LOCUS       sequence-173223-        5112 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  N-terminal PCR tagging cassette for yeast codon optimized split 
            GFP1-10 with CDC42 promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5112)
  AUTHORS   Smoyer CJ, Katta SS, Gardner JM, Stoltz L, McCroskey S, Bradford WD,
            McClain M, Smith SE, Slaughter BD, Unruh JR, Jaspersen SL
  TITLE     Analysis of membrane proteins localizing to the inner nuclear 
            envelope in living cells.
  JOURNAL   J Cell Biol. 2016 Nov 21;215(4):575-590. Epub 2016 Nov 9.
  PUBMED    27831485
REFERENCE   2  (bases 1 to 5112)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "J Cell 
            Biol."; date: "2016-11-21"; volume: "215(4)"; pages: "575-590. Epub 
            2016 Nov 9"
COMMENT     Sequence Label: pS1643 (pFA6-NATMX-CDC42pr-yGFP1-10)
FEATURES             Location/Qualifiers
     source          1..5112
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(103..207)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     275..293
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(331..353)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     453..472
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        553..571
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     553..570
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     gene            684..1803
                     /label=natMX6
                     /note="yeast selectable marker conferring nourseothricin 
                     resistance"
     promoter        684..1027
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             1028..1600
                     /codon_start=1
                     /gene="Streptomyces noursei nat1"
                     /product="nourseothricin acetyltransferase"
                     /label=NrsR
                     /note="confers resistance to nourseothricin"
                     /translation="MGTTLDDTAYRYRTSVPGDAEAIEALDGSFTTDTVFRVTATGDGF
                     TLREVPVDPPLTKVFPDDESDDESDDGEDGDPDSRTFVAYGDDGDLAGFVVVSYSGWNR
                     RLTVEDIEVAPEHRGHGVGRALMGLATEFARERGAGHLWLEVTNVNAPAIHAYRRMGFT
                     LCGLDTALYDGTASDGEQALYMSMPCP"
     terminator      1606..1803
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     primer_bind     complement(3318..3337)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3319..3337)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3424..3441)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3595..4183)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3675..3694)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4354..5112,1..33,34..102))
                     /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     4977..4996
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc
       61 aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc tttttcaata
      121 ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg aatgtattta
      181 gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac ctgacgtcta
      241 agaaaccatt attatcatga cattaaccta taaaaatagg cgtatcacga ggccctttcg
      301 tctcgcgcgt ttcggtgatg acggtgaaaa cctctgacac atgcagctcc cggagacggt
      361 cacagcttgt ctgtaagcgg atgccgggag cagacaagcc cgtcagggcg cgtcagcggg
      421 tgttggcggg tgtcggggct ggcttaacta tgcggcatca gagcagattg tactgagagt
      481 gcaccatatg gacatattgt cgttagaacg cggctacaat taatacataa ccttatgtat
      541 catacacata cgatttaggt gacactatag aacgcggccg ccagctgaag cttcgtacgc
      601 tgcaggtcga cggatccccg ggttaattaa ggcgcgccag atctgtttag cttgcctcgt
      661 ccccgccggg tcacccggcc agcgacatgg aggcccagaa taccctcctt gacagtcttg
      721 acgtgcgcag ctcaggggca tgatgtgact gtcgcccgta catttagccc atacatcccc
      781 atgtataatc atttgcatcc atacattttg atggccgcac ggcgcgaagc aaaaattacg
      841 gctcctcgct gcagacctgc gagcagggaa acgctcccct cacagacgcg ttgaattgtc
      901 cccacgccgc gcccctgtag agaaatataa aaggttagga tttgccactg aggttcttct
      961 ttcatatact tccttttaaa atcttgctag gatacagttc tcacatcaca tccgaacata
     1021 aacaaccatg ggtaccactc ttgacgacac ggcttaccgg taccgcacca gtgtcccggg
     1081 ggacgccgag gccatcgagg cactggatgg gtccttcacc accgacaccg tcttccgcgt
     1141 caccgccacc ggggacggct tcaccctgcg ggaggtgccg gtggacccgc ccctgaccaa
     1201 ggtgttcccc gacgacgaat cggacgacga atcggacgac ggggaggacg gcgacccgga
     1261 ctcccggacg ttcgtcgcgt acggggacga cggcgacctg gcgggcttcg tggtcgtctc
     1321 gtactccggc tggaaccgcc ggctgaccgt cgaggacatc gaggtcgccc cggagcaccg
     1381 ggggcacggg gtcgggcgcg cgttgatggg gctcgcgacg gagttcgccc gcgagcgggg
     1441 cgccgggcac ctctggctgg aggtcaccaa cgtcaacgca ccggcgatcc acgcgtaccg
     1501 gcggatgggg ttcaccctct gcggcctgga caccgccctg tacgacggca ccgcctcgga
     1561 cggcgagcag gcgctctaca tgagcatgcc ctgcccctaa tcagtactga caataaaaag
     1621 attcttgttt tcaagaactt gtcatttgta tagttttttt atattgtagt tgttctattt
     1681 taatcaaatg ttagcgtgat ttatattttt tttcgcctcg acatcatctg cccagatgcg
     1741 aagttaagtg cgcagaaagt aatatcatgc gtcaatcgta tgtgaatgct ggtcgctata
     1801 ctgctgtcga ttcgatacta acgccgccat ccagtgtcga aaacgagctc gccgctctag
     1861 acaggccgga actcaaaagg gtaatttcgt gaaaaacaat catctacggt ataaataaca
     1921 atttaattta cgtctctttc gaaaatgctc gtgtagctca gtggttagag cttcgtgctt
     1981 atagcaacat tcggtttccg aagtttctgt gccaaagacc tttcaaacag gcctttaaaa
     2041 gcaacgcgac cgtcgtgggt tcaatcccca cctcgagcac tttctctttt ttttttaaaa
     2101 aaagttgcat tatttctatc aactgttgta gctacgttac acctaatttt gtattgtaga
     2161 ctgttagata atccacatat acaggtaaat acggcacatt gtcttctaaa catcgcatta
     2221 aagatgtctt ccaccgtcga ttcaagggtc attcaatacg ggtaatactg cttccattgg
     2281 gcccttccaa acgaaacaag atacgggaaa aagtgataaa aaaaagcaaa caaggttgat
     2341 tgatcgatga tagacgcgat tcttggaaaa ggcaagagca gcgttttgat aaataggttc
     2401 caggtgtacc gagatatact gattatcctt ctctgtcatt cttcactttt tgcaataggt
     2461 ttcctttgtc gcaaaaaatt atattcgtta tttattatac tattctattt tcctgaggag
     2521 ataggttaac aaacgaatta gagaagcaaa acttataaaa caagaaataa acgtattagc
     2581 tcttccacaa aggatccatg tctaaaggtg aagaattgtt tactggtgtt gttccaattt
     2641 tggttgaatt ggatggtgat gttaatggtc ataaattttc tgttagaggt gaaggtgaag
     2701 gtgatgctac tattggtaaa ttgactttga aatttatttg tactactggt aaattgccag
     2761 ttccatggcc aactttggtt actactttga cttatggtgt tcaatgtttt tctagatatc
     2821 cagatcatat gaaacaacat gattttttta aatctgctat gccagaaggt tatgttcaag
     2881 aaagaactat ttcttttaaa gatgatggta aatataaaac tagagctgtt gttaaatttg
     2941 aaggtgatac tttggttaat agaattgaat tgaaaggtac tgattttaaa gaagatggta
     3001 atattttggg tcataaattg gaatataatt ttaattctca taatgtttat attactgctg
     3061 ataaacaaaa gaatggtatt aaagctaatt ttactgttag acataatgtt gaagatggtt
     3121 ctgttcaatt ggctgatcat tatcaacaaa atactccaat tggtgatggt ccagttttgt
     3181 tgccagataa tcattatttg tctactcaaa ctgttttgtc taaagatcca aatgaaaaag
     3241 gtggtggttc tggtggtggt tctcatcgat gatatcagat ccactagtgg cctatgcggc
     3301 cgcggatctg ccggtctccc tatagtgagt cgtattaatt tcgataagcc aggttaacct
     3361 gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc
     3421 ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca
     3481 ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg
     3541 agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca
     3601 taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa
     3661 cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc
     3721 tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc
     3781 gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct
     3841 gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg
     3901 tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag
     3961 gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta
     4021 cggctacact agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg
     4081 aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt
     4141 tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt
     4201 ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag
     4261 attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat
     4321 ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc
     4381 tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat
     4441 aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc
     4501 acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag
     4561 aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag
     4621 agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt
     4681 ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg
     4741 agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt
     4801 tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc
     4861 tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc
     4921 attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa
     4981 taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg
     5041 aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc
     5101 caactgatct tc
//
