LOCUS       sequence-404698-        8585 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Lentiviral conditional expression of mitochondrial-targeted eGFP in 
            mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8585)
  AUTHORS   Michaelis JB, Bozkurt S, Schafer JA, Munch C
  TITLE     Monitoring Mitochondrial Protein Import Using Mitochondrial 
            Targeting Sequence (MTS)-eGFP.
  JOURNAL   Bio Protoc. 2022 Dec 20;12(24):e4578. doi: 10.21769/BioProtoc.4578. 
            eCollection 2022 Dec 20.
  PUBMED    36618095
REFERENCE   2  (bases 1 to 8585)
  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: "Bio 
            Protoc."; date: "2022-12-20"; volume: "12(24):e4578. doi"; pages: " 
            10.21769/BioProtoc.4578. eCollection 2022 Dec 20"
COMMENT     Sequence Label: pLD-puro-MTS-EGFP
FEATURES             Location/Qualifiers
     source          1..8585
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    470..703
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             888..932
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
     misc_feature    1145..1262
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        1316..1630
                     /label=tight TRE promoter
                     /note="Tet-responsive promoter PTight, consisting of seven 
                     tet operator sequences followed by the minimal CMV 
                     promoter"
     protein_bind    1324..1342
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1360..1378
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1395..1413
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1431..1449
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1467..1485
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1502..1520
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1538..1556
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     1603..1627
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     regulatory      1861..1870
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1867..1869,1870..1872,1873..2586)
                     /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             join(1867..1869,1870..1872,1873..2583)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1912..1933)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2173..2192)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2520..2541
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             2604..2621
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     promoter        2631..3141
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     3060..3078
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     CDS             3151..3747
                     /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(3151..3170)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     3647..3667
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     CDS             3748..3801
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             join(3802..4419,4420..4425,4426..4461,4462..4464,4465..4500,
                     4501..4503,4504..4539,4540..4548)
                     /codon_start=1
                     /product="improved tetracycline-controlled transactivator"
                     /label=rtTA-Advanced
                     /note="In the Tet-On(R) system, rtTA-Advanced binds to the 
                     Tet-responsive element and stimulates transcription only in
                     the presence of tetracycline or doxycycline."
                     /translation="MSRLDKSKVINGALELLNGVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALPIEMLDRHHTHFCPLEGESWQDFLRNNAKSYRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEEQEHQVAKEERETPTT
                     DSMPPLLRQAIELFDRQGAEPAFLFGLELIICGLEKQLKCESGGPTDALDDFDLDMLPA
                     DALDDFDLDMLPADALDDFDLDMLPG"
     primer_bind     complement(4588..4608)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(5006..5017)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             5222..5455
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    5527..5648
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5564..5583)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5618..5637
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      5688..5823
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     promoter        5823..5894
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5895..5963,5964..6755)
                     /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     complement(6113..6132)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6926..7514
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7415..7434
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7668..7685
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    7802..7823
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(7838..7843,7844..7861,7862..7868)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7876..7892
                     /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)."
     primer_bind     7881..7903
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     7900..7916
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7900..7916
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     7935..7955
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        7937..7955
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        7983..8209
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             8210..8390
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    8437..8562
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
ORIGIN
        1 atcgcgatgg gaaaaaattc ggttaaggcc agggggaaag aaaaaatata aattaaaaca
       61 tatagtatgg gcaagcaggg agctagaacg attcgcagtt aatcctggcc tgttagaaac
      121 atcagaaggc tgtagacaaa tactgggaca gctacaacca tcccttcaga caggatcaga
      181 agaacttaga tcattatata atacagtagc aaccctctat tgtgtgcatc aaaggataga
      241 gataaaagac accaaggaag ctttagacaa gatagaggaa gagcaaaaca aaagtaagac
      301 caccgcacag caagcggccg ctgatcttca gacctggagg aggagatatg agggacaatt
      361 ggagaagtga attatataaa tataaagtag taaaaattga accattagga gtagcaccca
      421 ccaaggcaaa gagaagagtg gtgcagagag aaaaaagagc agtgggaata ggagctttgt
      481 tccttgggtt cttgggagca gcaggaagca ctatgggcgc agcgtcaatg acgctgacgg
      541 tacaggccag acaattattg tctggtatag tgcagcagca gaacaatttg ctgagggcta
      601 ttgaggcgca acagcatctg ttgcaactca cagtctgggg catcaagcag ctccaggcaa
      661 gaatcctggc tgtggaaaga tacctaaagg atcaacagct cctggggatt tggggttgct
      721 ctggaaaact catttgcacc actgctgtgc cttggaatgc tagttggagt aataaatctc
      781 tggaacagat ttggaatcac acgacctgga tggagtggga cagagaaatt aacaattaca
      841 caagcttaat acactcctta attgaagaat cgcaaaacca gcaagaaaag aatgaacaag
      901 aattattgga attagataaa tgggcaagtt tgtggaattg gtttaacata acaaattggc
      961 tgtggtatat aaaattattc ataatgatag taggaggctt ggtaggttta agaatagttt
     1021 ttgctgtact ttctatagtg aatagagtta ggcagggata ttcaccatta tcgtttcaga
     1081 cccacctccc aaccccgagg ggacaattct cgacctcgag acaaatggca gtattcatcc
     1141 acaattttaa aagaaaaggg gggattgggg ggtacagtgc aggggaaaga atagtagaca
     1201 taatagcaac agacatacaa actaaagaat tacaaaaaca aattacaaaa attcaaaatt
     1261 ttcgggttta ttacagggac agcagagatc cactttggcc gcgaatcgat atgtcgagtt
     1321 tactccctat cagtgataga gaacgtatgt cgagtttact ccctatcagt gatagagaac
     1381 gatgtcgagt ttactcccta tcagtgatag agaacgtatg tcgagtttac tccctatcag
     1441 tgatagagaa cgtatgtcga gtttactccc tatcagtgat agagaacgta tgtcgagttt
     1501 atccctatca gtgatagaga acgtatgtcg agtttactcc ctatcagtga tagagaacgt
     1561 atgtcgaggt aggcgtgtac ggtgggaggc ctatataagc agagctcgtt tagtgaaccg
     1621 tcagatcgcc tggagaattg gaccatggcc tccactcgtg tcctcgcctc tcgcctggcc
     1681 tcccggatgg ctgcttccgc caaggttgcc cgccctgctg tccgcgttgc tcaggtcagc
     1741 aagcgcacca tccagactgg ctcccccctc cagaccctca agcgcaccca gatgacctcc
     1801 atcgtcaacg ccaccacccg ccaggctttc cagaagcgcg cctactcttc cggatccatc
     1861 gccaccatgg tgagcaaggg cgaggagctg ttcaccgggg tggtgcccat cctggtcgag
     1921 ctggacggcg acgtaaacgg ccacaagttc agcgtgtccg gcgagggcga gggcgatgcc
     1981 acctacggca agctgaccct gaagttcatc tgcaccaccg gcaagctgcc cgtgccctgg
     2041 cccaccctcg tgaccaccct gacctacggc gtgcagtgct tcagccgcta ccccgaccac
     2101 atgaagcagc acgacttctt caagtccgcc atgcccgaag gctacgtcca ggagcgcacc
     2161 atcttcttca aggacgacgg caactacaag acccgcgccg aggtgaagtt cgagggcgac
     2221 accctggtga accgcatcga gctgaagggc atcgacttca aggaggacgg caacatcctg
     2281 gggcacaagc tggagtacaa ctacaacagc cacaacgtct atatcatggc cgacaagcag
     2341 aagaacggca tcaaggtgaa cttcaagatc cgccacaaca tcgaggacgg cagcgtgcag
     2401 ctcgccgacc actaccagca gaacaccccc atcggcgacg gccccgtgct gctgcccgac
     2461 aaccactacc tgagcaccca gtccgccctg agcaaagacc ccaacgagaa gcgcgatcac
     2521 atggtcctgc tggagttcgt gaccgccgcc gggatcactc tcggcatgga cgagctgtac
     2581 aagtaattag taatgaaccg gtccaccacc accaccacca ctaaggatcc ggggttgggg
     2641 ttgcgccttt tccaaggcag ccctgggttt gcgcagggac gcggctgctc tgggcgtggt
     2701 tccgggaaac gcagcggcgc cgaccctggg tctcgcacat tcttcacgtc cgttcgcagc
     2761 gtcacccgga tcttcgccgc tacccttgtg ggccccccgg cgacgcttcc tgctccgccc
     2821 ctaagtcggg aaggttcctt gcggttcgcg gcgtgccgga cgtgacaaac ggaagccgca
     2881 cgtctcacta gtaccctcgc agacggacag cgccagggag caatggcagc gcgccgaccg
     2941 cgatgggctg tggccaatag cggctgctca gcagggcgcg ccgagagcag cggccgggaa
     3001 ggggcggtgc gggaggcggg gtgtggggcg gtagtgtggg ccctgttcct gcccgcgcgg
     3061 tgttccgcat tctgcaagcc tccggagcgc acgtcggcag tcggctccct cgttgaccga
     3121 atcaccgacc tctctcccca gcaattcacc atgaccgagt acaagcccac ggtgcgcctc
     3181 gccacccgcg acgacgtccc cagggccgta cgcaccctcg ccgccgcgtt cgccgactac
     3241 cccgccacgc gccacaccgt cgatccggac cgccacatcg agcgggtcac cgagctgcaa
     3301 gaactcttcc tcacgcgcgt cgggctcgac atcggcaagg tgtgggtcgc ggacgacggc
     3361 gccgcggtgg cggtctggac cacgccggag agcgtcgaag cgggggcggt gttcgccgag
     3421 atcggcccgc gcatggccga gttgagcggt tcccggctgg ccgcgcagca acagatggaa
     3481 ggcctcctgg cgccgcaccg gcccaaggag cccgcgtggt tcctggccac cgtcggcgtc
     3541 tcgcccgacc accagggcaa gggtctgggc agcgccgtcg tgctccccgg agtggaggcg
     3601 gccgagcgcg ccggggtgcc cgccttcctg gagacctccg cgccccgcaa cctccccttc
     3661 tacgagcggc tcggcttcac cgtcaccgcc gacgtcgagg tgcccgaagg accgcgcacc
     3721 tggtgcatga cccgcaagcc cggtgccgaa ggtagaggtt ctctcctcac ttgtggtgat
     3781 gttgaagaaa accctggtcc aatgtctaga ctggacaaga gcaaagtcat aaacggagct
     3841 ctggaattac tcaatggtgt cggtatcgaa ggcctgacga caaggaaact cgctcaaaag
     3901 ctgggagttg agcagcctac cctgtactgg cacgtgaaga acaagcgggc cctgctcgat
     3961 gccctgccaa tcgagatgct ggacaggcat catacccact tctgccccct ggaaggcgag
     4021 tcatggcaag actttctgcg gaacaacgcc aagtcatacc gctgtgctct cctctcacat
     4081 cgcgacgggg ctaaagtgca tctcggcacc cgcccaacag agaaacagta cgaaaccctg
     4141 gaaaatcagc tcgcgttcct gtgtcagcaa ggcttctccc tggagaacgc actgtacgct
     4201 ctgtccgccg tgggccactt tacactgggc tgcgtattgg aggaacagga gcatcaagta
     4261 gcaaaagagg aaagagagac acctaccacc gattctatgc ccccacttct gagacaagca
     4321 attgagctgt tcgaccggca gggagccgaa cctgccttcc ttttcggcct ggaactaatc
     4381 atatgtggcc tggagaaaca gctaaagtgc gaaagcggcg ggccgaccga cgcccttgac
     4441 gattttgact tagacatgct cccagccgat gcccttgacg actttgacct tgatatgctg
     4501 cctgctgacg ctcttgacga ttttgacctt gacatgctcc ccgggtaact aagtaaggat
     4561 cgatccaaga tatcgtattc ttaactatgt tgctcctttt acgctatgtg gatacgctgc
     4621 tttaatgcct ttgtatcatg ctattgcttc ccgtatggct ttcattttct cctccttgta
     4681 taaatcctgg ttgctgtctc tttatgagga gttgtggccc gttgtcaggc aacgtggcgt
     4741 ggtgtgcact gtgtttgctg acgcaacccc cactggttgg ggcattgcca ccacctgtca
     4801 gctcctttcc gggactttcg ctttccccct ccctattgcc acggcggaac tcatcgccgc
     4861 ctgccttgcc cgctgctgga caggggctcg gctgttgggc actgacaatt ccgtggtgtt
     4921 gtcggggaag ctgacgtcct ttccatggct gctcgcctgt gttgccacct ggattctgcg
     4981 cgggacgtcc ttctgctacg tcccttcggc cctcaatcca gcggaccttc cttcccgcgg
     5041 cctgctgccg gctctgcggc ctcttccgcg tcttcgcctt cgccctcaga cgagtcggat
     5101 ctccctttgg gccgcctccc cgcctgtttc gcctcggcgt ccggactaga ggtaccttta
     5161 agaccaatga cttacaaggc agctgtagat cttagccact ttttaaaaga aaagggggga
     5221 ctggaagggc taattcactc ccaacgaaga caagatctgc tttttgcttg tactgggtct
     5281 ctctggttag accagatctg agcctgggag ctctctggct aactagggaa cccactgctt
     5341 aagcctcaat aaagcttgcc ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac
     5401 tctggtaact agagatccct cagacccttt tagtcagtgt ggaaaatctc tagcagtagt
     5461 agttcatgtc atcttattat tcagtattta taacttgcaa agaaatgaat atcagagagt
     5521 gagaggaact tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat
     5581 ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat
     5641 gtatcttatc atgtctggct ctagctatcc cgcccctaac tccgcccatc ccgcccctaa
     5701 ctccgcccag ttccgcccat tctccgcccc atggctgact aatttttttt atttatgcag
     5761 aggccgaggc cgcctcggcc tctgagctat tccagaagta gtgaggaggc ttttttggag
     5821 gcctttcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt
     5881 gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg
     5941 cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag
     6001 atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt aagatccttg
     6061 agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg
     6121 gcgcggtatt atcccgtatt gacgccgggc aagagcaact cggtcgccgc atacactatt
     6181 ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg gatggcatga
     6241 cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac
     6301 ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc
     6361 atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc
     6421 gtgacaccac gatgcctgta gcaatggcaa caacgttgcg caaactatta actggcgaac
     6481 tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat aaagttgcag
     6541 gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg
     6601 gtgagcgtgg gtctcgcggt atcattgcag cactggggcc agatggtaag ccctcccgta
     6661 tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg
     6721 ctgagatagg tgcctcactg attaagcatt ggtaactgtc agaccaagtt tactcatata
     6781 tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt
     6841 ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc
     6901 ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct
     6961 tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa
     7021 ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact gttcttctag
     7081 tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc
     7141 tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg
     7201 actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca
     7261 cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat
     7321 gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg
     7381 tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc
     7441 ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc
     7501 ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc
     7561 cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg
     7621 cctttgagtg agctgatacc gctcgccgca gccgaacgac cgagcgcagc gagtcagtga
     7681 gcgaggaagc ggaagagcgc ccaatacgca aaccgcctct ccccgcgcgt tggccgattc
     7741 attaatgcag ctggcacgac aggtttcccg actggaaagc gggcagtgag cgcaacgcaa
     7801 ttaatgtgag ttagctcact cattaggcac cccaggcttt acactttatg cttccggctc
     7861 gtatgttgtg tggaattgtg agcggataac aatttcacac aggaaacagc tatgaccatg
     7921 attacgccaa gcgcgcaatt aaccctcact aaagggaaca aaagctggag ctgcaagctt
     7981 aatgtagtct tatgcaatac tcttgtagtc ttgcaacatg gtaacgatga gttagcaaca
     8041 tgccttacaa ggagagaaaa agcaccgtgc atgccgattg gtggaagtaa ggtggtacga
     8101 tcgtgcctta ttaggaaggc aacagacggg tctgacatgg attggacgaa ccactgaatt
     8161 gccgcattgc agagatattg tatttaagtg cctagctcga tacataaacg ggtctctctg
     8221 gttagaccag atctgagcct gggagctctc tggctaacta gggaacccac tgcttaagcc
     8281 tcaataaagc ttgccttgag tgcttcaagt agtgtgtgcc cgtctgttgt gtgactctgg
     8341 taactagaga tccctcagac ccttttagtc agtgtggaaa atctctagca gtggcgcccg
     8401 aacagggact tgaaagcgaa agggaaacca gaggagctct ctcgacgcag gactcggctt
     8461 gctgaagcgc gcacggcaag aggcgagggg cggcgactgg tgagtacgcc aaaaattttg
     8521 actagcggag gctagaagga gagagatggg tgcgagagcg tcagtattaa gcgggggaga
     8581 attag
//
