LOCUS       Exported                8743 bp ds-DNA     circular SYN 13-MAY-2021
DEFINITION  Express  in MT1-MMP mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    pLPCX2-MT1-MMP-eGFP
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8743)
  AUTHORS   Sun J, Lu F, He H, Shen J, Messina J, Mathew R, Wang D, Sarnaik AA, 
            Chang WC, Kim M, Cheng H, Yang S
  TITLE     STIM1- and Orai1-mediated Ca(2+) oscillation orchestrates 
            invadopodium formation and melanoma invasion.
  JOURNAL   J Cell Biol. 2014 Nov 24;207(4):535-48. doi: 10.1083/jcb.201407082. 
            Epub 2014 Nov 17.
  PUBMED    25404747
REFERENCE   2  (bases 1 to 8743)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1083/jcb.201407082"; journalName: "J Cell Biol"; date: 
            "2014-11-24- 24"; volume: "207"; issue: "4"; pages: "535-48"
FEATURES             Location/Qualifiers
     source          1..8743
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     enhancer        1..304
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        305..508
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     459..479
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             complement(1150..1161)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     regulatory      2353..2362
                     /regulatory_class="other"
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             2359..3078
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             2359..3075
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2404..2425)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2665..2684)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3012..3033
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     LTR             3144..3737
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     primer_bind     complement(3840..3862)
                     /label=pGEX 3
                     /note="pGEX vectors, reverse primer"
     protein_bind    complement(3990..4023)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     protein_bind    complement(4083..4116)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     primer_bind     4136..4155
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4236..4253)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4407..4995)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4487..4506)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(5166..6026)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     5789..5808
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6027..6131)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6199..6217
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     LTR             6819..6994
                     /label=5 LTR (truncated)
                     /note="truncated long terminal repeat from Moloney murine 
                     sarcoma virus"
     misc_feature    7057..7256
                     /label=MMLV Psi
                     /note="packaging signal of Moloney murine leukemia virus 
                     (MMLV)"
     CDS             7457..7873
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     7803..7825
                     /label=pLXSN 5
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     7841..7857
                     /label=pBABE 5
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     CDS             7970..8569
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(7970..7989)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     8466..8486
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"