LOCUS       Exported                3494 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  Expresses 3Xflag::tagRFP::myc in bacteria.
ACCESSION   .
VERSION     .
KEYWORDS    AP575-1
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3494)
  AUTHORS   Paix A, Folkmann A, Rasoloson D, Seydoux G
  TITLE     High Efficiency, Homology-Directed Genome Editing in Caenorhabditis 
            elegans Using CRISPR-Cas9 Ribonucleoprotein Complexes.
  JOURNAL   Genetics. 2015 Sep;201(1):47-54. doi: 10.1534/genetics.115.179382. 
            Epub 2015 Jul 17.
  PUBMED    26187122
REFERENCE   2  (bases 1 to 3494)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..3494
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     primer_bind     complement(29..51)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     151..170
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     364..386
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     378..395
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     379..395
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             422..487
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             488..1195
                     /codon_start=1
                     /product="monomeric derivative of red fluorescent protein 
                     from Entacmaea quadricolor (Merzlyak et al., 2007)"
                     /label=TagRFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELIKENMHMKLYMEGTVNNHHFKCTSEGEGKPYEGTQTMR
                     IKVVEGGPLPFAFDILATSFMYGSRTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGV
                     LTATQDTSLQDGCLIYNVKIRGVNFPSNGPVMQKKTLGWEANTEMLYPADGGLEGRSDM
                     ALKLVGGGHLICNFKTTYRSKKPAKNLKMPGVYYVDHRLERIKEADKETYVEQHEVAVA
                     RYCDLPSKLGHKLN"
     CDS             1196..1225
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     primer_bind     complement(1273..1289)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1273..1289)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1286..1308)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1297..1313
                     /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(1321..1351)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1366..1387
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1504..1521)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1675..2263)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1755..1774)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(2434..3294)
                     /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     3057..3076
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3295..3399)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3467..3485
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca
       61 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
      121 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
      181 accatatgcg gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc atcaggcgcc
      241 attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat
      301 tacgccagct ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta acgccagggt
      361 tttcccagtc acgacgttgt aaaacgacgg ccagtgaatt cgagctcggt acccggggat
      421 cgactacaaa gaccatgacg gtgattataa agatcatgat atcgattaca aggatgacga
      481 tgacaaggtg tctaagggcg aagagctgat taaggagaac atgcacatga agctgtacat
      541 ggagggcacc gtgaacaacc accacttcaa atgcacatcc gagggcgaag gcaagcccta
      601 cgagggcacc cagaccatga gaatcaaggt ggtcgagggc ggccctctcc ccttcgcctt
      661 tgacatcctg gctaccagct tcatgtacgg cagcagaacc ttcatcaacc acacccaggg
      721 catccccgac ttctttaagc agtcctttcc tgagggcttc acatgggaga gagtcaccac
      781 atacgaagac gggggcgtgc tgaccgctac ccaggacacc agcctccagg acggctgtct
      841 catctacaac gtcaagatca gaggggtgaa cttcccatcc aacggccctg tgatgcagaa
      901 gaaaacactc ggctgggagg ccaataccga gatgctgtac cccgctgacg gcggcctgga
      961 aggcagaagc gacatggccc tgaagctcgt gggcgggggc cacctgatct gcaatttcaa
     1021 gaccacatac agatccaaga aacccgctaa gaacctcaag atgcccggcg tctactatgt
     1081 ggaccacaga ctggaaagaa tcaaagaggc cgacaaagag acctacgtcg agcagcacga
     1141 ggtggctgtg gccagatact gcgacctccc tagcaaactg gggcacaaac ttaatgaaca
     1201 aaaactgata tcagaagagg atctggatcc tctagagtcg acctgcaggc atgcaagctt
     1261 ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca
     1321 caacatacga gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact
     1381 cacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct
     1441 gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc
     1501 ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca
     1561 ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg
     1621 agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca
     1681 taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa
     1741 cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc
     1801 tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc
     1861 gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct
     1921 gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg
     1981 tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag
     2041 gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta
     2101 cggctacact agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg
     2161 aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt
     2221 tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt
     2281 ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag
     2341 attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat
     2401 ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc
     2461 tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat
     2521 aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc
     2581 acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag
     2641 aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag
     2701 agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt
     2761 ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg
     2821 agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt
     2881 tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc
     2941 tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc
     3001 attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa
     3061 taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg
     3121 aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc
     3181 caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag
     3241 gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt
     3301 cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt
     3361 tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc
     3421 acctgacgtc taagaaacca ttattatcat gacattaacc tataaaaata ggcgtatcac
     3481 gaggcccttt cgtc
//
