 2016 May 3;2:16009. doi: 10.1038/celldisc.2016.9. 
            eCollection 2016.
  PUBMED    27462456
REFERENCE   2  (bases 1 to 8104)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Cell 
            Discov."; date: "2016-05-3"; volume: "2:16009. doi"; pages: " 
            10.1038/celldisc.2016.9. eCollection 2016"
FEATURES             Location/Qualifiers
     source          1..8104
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             926..1312
                     /codon_start=1
                     /product="bacteriophage MS2 coat protein"
                     /label=MS2-N55K
                     /note="binds with high affinity to a specific stem-loop 
                     structure in the viral RNA (Lim et al., 1994)"
                     /translation="ASNFTQFVLVDNGGTGDVTVAPSNFANGVAEWISSNSRSQAYKVT
                     CSVRQSSAQKRKYTIKVEVPKVATQTVGGVELPVAAWRSYLNMELTIPIFATNSDCELI
                     VKAMQGLLKDGNPIPSAIAANSGIY"
     CDS             1364..1384
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             3461..3484
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     primer_bind     complement(3525..3542)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3531..3755
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3801..4229
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3888..3907)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4098..4119
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4238..4258)
                     /label=pBABE 3
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4243..4573
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4424..4559
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4486..4505
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4622..5647
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     polyA_signal    5777..5898
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5814..5833)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5868..5887
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5947..5963)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5947..5963)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5960..5982)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5971..5987
                     /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(5995..6025)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6040..6061
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6178..6195)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6349..6937)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6429..6448)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(7108..7968)
                     /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     7731..7750
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7969..8073)
                     /gene="bla"
                     /label=AmpR promoter