LOCUS       sequence-62604-e        8645 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8645)
  AUTHORS   Gascoigne KE, Takeuchi K, Suzuki A, Hori T, Fukagawa T, Cheeseman IM
  TITLE     Induced ectopic kinetochore assembly bypasses the requirement for 
            CENP-A nucleosomes.
  JOURNAL   Cell. 2011 Apr 29;145(3):410-22. doi: 10.1016/j.cell.2011.03.031.
  PUBMED    21529714
REFERENCE   2  (bases 1 to 8645)
  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.cell.2011.03"; journalName: "Cell"; date: "2011-04-29- 
            29"; volume: "145"; issue: "3"; pages: "410-22"
COMMENT     Sequence Label: pKG209
FEATURES             Location/Qualifiers
     source          1..8645
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     LTR             8..477
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     misc_feature    542..741
                     /label=MMLV Psi
                     /note="packaging signal of Moloney murine leukemia virus 
                     (MMLV)"
     CDS             934..1350
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     1318..1334
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     regulatory      1385..1394
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1391..1393,1394..1396,1397..2107)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(1391..1393,1394..1396,1397..2107)
                     /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(1436..1457)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1697..1716)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2044..2065
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             2153..2173
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             2186..2230
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     primer_bind     complement(3432..3451)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             4493..4513
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     intron          4675..4740
                     /label=small t intron
                     /note="SV40 (simian virus 40) small t antigen intron"
     primer_bind     complement(5060..5080)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        5065..5394
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5245..5380
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     promoter        5409..5456
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             5475..5873
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     LTR             5926..6519
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     primer_bind     complement(6720..6737)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6891..7479)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6971..6990)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7657..8448,8449..8517))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     8280..8299
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8518..8622)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ctgcagcctg aatatgggcc aaacaggata tctgtggtaa gcagttcctg ccccggctca
       61 gggccaagaa cagatggaac agctgaatat gggccaaaca ggatatctgt ggtaagcagt
      121 tcctgccccg gctcagggcc aagaacagat ggtccccaga tgcggtccag ccctcagcag
      181 tttctagaga accatcagat gtttccaggg tgccccaagg acctgaaatg accctgtgcc
      241 ttatttgaac taaccaatca gttcgcttct cgcttctgtt cgcgcgcttc tgctccccga
      301 gctcaataaa agagcccaca acccctcact cggggcgcca gtcctccgat tgactgagtc
      361 gcccgggtac ccgtgtatcc aataaaccct cttgcagttg catccgactt gtggtctcgc
      421 tgttccttgg gagggtctcc tctgagtgat tgactacccg tcagcggggg tctttcattt
      481 gggggctcgt ccgggatcgg gagacccctg cccagggacc accgacccac caccgggagg
      541 taagctggcc agcaacttat ctgtgtctgt ccgattgtct agtgtctatg actgatttta
      601 tgcgcctgcg tcggtactag ttagctaact agctctgtat ctggcggacc cgtggtggaa
      661 ctgacgagtt ctgaacaccc ggccgcaacc ctgggagacg tcccagggac tttgggggcc
      721 gtttttgtgg cccgacctga ggaagggagt cgatgtggaa tccgaccccg tcaggatatg
      781 tggttctggt aggagacgag aacctaaaac agttcccgcc tccgtctgaa tttttgcttt
      841 cggtttggaa ccgaagccgc gcgtcttgtc tgctgcagca tcgttctgtg ttgtctctgt
      901 ctgactgtgt ttctgtattt gtctgaaaat tagggccaga ctgttaccac tcccttaagt
      961 ttgaccttag atcactggaa agatgtcgag cggctcgctc acaaccagtc ggtagatgtc
     1021 aagaagagac gttgggttac cttctgctct gcagaatggc caacctttaa cgtcggatgg
     1081 ccgcgagacg gcacctttaa ccgagacctc atcacccagg ttaagatcaa ggtcttttca
     1141 cctggcccgc atggacaccc agaccaggtc ccctacatcg tgacctggga agccttggct
     1201 tttgaccccc ctccctgggt caagcccttt gtacacccta agcctccgcc tcctcttctt
     1261 ccatccgcgc cgtctctccc ccttgaacct cctctttcga ccccgcctca atcctccctt
     1321 tatccagccc tcactccttc tctaggcgcc ggccggatcc cagtgtggtg gtacgctacc
     1381 ggtcgccacc atggtgagca agggcgagga gctgttcacc ggggtggtgc ccatcctggt
     1441 cgagctggac ggcgacgtaa acggccacaa gttcagcgtg tccggcgagg gcgagggcga
     1501 tgccacctac ggcaagctga ccctgaagtt catctgcacc accggcaagc tgcccgtgcc
     1561 ctggcccacc ctcgtgacca ccctgaccta cggcgtgcag tgcttcagcc gctaccccga
     1621 ccacatgaag cagcacgact tcttcaagtc cgccatgccc gaaggctacg tccaggagcg
     1681 caccatcttc ttcaaggacg acggcaacta caagacccgc gccgaggtga agttcgaggg
     1741 cgacaccctg gtgaaccgca tcgagctgaa gggcatcgac ttcaaggagg acggcaacat
     1801 cctggggcac aagctggagt acaactacaa cagccacaac gtctatatca tggccgacaa
     1861 gcagaagaac ggcatcaagg tgaacttcaa gatccgccac aacatcgagg acggcagcgt
     1921 gcagctcgcc gaccactacc agcagaacac ccccatcggc gacggccccg tgctgctgcc
     1981 cgacaaccac tacctgagca cccagtccgc cctgagcaaa gaccccaacg agaagcgcga
     2041 tcacatggtc ctgctggagt tcgtgaccgc cgccgggatc actctcggca tggacgagct
     2101 gtacaagtcc ggaggggatc ggtggagctc caccggtggc ggccgctcta gagagaatct
     2161 ttattttcag ggcgccgcca aattcaaaga aaccgctgct gctaaattcg aacgccagca
     2221 catggacagc ggaggtggag gtactagtct cgagatggca gaccacaacc ctgacagcga
     2281 ttctacacct cgtacactgc tgcgtcgtgt gctggatact gctgatcctc gtactcctcg
     2341 tcgtcctcgt agcgctcgtg ctggtgctcg ccgtgccctg ctggaaacag cctctcctcg
     2401 taaactgagt ggccagaccc gtactattgc ccgtgggcgt tcacatggag cccgctcggt
     2461 cggtcgtagt gctcacattc aagcctcggg acacctggaa gaacaaaccc ctcgtaccct
     2521 gctgaaaaac atcctgctga ctgcccctga aagctctatt ctgatgccgg aaagcgtcgt
     2581 taaaccggtt cctgctcctc aagccgttca accttctcgt caagagtcct cctgtggatc
     2641 tctggagctg caactgcctg aactggagcc tcctacaaca ctggcaccgg ggctgctggc
     2701 gcctggtcgt cgtaaacaac gtctgcgtct gagcgtgttt caacaaggtg ttgatcaggg
     2761 cctgagtctg tctcaagaac ctcaaggcaa cgctgatgct tctagtctga cccgttctct
     2821 gaacctgacc tttgctactc cactgcaacc tcaaagcgtc caacgtcctg gactggctcg
     2881 ccgccctcct gctcgccgtg ccgttgatgt gggtgctttt ctgcgtgacc tgcgtgatac
     2941 aagcctggca cctccgaata ttgtgctgga ggatacacag ccgttttcac agcctatggt
     3001 tggtagccct aacgtttatc actcactgcc gtgtacccct cacactggtg ccgaagatgc
     3061 tgaacaagct gctggtcgta aaacccaaag ctctggaccg ggactgcaaa aaaactctcc
     3121 ggggaaacct gcccaattcc tggcgggtga agctgaagaa gtcaacgcct ttgctctggg
     3181 atttctgtct accagctcgg gtgttagcgg agaagatgag gtggaacctc tgcatgatgg
     3241 tgtggaagaa gccgagaaga aaatggaaga ggaaggggtg tctgtctctg aaatggaagc
     3301 gactggtgct caaggaccaa gtcgtgtcga agaggcagaa ggccataccg aggttaccga
     3361 agccgaagga agccaagggg aattcccgcg gggaagtgga agtggatcca tggtgaaacc
     3421 agtaacgtta tacgatgtcg cagagtatgc cggtgtctct tatcagaccg tttcccgcgt
     3481 ggtgaaccag gccagccacg tttctgcgaa aacgcgggaa aaagtggaag cggcgatggc
     3541 ggagctgaat tacattccca accgcgtggc acaacaactg gcgggcaaac agtcgttgct
     3601 gattggcgtt gccacctcca gtctggccct gcacgcgccg tcgcaaattg tcgcggcgat
     3661 taaatctcgc gccgatcaac tgggtgccag cgtggtggtg tcgatggtag aacgaagcgg
     3721 cgtcgaagcc tgtaaagcgg cggtgcacaa tcttctcgcg caacgcgtca gtgggctgat
     3781 cattaactat ccgctggatg accaggatgc cattgctgtg gaagctgcct gcactaatgt
     3841 tccggcgtta tttcttgatg tctctgacca gacacccatc aacagtatta ttttctccca
     3901 tgaagacggt acgcgactgg gcgtggagca tctggtcgca ttgggtcacc agcaaatcgc
     3961 gctgttagcg ggcccattaa gttctgtctc ggcgcgtctg cgtctggctg gctggcataa
     4021 atatctcact cgcaatcaaa ttcagccgat agcggaacgg gaaggcgact ggagtgccat
     4081 gtccggtttt caacaaacca tgcaaatgct gaatgagggc atcgttccca ctgcgatgct
     4141 ggttgccaac gatcagatgg cgctgggcgc aatgcgcgcc attaccgagt ccgggctgcg
     4201 cgttggtgcg gatatctcgg tagtgggata cgacgatacc gaagacagct catgttatat
     4261 cccgccgtta accaccatca aacaggattt tcgcctgctg gggcaaacca gcgtggaccg
     4321 cttgctgcaa ctctctcagg gccaggcggt gaagggcaat cagctgttgc ccgtctcact
     4381 ggtgaaaaga aaaaccaccc tggcgcccaa tacgcaaacc gcctctcccc gcgcgttggc
     4441 cgattcatta atgcagctgg cacgacaggt ttcccgaagc agcctgaggc ctcctaagaa
     4501 gaagaggaag gtttgagcgc aacgcaatta atgtaagtta gctcactcat taggcacccc
     4561 aggctttaca ctttatgctt ccgacctgca gccccttgga tctttgtgaa ggaaccttac
     4621 ttctgtggtg tgacataatt ggacaaacta cctacagaga tttaaagctc taaggtaaat
     4681 ataaaatttt taagtgtata atgtgttaaa ctactgattc taattgtttg tgtattttag
     4741 atcacagtcc caaggctcat ttcaggcccc tcagtcctca cagtctgttc atgatcataa
     4801 tcagccatac cacatttgta gaggttttac ttgctttaaa aaacctccca cacctccccc
     4861 tgaacctgaa acataaaatg aatgcaattc tgcagtcgac ggtaccgcgg gcccgggatc
     4921 caccggatct agataactga tcataatcag ccataccaca tttgtagagg ttttacttgc
     4981 tttaaaaaac ctcccacacc tccccctgaa cctgaaacat aaaatgaatg caattcgcca
     5041 gcacagtggt cgaccctgtg gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc
     5101 ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccag gtgtggaaag
     5161 tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc
     5221 atagtcccgc ccctaactcc gcccatcccg cccctaactc cgcccagttc cgcccattct
     5281 ccgccccatg gctgactaat tttttttatt tatgcagagg ccgaggccgc ctcggcctct
     5341 gagctattcc agaagtagtg aggaggcttt tttggaggcc taggcttttg caaaaagctt
     5401 gagcacgtgt tgacaattaa tcatcggcat agtatatcgg catagtataa tacgacaagg
     5461 tgaggaacta aaccatggcc aagcctttgt ctcaagaaga atccaccctc attgaaagag
     5521 caacggctac aatcaacagc atccccatct ctgaagacta cagcgtcgcc agcgcagctc
     5581 tctctagcga cggccgcatc ttcactggtg tcaatgtata tcattttact gggggacctt
     5641 gtgcagaact cgtggtgctg ggcactgctg ctgctgcggc agctggcaac ctgacttgta
     5701 tcgtcgcgat cggaaatgag aacaggggca tcttgagccc ctgcggacgg tgccgacagg
     5761 tgcttctcga tctgcatcct gggatcaaag ccatagtgaa ggacagtgat ggacagccga
     5821 cggcagttgg gattcgtgaa ttgctgccct ctggttatgt gtgggagggc taagcacttc
     5881 atcgataaaa taaaagattt tatttagtct ccagaaaaag gggggaatga aagaccccac
     5941 ctgtaggttt ggcaagctag cttaagtaac gccattttgc aaggcatgga aaaatacata
     6001 actgagaata gagaagttca gatcaaggtc aggaacagat ggaacagctg aatatgggcc
     6061 aaacaggata tctgtggtaa gcagttcctg ccccggctca gggccaagaa cagatggaac
     6121 agctgaatat gggccaaaca ggatatctgt ggtaagcagt tcctgccccg gctcagggcc
     6181 aagaacagat ggtccccaga tgcggtccag ccctcagcag tttctagaga accatcagat
     6241 gtttccaggg tgccccaagg acctgaaatg accctgtgcc ttatttgaac taaccaatca
     6301 gttcgcttct cgcttctgtt cgcgcgcttc tgctccccga gctcaataaa agagcccaca
     6361 acccctcact cggggcgcca gtcctccgat tgactgagtc gcccgggtac ccgtgtatcc
     6421 aataaaccct cttgcagttg catccgactt gtggtctcgc tgttccttgg gagggtctcc
     6481 tctgagtgat tgactacccg tcagcggggg tctttcattt ccgacttgtg gtctcgctgc
     6541 cttgggaggg tctcctctga gtgattgact acccgtcagc gggggtcttc acatgcagca
     6601 tgtatcaaaa ttaatttggt tttttttctt aagtatttac attaaatggc catagttgca
     6661 ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attggcgctc ttccgcttcc
     6721 tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca
     6781 aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca
     6841 aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg
     6901 ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg
     6961 acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt
     7021 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt
     7081 tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc
     7141 tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt
     7201 gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt
     7261 agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc
     7321 tacactagaa ggacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa
     7381 agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt
     7441 tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct
     7501 acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta
     7561 tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagtttgcg gccgcaaatc
     7621 aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc
     7681 acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta
     7741 gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga
     7801 cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg
     7861 cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc
     7921 tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat
     7981 cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag
     8041 gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat
     8101 cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa
     8161 ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa
     8221 gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caacacggga
     8281 taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg
     8341 gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc
     8401 acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg
     8461 aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact
     8521 cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat
     8581 atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt
     8641 gccac
//
