LOCUS       sequence-270304-        6601 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Retroviral expression of GFP-Ska1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6601)
  AUTHORS   Monda JK, Whitney IP, Tarasovetc EV, Wilson-Kubalek E, Milligan RA, 
            Grishchuk EL, Cheeseman IM
  TITLE     Microtubule Tip Tracking by the Spindle and Kinetochore Protein Ska1
            Requires Diverse Tubulin-Interacting Surfaces.
  JOURNAL   Curr Biol. 2017 Dec 4;27(23):3666-3675.e6. doi: 
            10.1016/j.cub.2017.10.018. Epub 2017 Nov 16.
  PUBMED    29153323
REFERENCE   2  (bases 1 to 6601)
  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"; doi: 
            "10.1016/j.cub.2017.10.018"; journalName: "Curr Biol"; date: 
            "2017-12-4- 4"; volume: "27"; issue: "23"; pages: "3666-3675.e6"
COMMENT     Sequence Label: pIW082
FEATURES             Location/Qualifiers
     source          1..6601
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             111..527
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     457..479
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     495..511
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     regulatory      562..571
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(568..570,571..573,574..1284)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(568..570,571..573,574..1284)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(613..634)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(874..893)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1221..1242
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             1330..1350
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             1363..1407
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     primer_bind     complement(2355..2375)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2360..2689
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2540..2675
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             2697..3095
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     LTR             3264..3733
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     primer_bind     complement(3863..3880)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4034..4622)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4114..4133)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4782..5573,5574..5642))
                     /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     5405..5424
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5643..5747)
                     /gene="bla"
                     /label=AmpR promoter
     LTR             5778..6247
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     misc_feature    6312..6511
                     /label=MMLV Psi
                     /note="packaging signal of Moloney murine leukemia virus 
                     (MMLV)"
ORIGIN
        1 ttttgctttc ggtttggaac cgaagccgcg cgtcttgtct gctgcagcgc tgcagcatcg
       61 ttctgtgttg tctctgtctg actgtgtttc tgtatttgtc tgaaaattag ggccagactg
      121 ttaccactcc cttaagtttg accttaggtc actggaaaga tgtcgagcgg atcgctcaca
      181 accagtcggt agatgtcaag aagagacgtt gggttacctt ctgctctgca gaatggccaa
      241 cctttaacgt cggatggccg cgagacggca cctttaaccg agacctcatc acccaggtta
      301 agatcaaggt cttttcacct ggcccgcatg gacacccaga ccaggtcccc tacatcgtga
      361 cctgggaagc cttggctttt gacccccctc cctgggtcaa gccctttgta caccctaagc
      421 ctccgcctcc tcttcctcca tccgccccgt ctctccccct tgaacctcct cgttcgaccc
      481 cgcctcgatc ctccctttat ccagccctca ctccttctct aggcgccggc cggatcccag
      541 tgtggtggta cgctaccggt cgccaccatg gtgagcaagg gcgaggagct gttcaccggg
      601 gtggtgccca tcctggtcga gctggacggc gacgtaaacg gccacaagtt cagcgtgtcc
      661 ggcgagggcg agggcgatgc cacctacggc aagctgaccc tgaagttcat ctgcaccacc
      721 ggcaagctgc ccgtgccctg gcccaccctc gtgaccaccc tgacctacgg cgtgcagtgc
      781 ttcagccgct accccgacca catgaagcag cacgacttct tcaagtccgc catgcccgaa
      841 ggctacgtcc aggagcgcac catcttcttc aaggacgacg gcaactacaa gacccgcgcc
      901 gaggtgaagt tcgagggcga caccctggtg aaccgcatcg agctgaaggg catcgacttc
      961 aaggaggacg gcaacatcct ggggcacaag ctggagtaca actacaacag ccacaacgtc
     1021 tatatcatgg ccgacaagca gaagaacggc atcaaggtga acttcaagat ccgccacaac
     1081 atcgaggacg gcagcgtgca gctcgccgac cactaccagc agaacacccc catcggcgac
     1141 ggccccgtgc tgctgcccga caaccactac ctgagcaccc agtccgccct gagcaaagac
     1201 cccaacgaga agcgcgatca catggtcctg ctggagttcg tgaccgccgc cgggatcact
     1261 ctcggcatgg acgagctgta caagtccgga ggggatcggt ggagctccac cggtggcggc
     1321 cgctctagag agaatcttta ttttcagggc gccgccaaat tcaaagaaac cgctgctgct
     1381 aaattcgaac gccagcacat ggacagcgga ggtggaggta ctagtctcga gatggcaagc
     1441 agcgacctgg agcagctgtg ctctcatgtt aatgaaaaga ttggcaatat taagaaaacc
     1501 ttatcattaa gaaactgtgg acaagagccc acactgaaga ccgtattaaa taaaatagga
     1561 gatgagatca ttgtaataaa tgaacttcta aataaattgg aattggaaat tcagtatcaa
     1621 gaacaaacca acaattcact caaggaactc tgtgaatctc ttgaagaaga ttacaaagac
     1681 atagaacatc ttaaagaaaa cgttccttcc catttgcctc aagtaacagt aacccagagc
     1741 tgtgttaagg gatcagatct tgatcctgaa gaaccaatca aagttgaaga acctgaaccc
     1801 gtaaagaagc ctcccaaaga gcaaagaagt attaaggaaa tgccatttat aacttgtgat
     1861 gagtttaacg gggtgccaag ttacatgaaa tcgcgactga catacaacca gattaatgat
     1921 gttattaaag aaatcaacaa ggcagtaatt agtaaatata aaatcctaca tcagccaaaa
     1981 aagtctatga actctgtgac cagaaatctc tatcacagat ttattgatga agaaacgaag
     2041 gataccaaag gtcgttattt catcgtagag gcagatatta aggagttcac aactttgaaa
     2101 gctgacaaga agtttcacgt gttactgaat attttagccc actgccggag gctatcagag
     2161 gtccgagggg gaggtttaac acgatacgta ataacctgac cgcgggcccg ggatccaccg
     2221 gatctagata actgatcata atcagccata ccacatttgt agaggtttta cttgctttaa
     2281 aaaacctccc acacctcccc ctgaacctga aacataaaat gaatgcaatt cgccagcaca
     2341 gtggtcgacc ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag gctccccagc
     2401 aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc
     2461 aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccatagt
     2521 cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc attctccgcc
     2581 ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctcgg cctctgagct
     2641 attccagaag tagtgaggag gcttttttgg aggcctaggc ttttgcaaaa agcttcatgg
     2701 ccaagccttt gtctcaagaa gaatccaccc tcattgaaag agcaacggct acaatcaaca
     2761 gcatccccat ctctgaagac tacagcgtcg ccagcgcagc tctctctagc gacggccgca
     2821 tcttcactgg tgtcaatgta tatcatttta ctgggggacc ttgtgcagaa ctcgtggtgc
     2881 tgggcactgc tgctgctgcg gcagctggca acctgacttg tatcgtcgcg atcggaaatg
     2941 agaacagggg catcttgagc ccctgcggac ggtgccgaca ggtgcttctc gatctgcatc
     3001 ctgggatcaa agccatagtg aaggacagtg atggacagcc gacggcagtt gggattcgtg
     3061 aattgctgcc ctctggttat gtgtgggagg gctaagatcg ataaaataaa agattttatt
     3121 tagtctccag aaaaaggggg gaatgaaaga ccccacctgt aggtttggca agctagctta
     3181 agtaacgcca ttttgcaagg catggaaaaa tacataactg agaatagaga agttcagatc
     3241 aaggtcagga acagatggaa cagctgaata tgggccaaac aggatatctg tggtaagcag
     3301 ttcctgcccc ggctcagggc caagaacaga tggaacagct gaatatgggc caaacaggat
     3361 atctgtggta agcagttcct gccccggctc agggccaaga acagatggtc cccagatgcg
     3421 gtccagccct cagcagtttc tagagaacca tcagatgttt ccagggtgcc ccaaggacct
     3481 gaaatgaccc tgtgccttat ttgaactaac caatcagttc gcttctcgct tctgttcgcg
     3541 cgcttctgct ccccgagctc aataaaagag cccacaaccc ctcactcggg gcgccagtcc
     3601 tccgattgac tgagtcgccc gggtacccgt gtatccaata aaccctcttg cagttgcatc
     3661 cgacttgtgg tctcgctgtt ccttgggagg gtctcctctg agtgattgac tacccgtcag
     3721 cgggggtctt tcacatgcag catgtatcaa aattaatttg gttttttttc ttaagtattt
     3781 acattaaatg gccatagttg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc
     3841 gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc
     3901 ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata
     3961 acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg
     4021 cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct
     4081 caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa
     4141 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc
     4201 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt
     4261 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg
     4321 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg
     4381 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct
     4441 tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc
     4501 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg
     4561 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc
     4621 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt
     4681 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttgcggccgc
     4741 aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt
     4801 gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc
     4861 gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg
     4921 cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc
     4981 gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg
     5041 gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca
     5101 ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga
     5161 tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct
     5221 ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg
     5281 cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca
     5341 accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata
     5401 cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct
     5461 tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact
     5521 cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa
     5581 acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc
     5641 atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga
     5701 tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga
     5761 aaagtgccac ctgcagcctg aatatgggcc aaacaggata tctgtggtaa gcagttcctg
     5821 ccccggctca gggccaagaa cagatggaac agctgaatat gggccaaaca ggatatctgt
     5881 ggtaagcagt tcctgccccg gctcagggcc aagaacagat ggtccccaga tgcggtccag
     5941 ccctcagcag tttctagaga accatcagat gtttccaggg tgccccaagg acctgaaatg
     6001 accctgtgcc ttatttgaac taaccaatca gttcgcttct cgcttctgtt cgcgcgcttc
     6061 tgctccccga gctcaataaa agagcccaca acccctcact cggggcgcca gtcctccgat
     6121 tgactgagtc gcccgggtac ccgtgtatcc aataaaccct cttgcagttg catccgactt
     6181 gtggtctcgc tgttccttgg gagggtctcc tctgagtgat tgactacccg tcagcggggg
     6241 tctttcattt gggggctcgt ccgggatcgg gagacccctg cccagggacc accgacccac
     6301 caccgggagg caagctggcc agcaacttat ctgtgtctgt ccgattgtct agtgtctatg
     6361 actgatttta tgcgcctgcg tcggtactag ttagctaact agctctgtat ctggcggacc
     6421 cgtggtggaa ctgacgagtt ctgaacaccc ggccgcaacc ctgggagacg tcccagggac
     6481 tttgggggcc gtttttgtgg cccgacctga ggaagggagt cgatgtggaa tccgaccccg
     6541 tcaggatatg tggttctggt aggagacgag aacctaaaac agttcccgcc tccgtctgaa
     6601 t
//
