The complete nucleotide sequence for transposon Tn5

Paul N Hengen pnh at fcsparc6.ncifcrf.gov
Mon May 16 16:46:43 EST 1994


    ****** Complete nucleotide sequence of transposon Tn5 *******

Transposon Tn5 is a 5.8 kb composite mobile genetic element encoding kanamycin,
bleomycin, and streptomycin resistance (for a review, see reference 1).
Despite wide use of Tn5 for the genetic study of many species of Gram-negative
bacteria, and that the nucleotide sequence of the entire genome of Tn5 has been
known since 1985, the correct completed nucleotide sequence entry has never
been added to GenBank.  Conflicting data and unmerged sequences have made
retrieval and use of the sequence difficult for molecular biologists.  Upon
accessing the entries related to Tn5 from GenBank, one must reconstruct the
genome of the transposon from those fragments which have been sent to the
database. This is a formidable task for those unfamiliar with the vast
published material on Tn5.  Moreover, Tn5 sequences derived from unidentified
sources have been passed from scientist to scientist and these contain sequence
and feature errors. The compilation of the correct DNA sequence of Tn5 is
therefore of immense value to the scientific community.

I have submitted a merged version of the entire DNA sequence of Tn5 and the
relevant biological features to the GenBank database. However, nearly a year
has passed and this entry has not yet been made available to biologists.  They
are therefore faced with the burden of collecting and merging all the sequence
data individually.

The Tn5 sequence data has been merged as follows:

Nucleotide bases 1 through 1400 are those reported for the left inverted repeat
[accession no. V00615] (2). This was chosen as the merge point to avoid a
missing "C" at Tn5 position 1684 which was later corrected (3).

Bases 1401 through 2300 are those reported for the kanamycin resistance
segment [accession nos. V00618, J01834] (3) and bases 2301 through 4218 are
from the central portion encoding bleomycin and streptomycin resistance
[accession no. X01702] (4).

Bases 4219 through 5818 are from the complement of those reported for the
terminal end of the right inverted repeat up to the overlap with the central
portion [accession no.  V00617] (2).

References:

1. Berg, D. E. (1989) In Berg, D. E., and Howe, M. (eds.),
   Mobile DNA, ASM Press, Washington, D.C., pp. 185-210.

2. Auerswald, E. A., Ludwig, G., and Schaller, H. (1981)
   Cold Spring Harbor Symposia on Quantitative Biology 45, 107-113.

3. Beck, E., Ludwig, G., Auerswald, E. A., Reiss, B., and Schaller, H.
   (1982) Gene 19, 327-336.
 
4. Mazodier, P., Cossart, P., Giraud, E., and Gasser, F.
   (1985) Nucleic Acids Research 13, 195-205.

================================================================================

LOCUS       TN5          5818 bp DNA    BCT             08-JUL-1993
DEFINITION  Complete sequence of E. coli transposon Tn5
ACCESSION   U00004
KEYWORDS    transposon Tn5; aminoglycoside-3'-O-phosphotransferase;
            kanamycin resistance; neomycin resistance;
            bleomycin resistance; streptomycin resistance;
            streptomycin phosphotransferase;
            transposase; transposition inhibitor.
SOURCE      Escherichia coli DNA.
  ORGANISM  Escherichia coli
            Prokaryota; Bacteria; Gracilicutes; Scotobacteria; Facultatively
            anaerobic rods; Enterobacteriaceae.
REFERENCE   1  (bases 1 to 1400; 4219 to 5818)
  AUTHORS   Auerswald, E. A., Ludwig, G., and Schaller, H.
  TITLE     Structural analysis of Tn5
  JOURNAL   Cold Spring Harb. Symp. Quant. Biol. 45, 107-113
            (1981)
  STANDARD  full automatic
REFERENCE   2  (bases 1401 to 2300)
  AUTHORS   Beck, E., Ludwig, G., Auerswald, E. A., Reiss, B., and Schaller, H.
  TITLE     Nucleotide sequence and exact localization of the neomycin
            phosphotransferase gene from Tn5
  JOURNAL   Gene 19, 327-336 (1982)
  STANDARD  full automatic
REFERENCE   3  (bases 2301 to 4218)
  AUTHORS   Mazodier, P., Cossart, P., Giraud, E., and Gasser, F.
  TITLE     Completion of the nucleotide sequence of the central region of Tn5
            confirms the presence of three resistance genes
  JOURNAL   Nucleic Acids Research 13, 195-205 (1985)
  STANDARD  full automatic
REFERENCE   4
  AUTHORS   Johnson, R. C., Yin, J. C. P., and Reznikoff, W. S.
  TITLE     Control of Tn5 transposition in Escherichia coli is mediated by
            protein from the right repeat
  JOURNAL   Cell 30, 873-882 (1982)
  STANDARD  full automatic
REFERENCE   5
  AUTHORS   Rothstein, S. J., Jorgensen, R. A., Yin, J. C.-P., Yong-Di, Z.,
            Johnson, R. C., and Reznikoff, W. S.
  TITLE     Genetic organization of Tn5
  JOURNAL   Cold Spring Harb. Symp. Quant. Biol. 45, 99-105
            (1981)
  STANDARD  full automatic
REFERENCE   6
  AUTHORS   Rothstein, S. J., and Reznikoff, W. S.
  TITLE     The functional differences in the inverted repeats of Tn5 are
            caused by a single base pair nonhomology
  JOURNAL   Cell 23, 191-199 (1981)
  STANDARD  full automatic
REFERENCE   7
  AUTHORS   Johnson, R. C., and Reznikoff, W. S.
  TITLE     DNA sequences at the ends of transposon Tn5 required for
            transposition
  JOURNAL   Nature 304, 280-282 (1983)
  STANDARD  full automatic
REFERENCE   8
  AUTHORS   Berg, D. E.
  TITLE     Transposon Tn5
  JOURNAL   (in) Berg, D. E., and Howe, M. (Eds.);
            Mobile DNA: 185-210, American Society for Microbiology Press,
            Washington, D.C. (1989)
  STANDARD  full automatic
COMMENT
            These data kindly submitted in computer readable form by:

            Paul N. Hengen
            Laboratory of Mathematical Biology
            P.O. Box B
            National Cancer Institute
            Frederick Cancer Research and Development Center
            Frederick, Maryland 21702-1201 U.S.A.
            Phone:  301-846-5581
            Fax:    301-846-5598
            Email:  pnh at ncifcrf.gov

                    -and-

            Andrew R. Bradbury
            International School for Advanced Studies (SISSA)
            Via Beirut 2-4,
            Trieste 34013,
            Italy.
            Phone:  +39 40 398995
            Fax:    +39 40 398991
            Email:  bradbury at icgeb.trieste.it


FEATURES            LOCATION/QUALIFIERS
     promoter        1413..1462
                     /function="aminoglycoside phosphotransferase 
                     promoter region"
                     /evidence= EXPERIMENTAL
                     /citation=[5]
     mRNA            (1.80)..(1500.1550)
     mRNA            (1460.1540)..(3600.3650)
     mRNA            complement((4300.4350)..(5738.5818))
     CDS             92..1444
                     /codon_start=1
                     /function="non-functional"
                     /product="protein #3"
                     /evidence=EXPERIMENTAL
                     /citation=[4]
                     /note= "A Thymidine at position 1442 causes an ochre 
                     stop codon which prematurely terminates the protein
                     at this point. Ochre suppressing strains allow readthrough
                     and express a functional product identical to the transposase."
                     /translation="MITSALHRAADWAKSVFSSAALGDPRRTARLVNVAAQLAKYSGK
                     SITISSEGSEAMQEGAYRFYRNPNVSAEAIRKAGAMQTVKLAQEFPELLAIEDTTSLS
                     YRHQVAEELGKLGSIQDKSRGWWVHSVLLLEATTFRTVGLLHQEWWMRPDDPADADEK
                     ESGKWLAAAATSRLRMGSMMSNVIAVCDREADIHAYLQDRLAHNERFVVRSKHPRKDV
                     ESGLYLIDHLKNQPELGGYQISIPQKGVVDKRGKRKNRPARKASLSLRSGRITLKQGN
                     ITLNAVLAEEINPPKGETPLKWLLLTGEPVESLAQALRVIDIYTHRWRIEEFHKAWKT
                     GAGAERQRMEEPDNLERMVSILSFVAVRLLQLRESFTLPQALRAQGLLKEAEHVESQS
                     AETVLTPDECQLLGYLDKGKRKRKEKAGSLQWAYMAIARLGGFMDSKRTGIASWGALW
                     "
     CDS             257..1444
                     /codon_start= 1
                     /function="non-functional"
                     /product="protein #4"
                     /evidence=EXPERIMENTAL
                     /citation=[4]
                     /note= "A Thymidine at position 1442 causes an ochre 
                     stop codon which prematurely terminates the protein
                     at this point. Ochre suppressing strains allow readthrough
                     and express a functional product identical to the transposition
                     inhibitor."
                     /translation="MQEGAYRFYRNPNVSAEAIRKAGAMQTVKLAQEFPELLAIEDTTSLS
                     YRHQVAEELGKLGSIQDKSRGWWVHSVLLLEATTFRTVGLLHQEWWMRPDDPADADEK
                     ESGKWLAAAATSRLRMGSMMSNVIAVCDREADIHAYLQDRLAHNERFVVRSKHPRKDV
                     ESGLYLIDHLKNQPELGGYQISIPQKGVVDKRGKRKNRPARKASLSLRSGRITLKQGN
                     ITLNAVLAEEINPPKGETPLKWLLLTGEPVESLAQALRVIDIYTHRWRIEEFHKAWKT
                     GAGAERQRMEEPDNLERMVSILSFVAVRLLQLRESFTLPQALRAQGLLKEAEHVESQS
                     AETVLTPDECQLLGYLDKGKRKRKEKAGSLQWAYMAIARLGGFMDSKRTGIASWGALW
                     "
     CDS             1551..2345
                     /codon_start= 1
                     /function= "neomycin/kanamycin resistance"
                     /product= "aminoglycoside-3'-O-phosphotransferase"
                     /evidence= EXPERIMENTAL
                     /citation= ([2])
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGR
                     PVLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDL
                     LSSHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDE
                     EHQGLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRY
                     QDIALATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             2366..2746
                     /codon_start= 1
                     /function= "bleomycin resistance"
                     /product= "bleomycin resistance"
                     /evidence= EXPERIMENTAL
                     /citation= ([3])
                     /translation="MTDQATPNLPSRDFDSTAAFYERLGFGIVFRDAGWMILQRGDLM
                     LEFFAHPGLDPLASWFSCCLRLDDLAEFYRQCKSVGIQETSSGYPRIHAPELQEWGGT
                     MAALVDPDGTLLRLIQNELLAGIS"
     CDS             2785..3585
                     /codon_start= 1
                     /function= "streptomycin resistance"
                     /product= "streptomycin phosphotransferase"
                     /evidence= EXPERIMENTAL
                     /citation= ([3])
                     /note= "This protein confers streptomycin resistance in
                     some species of Gram-negative bacteria, but is cryptic in
                     Escherichia coli."
                     /translation="MERWRLLRDGELLTTHSSWILPVRQGDMPAMLKVARIPDEEAGY
                     RLLTWWDGQGAARVFASAAGALLMERASGAGDLAQIAWSGQDDEACRILCDTAARLHA
                     PRSGPPPDLHPLQEWFQPLFRLAAEHAALAPAASVARQLLAAPREVCPLHGDLHHENV
                     LDFGDRGWLAIDPHGLLGERTFDYANIFTNPDLSDPGRPLAILPGRLEARLSIVVATT
                     GFEPERLLRWIIAWTGLSAAWFIGDGDGEGEGAAIDLAVNAMARRLLD"
     CDS             complement(4297..5727)
                     /codon_start= 1
                     /function= "transposase"
                     /product= "protein #1"
                     /evidence= EXPERIMENTAL
                     /citation= ([4])
                     /translation="MITSALHRAADWAKSVFSSAALGDPRRTARLVNVAAQLAKYSGK
                     SITISSEGSEAMQEGAYRFYRNPNVSAEAIRKAGAMQTVKLAQEFPELLAIEDTTSLS
                     YRHQVAEELGKLGSIQDKSRGWWVHSVLLLEATTFRTVGLLHQEWWMRPDDPADADEK
                     ESGKWLAAAATSRLRMGSMMSNVIAVCDREADIHAYLQDRLAHNERFVVRSKHPRKDV
                     ESGLYLIDHLKNQPELGGYQISIPQKGVVDKRGKRKNRPARKASLSLRSGRITLKQGN
                     ITLNAVLAEEINPPKGETPLKWLLLTGEPVESLAQALRVIDIYTHRWRIEEFHKAWKT
                     GAGAERQRMEEPDNLERMVSILSFVAVRLLQLRESFTLPQALRAQGLLKEAEHVESQS
                     AETVLTPDECQLLGYLDKGKRKRKEKAGSLQWAYMAIARLGGFMDSKRTGIASWGALW
                     EGWEALQSKLDGFLAAKDLMAQGIKI"
     CDS             complement(4297..5562)
                     /codon_start= 1
                     /function= "transposition inhibitor"
                     /product= "protein #2"
                     /evidence= EXPERIMENTAL
                     /citation= ([4])
                     /translation="MQEGAYRFYRNPNVSAEAIRKAGAMQTVKLAQEFPELLAIEDTTSLS
                     YRHQVAEELGKLGSIQDKSRGWWVHSVLLLEATTFRTVGLLHQEWWMRPDDPADADEK
                     ESGKWLAAAATSRLRMGSMMSNVIAVCDREADIHAYLQDRLAHNERFVVRSKHPRKDV
                     ESGLYLIDHLKNQPELGGYQISIPQKGVVDKRGKRKNRPARKASLSLRSGRITLKQGN
                     ITLNAVLAEEINPPKGETPLKWLLLTGEPVESLAQALRVIDIYTHRWRIEEFHKAWKT
                     GAGAERQRMEEPDNLERMVSILSFVAVRLLQLRESFTLPQALRAQGLLKEAEHVESQS
                     AETVLTPDECQLLGYLDKGKRKRKEKAGSLQWAYMAIARLGGFMDSKRTGIASWGALW
                     EGWEALQSKLDGFLAAKDLMAQGIKI"
     conflict        1684
                     /note= "A missing 'C' in the sequence published by Auerswald
                     et al.(1981) is corrected within that published by Beck
                     et al. (1982)."
                     /citation= ([1],[2])
     mutation        1442
                     /note= "A single base pair difference exists as a 'T' at
                     position 1442 and a 'G' at position 4377 within the
                     inverted repeat regions of IS50L and IS50R. This mutation
                     renders IS50L inactive by prematurely terminating the
                     transposase."
                     /citation= ([6])
     repeat_region   1..1534
                     /function= "insertion sequence IS50L"
                     /rpt_type= inverted
                     /evidence= EXPERIMENTAL
                     /rpt_unit= 1 .. 1534
                     /rpt_unit= 4285 .. 5818
                     /standard_name= "IS50L"
                     /note="IS50L is non-functional due to mutation at 1442"
                     /citation= ([1])
     repeat_region   4285..5818
                     /function= "insertion sequence IS50R"
                     /rpt_type= inverted
                     /evidence= EXPERIMENTAL
                     /rpt_unit= 1 .. 1534
                     /rpt_unit= 4285 .. 5818
                     /standard_name= "IS50R"
                     /note="IS50R is functional in transposition"
                     /citation= ([1])
     repeat_region   order(1..19,5800..5818)
                     /function= "outside end"
                     /rpt_type= inverted
                     /evidence= EXPERIMENTAL
                     /rpt_unit= 1 .. 19
                     /rpt_unit= 5800 .. 5818
                     /note= "minimum length required for transposition"
                     /citation= ([7])
     repeat_region   order(1515..1533,4286..4304)
                     /function= "inside end"
                     /rpt_type= inverted
                     /evidence= EXPERIMENTAL
                     /rpt_unit= 1515 .. 1533
                     /rpt_unit= 4286 .. 4304
                     /note= "minimum length required for transposition"
                     /citation= ([7])
     protein_bind    8..16
                     /note= "putative DnaA box"
                     /citation= ([8])
     protein_bind    5803..5811
                     /note= "putative DnaA box"
                     /citation= ([8])
     protein_bind    1510..1533
                     /note= "putative IHF binding site"
                     /citation= ([8])
     protein_bind    4286..4309
                     /note= "putative IHF binding site"
                     /citation= ([8])

BASE COUNT     1125 a   1719 c   1777 g   1197 t 
ORIGIN
        1 ctgactctta tacacaagta gcgtcctgaa cggaaccttt cccgttttcc aggatctgac 
       61 ttccatgtga cctcctaaca tggtaacgtt catgataact tctgctcttc atcgtgcggc 
      121 cgactgggct aaatctgtgt tctcttcggc ggcgctgggt gatcctcgcc gtactgcccg 
      181 cttggttaac gtcgccgccc aattggcaaa atattctggt aaatcaataa ccatctcatc 
      241 agagggtagt gaagccatgc aggaaggcgc ttaccgattt taccgcaatc ccaacgtttc 
      301 tgccgaggcg atcagaaagg ctggcgccat gcaaacagtc aagttggctc aggagtttcc 
      361 cgaactgctg gccattgagg acaccacctc tttgagttat cgccaccagg tcgccgaaga 
      421 gcttggcaag ctgggctcta ttcaggataa atcccgcgga tggtgggttc actccgttct 
      481 cttgctcgag gccaccacat tccgcaccgt aggattactg catcaggagt ggtggatgcg 
      541 cccggatgac cctgccgatg cggatgaaaa ggagagtggc aaatggctgg cagcggccgc 
      601 aactagccgg ttacgcatgg gcagcatgat gagcaacgtg attgcggtct gtgaccgcga 
      661 agccgatatt catgcttatc tgcaggacag gctggcgcat aacgagcgct tcgtggtgcg 
      721 ctccaagcac ccacgcaagg acgtagagtc tgggttgtat ctgatcgacc atctgaagaa 
      781 ccaaccggag ttgggtggct atcagatcag cattccgcaa aagggcgtgg tggataaacg 
      841 cggtaaacgt aaaaatcgac cagcccgcaa ggcgagcttg agcctgcgca gtgggcgcat 
      901 cacgctaaaa caggggaata tcacgctcaa cgcggtgctg gccgaggaga ttaacccgcc 
      961 caagggtgag accccgttga aatggttgtt gctgaccggc gaaccggtcg agtcgctagc 
     1021 ccaagccttg cgcgtcatcg acatttatac ccatcgctgg cggatcgagg agttccataa 
     1081 ggcatggaaa accggagcag gagccgagag gcaacgcatg gaggagccgg ataatctgga 
     1141 gcggatggtc tcgatcctct cgtttgttgc ggtcaggctg ttacagctca gagaaagctt 
     1201 cacgctgccg caagcactca gggcgcaagg gctgctaaag gaagcggaac acgtagaaag 
     1261 ccagtccgca gaaacggtgc tgaccccgga tgaatgtcag ctactgggct atctggacaa 
     1321 gggaaaacgc aagcgcaaag agaaagcagg tagcttgcag tgggcttaca tggcgatagc 
     1381 tagactgggc ggttttatgg acagcaagcg aaccggaatt gccagctggg gcgccctctg 
     1441 gtaaggttgg gaagccctgc aaagtaaact ggatggcttt cttgccgcca aggatctgat 
     1501 ggcgcagggg atcaagatct gatcaagaga caggatgagg atcgtttcgc atgattgaac 
     1561 aagatggatt gcacgcaggt tctccggccg cttgggtgga gaggctattc ggctatgact 
     1621 gggcacaaca gacaatcggc tgctctgatg ccgccgtgtt ccggctgtca gcgcaggggc 
     1681 gcccggttct ttttgtcaag accgacctgt ccggtgccct gaatgaactg caggacgagg 
     1741 cagcgcggct atcgtggctg gccacgacgg gcgttccttg cgcagctgtg ctcgacgttg 
     1801 tcactgaagc gggaagggac tggctgctat tgggcgaagt gccggggcag gatctcctgt 
     1861 catctcacct tgctcctgcc gagaaagtat ccatcatggc tgatgcaatg cggcggctgc 
     1921 atacgcttga tccggctacc tgcccattcg accaccaagc gaaacatcgc atcgagcgag 
     1981 cacgtactcg gatggaagcc ggtcttgtcg atcaggatga tctggacgaa gagcatcagg 
     2041 ggctcgcgcc agccgaactg ttcgccaggc tcaaggcgcg catgcccgac ggcgaggatc 
     2101 tcgtcgtgac ccatggcgat gcctgcttgc cgaatatcat ggtggaaaat ggccgctttt 
     2161 ctggattcat cgactgtggc cggctgggtg tggcggaccg ctatcaggac atagcgttgg 
     2221 ctacccgtga tattgctgaa gagcttggcg gcgaatgggc tgaccgcttc ctcgtgcttt 
     2281 acggtatcgc cgctcccgat tcgcagcgca tcgccttcta tcgccttctt gacgagttct 
     2341 tctgagcggg actctggggt tcgaaatgac cgaccaagcg acgcccaacc tgccatcacg 
     2401 agatttcgat tccaccgccg ccttctatga aaggttgggc ttcggaatcg ttttccggga 
     2461 cgccggctgg atgatcctcc agcgcgggga tctcatgctg gagttcttcg cccaccccgg 
     2521 gctcgatccc ctcgcgagtt ggttcagctg ctgcctgagg ctggacgacc tcgcggagtt 
     2581 ctaccggcag tgcaaatccg tcggcatcca ggaaaccagc agcggctatc cgcgcatcca 
     2641 tgcccccgaa ctgcaggagt ggggaggcac gatggccgct ttggtcgacc cggacgggac 
     2701 gctcctgcgc ctgatacaga acgaattgct tgcaggcatc tcatgagtgt gtcttcccgt 
     2761 tttccgcctg aggtcactgc gtggatggag cgctggcgcc tgctgcgcga cggcgagctg 
     2821 ctcaccaccc actcgagctg gatacttccc gtccgccagg gggacatgcc ggcgatgctg 
     2881 aaggtcgcgc gcattcccga tgaagaggcc ggttaccgcc tgttgacctg gtgggacggg 
     2941 cagggcgccg cccgagtctt cgcctcggcg gcgggcgctc tgctcatgga gcgcgcgtcc 
     3001 ggggccgggg accttgcaca gatagcgtgg tccggccagg acgacgaggc ttgcaggatc 
     3061 ctctgcgaca ccgccgctcg tctgcacgcg ccgcggtccg gaccgccgcc cgatctccat 
     3121 ccgctacagg aatggttcca gccgcttttc cggttggccg ctgagcacgc ggcacttgcg 
     3181 cccgccgcca gcgtagcgcg ccaacttctg gcggcgccgc gcgaggtgtg cccgctccac 
     3241 ggcgacctgc accacgagaa cgtgctcgac ttcggcgacc gcggctggct ggccatcgac 
     3301 ccgcacggac tgctcggcga gcgcaccttc gactatgcca acatcttcac gaatcccgat 
     3361 ctcagcgacc ccggtcgccc gcttgcgatc ctgccgggca ggctggaggc tcgactcagc 
     3421 attgtggtcg cgacgaccgg gtttgagccc gaacggcttc ttcgctggat cattgcatgg 
     3481 acgggcttgt cggcagcctg gttcatcggc gacggcgacg gcgagggcga gggcgctgcg 
     3541 attgatctgg ccgtaaacgc catggcacgc cggttgcttg actagcgcgg tcaccgatct 
     3601 cacctggtcg tcgagctagg tcaggccgtg tcgggcgtga tccgctggaa gtcgttgcgg 
     3661 gccacacccg ccgcctcgaa gccctgcacc aggccggcat cgtggtgtgc gtggccgagg 
     3721 gactatggaa ggtgccggac gatctgcccg agcagggccg ccgctatgac gcccagcgtc 
     3781 ttggtggcgt gacggtggag ctgaaatcgc acctgcccat cgagcggcag gcccgcgtga 
     3841 tcggtgccac ctggcttgac cagcagttga tcgacggtgg ctcgggcttg ggcgacctgg 
     3901 gctttagcag tgaggccaag taggcgatac agcagcgcgc ggacttcctg gccgaacagg 
     3961 gactggccga gcggcgcggg cagcgcgtga tcctcaccgg aatctgctcg gcagcagcgg 
     4021 gctcgggaac tggcgcaggc cgcgaaggac attgccgccg ataccggcct ggagcatcgc 
     4081 cccgtggccg acggccagcg cgttgccggc gtctaccggc gccccgtcat gctcgccagc 
     4141 gggcgaaatg ggatgcttga tgacgccaag gggtccagcc tcgtgcggtg gaagcccatc 
     4201 gaacagcggc ttggggagca gctcgccgcg acggtgcgcg gtggcggcgt gtcttgggag 
     4261 attggacgac agcgtgggcc ggcccctgtc tcttgatcag atcttgatcc cctgcgccat 
     4321 cagatccttg gcggcaagaa agccatccag tttactttgc agggcttccc aaccttccca 
     4381 gagggcgccc cagctggcaa ttccggttcg cttgctgtcc ataaaaccgc ccagtctagc 
     4441 tatcgccatg taagcccact gcaagctacc tgctttctct ttgcgcttgc gttttccctt 
     4501 gtccagatag cccagtagct gacattcatc cggggtcagc accgtttctg cggactggct 
     4561 ttctacgtgt tccgcttcct ttagcagccc ttgcgccctg agtgcttgcg gcagcgtgaa 
     4621 gctttctctg agctgtaaca gcctgaccgc aacaaacgag aggatcgaga ccatccgctc 
     4681 cagattatcc ggctcctcca tgcgttgcct ctcggctcct gctccggttt tccatgcctt 
     4741 atggaactcc tcgatccgcc agcgatgggt ataaatgtcg atgacgcgca aggcttgggc 
     4801 tagcgactcg accggttcgc cggtcagcaa caaccatttc aacggggtct cacccttggg 
     4861 cgggttaatc tcctcggcca gcaccgcgtt gagcgtgata ttcccctgtt ttagcgtgat 
     4921 gcgcccactg cgcaggctca agctcgcctt gcgggctggt cgatttttac gtttaccgcg 
     4981 tttatccacc acgccctttt gcggaatgct gatctgatag ccacccaact ccggttggtt 
     5041 cttcagatgg tcgatcagat acaacccaga ctctacgtcc ttgcgtgggt gcttggagcg 
     5101 caccacgaag cgctcgttat gcgccagcct gtcctgcaga taagcatgaa tatcggcttc 
     5161 gcggtcacag accgcaatca cgttgctcat catgctgccc atgcgtaacc ggctagttgc 
     5221 ggccgctgcc agccatttgc cactctcctt ttcatccgca tcggcagggt catccgggcg 
     5281 catccaccac tcctgatgca gtaatcctac ggtgcggaat gtggtggcct cgagcaagag 
     5341 aacggagtga acccaccatc cgcgggattt atcctgaata gagcccagct tgccaagctc 
     5401 ttcggcgacc tggtggcgat aactcaaaga ggtggtgtcc tcaatggcca gcagttcggg 
     5461 aaactcctga gccaacttga ctgtttgcat ggcgccagcc tttctgatcg cctcggcaga 
     5521 aacgttggga ttgcggtaaa atcggtaagc gccttcctgc atggcttcac taccctctga 
     5581 tgagatggtt attgatttac cagaatattt tgccaattgg gcggcgacgt taaccaagcg 
     5641 ggcagtacgg cgaggatcac ccagcgccgc cgaagagaac acagatttag cccagtcggc 
     5701 cgcacgatga agagcagaag ttatcatgaa cgttaccatg ttaggaggtc acatggaagt 
     5761 cagatcctgg aaaacgggaa aggttccgtt caggacgcta cttgtgtata agagtcag 
//



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