LOCUS       sequence-487602-       18045 bp    DNA     circular SYN 01-OCT-2025
DEFINITION  pgSIT 2.0 gRNA expressing plasmid with working HR5Ie1-EGFP SEPARATOR
            cassette.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 18045)
  TITLE     High-throughput production of genetically sterile males for 
            population suppression of Anopheles gambiae in Africa
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 18045)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Oct 1, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: 1197TCC_gZBF-HG
FEATURES             Location/Qualifiers
     source          1..18045
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(30..458)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(140..161)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     352..371
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     585..607
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     599..616
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     600..616
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     repeat_region   746..808
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     misc_RNA        1818..1893
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     protein_bind    1923..1956
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     enhancer        1974..2456
                     /gene="Autographa californica hr5"
                     /label=hr5 enhancer
                     /note="baculovirus early transcription enhancer"
     promoter        2460..3051
                     /label=IE1 promoter
                     /note="promoter of the ie1 gene from the baculovirus 
                     Autographa californica"
     CDS             join(5667..5669,5670..6383)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     CDS             join(5667..5669,5670..6380)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(5709..5730)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(5970..5989)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     6317..6338
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    6504..6625
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     misc_RNA        6980..7055
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     misc_RNA        7447..7522
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     CDS             10252..10929
                     /codon_start=1
                     /product="improved tetrameric variant of DsRed fluorescent 
                     protein"
                     /label=DsRed2
                     /note="mammalian codon-optimized"
                     /translation="MASSENVITEFMRFKVRMEGTVNGHEFEIEGEGEGRPYEGHNTVK
                     LKVTKGGPLPFAWDILSPQFQYGSKVYVKHPADIPDYKKLSFPEGFKWERVMNFEDGGV
                     ATVTQDSSLQDGCFIYKVKFIGVNFPSDGPVMQKKTMGWEASTERLYPRDGVLKGETHK
                     ALKLKDGGHYLVEFKSIYMAKKPVQLPGYYYVDAKLDITSHNEDYTIVEQYERTEGRHH
                     LFL"
     primer_bind     complement(10432..10452)
                     /label=DsRed1-N
                     /note="DsRed1, reverse primer"
     primer_bind     10844..10867
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     polyA_signal    11050..11131
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     11158..11174
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     11159..11175
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     repeat_region   11458..11492
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(11983..11999)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(11983..11999)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(11996..12018)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    12007..12023
                     /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)."
     promoter        complement(join(12031..12037,12038..12055,12056..12061))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    12076..12097
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(12214..12231)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(16367..16955)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(16447..16466)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(17173..17192)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             17184..17993
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(17251..17270)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
ORIGIN
        1 ctaaattgta agcgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc
       61 attttttaac caataggccg aaatcggcaa aatcccttat aaatcaaaag aatagaccga
      121 gatagggttg agtgttgttc cagtttggaa caagagtcca ctattaaaga acgtggactc
      181 caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg aaccatcacc
      241 ctaatcaagt tttttggggt cgaggtgccg taaagcacta aatcggaacc ctaaagggag
      301 cccccgattt agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa
      361 agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac
      421 cacacccgcc gcgcttaatg cgccgctaca gggcgcgtcc cattcgccat tcaggctgcg
      481 caactgttgg gaagggcgat cggtgcgggc ctcttcgcta ttacgccagc tggcgaaagg
      541 gggatgtgct gcaaggcgat taagttgggt aacgccaggg ttttcccagt cacgacgttg
      601 taaaacgacg gccagtgagc gcgcctcgtt cattcacgtt tttgaacccg tggaggacgg
      661 gcagactcgc ggtgcaaatg tgttttacag cgtgatggag cagatgaaga tgctcgacac
      721 gctgcagaac acgcagctag attaacccta gaaagataat catattgtga cgtacgttaa
      781 agataatcat gtgtaaaatt gacgcatgtg ttttatcggt ctgtatatcg aggtttattt
      841 attaatttga atagatatta agttttatta tatttacact tacatactaa taataaattc
      901 aacaaacaat ttatttatgt ttatttattt attaaaaaaa acaaaaactc aaaatttctt
      961 ctataaagta acaaaacttt tatcggtcgt gacgtagatt cgtcgccaca tatacactga
     1021 agcggtttgg ctcttggtta ttctagaaaa ctctgttttt gaattaatca taggaaaact
     1081 gtttattctt ttttgatatt tcaacgtatt acaatgaata gtaacttttg attaaaaaag
     1141 agatgaagca atacaagaaa tgttgaaaag tttatgaaac atcacattag cgtgagttac
     1201 ggcaggatca aaaccttata acagtcacac tcaggcaaaa aatccttctt gaatatcctt
     1261 tatgcatcgc tagagcaagg attgaaagcg caaagtatat atacaacctt ttttcccctc
     1321 gtccttgatc gagcgcgtcg cctggtagtt ccagagctat ggaaacatag caagttggaa
     1381 taaggcttag tccgtactca acttgaaaaa gtggcaccga gtcggtgcat ttttttcggt
     1441 gactatcccc cacggtaagt gtctatcacc cctagccaca tatacactga agcggtttgg
     1501 ctcttggtta ttctagaaaa ctctgttttt gaattaatca taggaaaact gtttattctt
     1561 ttttgatatt tcaacgtatt acaatgaata gtaacttttg attaaaaaag agatgaagca
     1621 atacaagaaa tgttgaaaag tttatgaaac atcacattag cgtgagttac ggcaggatca
     1681 aaaccttata acagtcacac tcaggcaaaa aatccttctt gaatatcctt tatgcatcgc
     1741 tagagcaagg attgaaagcg caaagtatat atacaacctt ttttcccctc gtccttgctg
     1801 ggccaaaggg cactacagtt ttagagctag aaatagcaag ttaaaataag gctagtccgt
     1861 tatcaacttg aaaaagtggc accgagtcgg tgcttttttg gccgtttccc ggatataaac
     1921 atataacttc gtataatgta tgctatacga agttatggat ccactagatc cggcgcgtaa
     1981 aacacaatca agtatgagtc ataagctgat gtcatgtttt gcacacggct cataaccgaa
     2041 ctggctttac gagtagaatt ctacttgtaa cgcacgatca gtggatgatg tcatttgttt
     2101 ttcaaatcga gatgatgtca tgttttgcac acggctcata aactcgcttt acgagtagaa
     2161 ttctacgtgt aacgcacgat cgattgatga gtcatttgtt ttgcaatatg atatcataca
     2221 atatgactca tttgtttttc aaaaccgaac ttgatttacg ggtagaattc tacttgtaaa
     2281 gcacaatcaa aaagatgatg tcatttgttt ttcaaaactg aactcgcttt acgagtagaa
     2341 ttctacgtgt aaaacacaat caagaaatga tgtcatttgt tataaaaata aaagctgatg
     2401 tcatgttttg cacatggctc ataactaaac tcgctttacg ggtagaattc tacgcgcgtc
     2461 gatgtctttg tgatgcgcgc gacatttttg taggttattg ataaaatgaa cggatacgtt
     2521 gcccgacatt atcattaaat ccttggcgta gaatttgtcg ggtccattgt ccgtgtgcgc
     2581 tagcatgccc gtaacggacc tcgtactttt ggcttcaaag gttttgcgca cagacaaaat
     2641 gtgccacact tgcagctctg catgtgtgcg cgttaccaca aatcccaacg gcgcagtgta
     2701 cttgttgtat gcaaataaat ctcgataaag gcgcggcgcg cgaatgcagc tgatcacgta
     2761 cgctcctcgt gttccgttca aggacggtgt tatcgacctc agattaatgt ttatcggccg
     2821 actgttttcg tatccgctca ccaaacgcgt ttttgcatta acattgtatg tcggcggatg
     2881 ttctatatct aatttgaata aataaacgat aaccgcgttg gttttagagg gcataataaa
     2941 agaaatattg ttatcgtgtt cgccattagg gcagtataaa ttgacgttca tgttggatat
     3001 tgtttcagtt gcaagttgac actggcggcg acaagatcgt gaacaaccaa gtgacttaat
     3061 taacaaaatg cccctgatgt acgtcatact aaagagcgcc gatggcgatg tacaaaaagc
     3121 acaccagcgg atcgacgaag gtaagctggc gatgatggtg tcgttcgaca tcactttcat
     3181 caccgtgtca gacatctact gtgcctagca ccgggtccag tggtcacagg gtgtagcaaa
     3241 aacgtgttct tttttgcgag agactctacc tcatgatgca gctgttaagg aaaggtttca
     3301 gttgaaggca atttttccta ggataagatg atcttaagtt acctgcgtat tagtgtttaa
     3361 cattgtcgtc tcaactccca agaatgtttt aatcgtctag ggttagttta tgtatactgt
     3421 tctcattgaa atgtcgttca atccaacatg ttaagttagc tagctcagac acgagaagtt
     3481 aggagtatct gcatcttgaa ggtagcggca tatggtgtta tgccacgttc actgacttca
     3541 aaattcgata tataaaaaaa acaaaatcaa aaacaaaatt gtgaattccg tcagccagca
     3601 gcagtgacct tcaaagcctt acctttccat tcatttatgt ttaacacagg tcaagcggtg
     3661 gtcaacgaat actcacgatt gcataatctg aacatgtttg atggcgtgga gttgcgcaat
     3721 accacccgtc agagtggatg ataaactttc cgcaccactg taactgtccg tatctttgta
     3781 tgtgggtgtg tgtatgtgtg tttggtgaaa cgaattcaat agttctgtgc tattttaaat
     3841 caagccgcgt gcgcaactga tgccgataag ttcaaactag tgtttaagga gtggagcgag
     3901 agagccgcac cacggtacag aagggcagca gaatgggtcg gcagcctagc tgcactggtg
     3961 cggtgcgtcc ggcatctcgg ggagagggcg gggaaattct agtgttaaat cggagcagca
     4021 aaaacaaaac agtggtcgtc ccgttcaaga aacggcctat acacacacag aaaacactgc
     4081 agcatgtttg tacatagtag atcctagagc aggtggtcgc tgctcctcga acgctctgga
     4141 cgcacgactt tgcgcgtact tgcgtagcgt tccgccgatc gtgggtattc gtactgccac
     4201 aagcccgctt tctcccatgc aatctctgca accaaaccaa caaacaacaa caaaaaacca
     4261 atcgacacaa tgaatcacac cccttttgta tcatctgtat atttcttgtt ctttgcgttc
     4321 ttttctatgt ggcccacgcc ccggcgggta cgtaattgcg tcgaaaaccc cgaaaacccc
     4381 ggcacataca gtgtacatac ggtttgagga caactttgac ctgcagccct tctggggttg
     4441 ccacgtgtag ctatacttgt gagatcgggc gccgacggtg taaagcgcga atggccgcca
     4501 cacagtgtgt ccactccaac actacccctc tggaactacc ccgtccaggg atgcaccggc
     4561 tcggctcatg cccctgcaaa acagtccggg ctccactgta gtagctccgg cgttgctctg
     4621 agagaaggat gcccttcgaa gtgtcgaaag cgtgcattgg gcgttcaagt gtgtgtgtgt
     4681 gtgttaggtt tagcgagaaa cagcagcagt tgcgtgtgct gaaaagcgaa ggagtaatag
     4741 agtgcataat gaaaatgaaa atgaaaatga agcaaaagta gaaggcggag gagagcaacc
     4801 tgtgttccac tagtagcgaa tagtttagtc tagtttcgtc accaatcaac cttccaacca
     4861 tcgttcaacc aatacctgag tcaacatcgt catcgttatc gtgccacaac tttattaaaa
     4921 atgaaccttg tccgcgccac cgtagggtga tctgagacga cctttcttac gggcgcgacc
     4981 cacatgccat cgtcaccttc tccaatcaaa accaacagcc tgtaccgatg gtgtgcaatt
     5041 gtgcgtgcgt gtgtgttatt agcaaaaaaa gagaaagaga cggctagaga gagatagatc
     5101 gagatcgaga gtacaaaaga gcagtagaaa tgttcgttgt ttgttttccg taacacagtt
     5161 gttcagccaa aatgggaatt ttcaacaatc ccgggggcgg ggaaatgcgg gaatactgcg
     5221 tacacacata catcaatcaa aaagaaaaat ccttgcgcta catcactacc gtttgcgcgg
     5281 tgctgatcta gagcagacca ctttccactc cactctacaa tcaatcaatc tgtgcagaag
     5341 gtatggtaag acggcctttg agcgagtcac ggtcgccacc ataacgccgt ccgacgaggg
     5401 ctgaatgcga actttgctaa tcgattttcc gctttctttt tatcccaccc ccctttctct
     5461 ctctctctct cttttgcacc gccccttgta acccccaaaa aggtaaacga cacattaaga
     5521 cctacgaagc gctggtgaag tcatcgctcg atccgaacag cgaccggctg acggaggacg
     5581 acgacgagga cgagaacatc tcggtgaccc gcaccaactc caccattcgg tcgaggtcca
     5641 gctcgctgtc gccggcaccg gcaccggtga gcaagggcga ggagctgttc accggggtgg
     5701 tgcccatcct ggtcgagctg gacggcgacg taaacggcca caagttcagc gtgtccggcg
     5761 agggcgaggg cgatgccacc tacggcaagc tgaccctgaa gttcatctgc accaccggca
     5821 agctgcccgt gccctggccc accctcgtga ccaccctgac ctacggcgtg cagtgcttca
     5881 gccgctaccc cgaccacatg aagcagcacg acttcttcaa gtccgccatg cccgaaggct
     5941 acgtccagga gcgcaccatc ttcttcaagg acgacggcaa ctacaagacc cgcgccgagg
     6001 tgaagttcga gggcgacacc ctggtgaacc gcatcgagct gaagggcatc gacttcaagg
     6061 aggacggcaa catcctgggg cacaagctgg agtacaacta caacagccac aacgtctata
     6121 tcatggccga caagcagaag aacggcatca aggtgaactt caagatccgc cacaacatcg
     6181 aggacggcag cgtgcagctc gccgaccact accagcagaa cacccccatc ggcgacggcc
     6241 ccgtgctgct gcccgacaac cactacctga gcacccagtc cgccctgagc aaagacccca
     6301 acgagaagcg cgatcacatg gtcctgctgg agttcgtgac cgccgccggg atcactctcg
     6361 gcatggacga gctgtacaag tagcggccgc gactctagat cataatcagc cataccacat
     6421 ttgtagaggt tttacttgct ttaaaaaacc tcccacacct ccccctgaac ctgaaacata
     6481 aaatgaatgc aattgttgtt gttaacttgt ttattgcagc ttataatggt tacaaataaa
     6541 gcaatagcat cacaaatttc acaaataaag catttttttc actgcattct agttgtggtt
     6601 tgtccaaact catcaatgta tcttaacctc tcaatcccca catatacact gaagcggttt
     6661 ggctcttggt tattctagaa aactctgttt ttgaattaat cataggaaaa ctgtttattc
     6721 ttttttgata tttcaacgta ttacaatgaa tagtaacttt tgattaaaaa agagatgaag
     6781 caatacaaga aatgttgaaa agtttatgaa acatcacatt agcgtgagtt acggcaggat
     6841 caaaacctta taacagtcac actcaggcaa aaaatccttc ttgaatatcc tttatgcatc
     6901 gctagagcaa ggattgaaag cgcaaagtat atatacaacc ttttttcccc tcgtccttgg
     6961 atccgatcac gcagtcgatg ttttagagct agaaatagca agttaaaata aggctagtcc
     7021 gttatcaact tgaaaaagtg gcaccgagtc ggtgcttttt tccaccactc ttcacgccta
     7081 aagccaaaga gacgcttatt atggccacat atacactgaa gcggtttggc tcttggttat
     7141 tctagaaaac tctgtttttg aattaatcat aggaaaactg tttattcttt tttgatattt
     7201 caacgtatta caatgaatag taacttttga ttaaaaaaga gatgaagcaa tacaagaaat
     7261 gttgaaaagt ttatgaaaca tcacattagc gtgagttacg gcaggatcaa aaccttataa
     7321 cagtcacact caggcaaaaa atccttcttg aatatccttt atgcatcgct agagcaagga
     7381 ttgaaagcgc aaagtatata tacaaccttt tttcccctcg tccttgcgac ggctcgttca
     7441 tcgctggttt tagagctaga aatagcaagt taaaataagg ctagtccgtt atcaacttga
     7501 aaaagtggca ccgagtcggt gctttttttg ctgacggcgg aagcgtcagc ttctagttct
     7561 agtgttccca caatgggcgc atgtgcttgt gtgtgaggcg tctctctctt cgtctctgtt
     7621 gcgcaaacgc atagactgca ctgaaaaaat cgattaccta ttttttatga atgaatattt
     7681 gcactattac tattcaaaac tattaagata gcaatcacat tcaatagcca aatactatac
     7741 cacctgagcg atgcaacgaa atgatcaatt tgagcaaaaa tgctgcatat ttaggacggc
     7801 atcattatag aaatgcttct tgctgtgtac ttttctctcg tctggcagct gtttcgccgt
     7861 tattgttaaa accggcttaa gttaggtgtg ttttctacga ctagtgaatg ccctactaga
     7921 agatgtgtgt tgcacaaaat gtccctggaa taaccaattt gaagtgcaga tagcagtaaa
     7981 cgtaagctaa tatgaatatt atttaactgt aatgttttaa tatcgctgga cattactaat
     8041 aaacccacta taaacacatg tacatatgta tgttttggca tacaatgagt agttggggaa
     8101 aaaatgtgta aaagcaccgt gaccatcaca gcataaagat aaccagctga agtatcgaat
     8161 atgagtaacc cccaaattga atcacatgcc gcaactgata ggacccatgg aagtacactc
     8221 ttcatggcga tatacaagac acacacaagc acgaacaccc agttgcggag gaaattctcc
     8281 gtaaatgaaa acccaatcgg cgaacaattc atacccatat atggtaaaag ttttgaacgc
     8341 gacttgagag cggagagcat tgcggctgat aaggttttag cgctaagcgg gctttataaa
     8401 acgggctgcg ggaccagttt tcatatcact accgtttgag ttcttgtgct gtgtggatac
     8461 tcctcccgac acaaagccgc tccatcagcc agcagtcgtc taatccagag acaccaaacc
     8521 gaaagactta atttatattt atttaattaa ttttaataaa acacaccaaa tgtaagtagc
     8581 tttccccttc ccaacaacaa aacaccatcg aaccactccc accaagaaaa agcaataatc
     8641 gagaaaagcc gcggaaaatg tgtgattttt tttgtaaaca aaacattttt ttatgtgcca
     8701 gtgctgaaag tgatcaaaaa atactagcca cgagctaaag agttattgta ttgaccaaaa
     8761 ctccaaaaat acccaagttt ggccctaaat tgtcaatcaa aataccaata ggtcgaaaga
     8821 catcaaaatt aacaaaacca gggtttcaaa taccataact caagaatcag gattacaact
     8881 gcagatttca ggatatatac atacaaatta tacgaaatta taaaaaccaa agcaattcaa
     8941 tagccccaac tcaaatgtta ggatctaata tagtgtttaa agccaagctc gctgatgtgg
     9001 gcgtgtcacg atttcaccca aagatatgcc aaattacgaa ttgcaaatca attcgccaac
     9061 actttctttt tttcccacgc cctaaaacac cagatcatcc ataaatgtac atacatacag
     9121 tatatgcata ttataatcta taaaactaga tcaggttctt gaaaatagtg acgtaggcag
     9181 ccgttttggc ttaagcagaa atttttgccg gtttttcaaa gttgtagttg caaaaatgga
     9241 gaaaaccttc gagcattcgt tcatgtacac acactcacgc gcaaaataac gagagagagt
     9301 gtatgtgtgt gtgagagagc gaaagccaga cgacggtttg cttttcgcct cgaaacatga
     9361 ccatatatgg tcacaaaact tggccgccgc aattcaacac accagcgctc tccttcgcac
     9421 ccatagcgac catggcgcgg agcgagcgag atggcgagag cgagcgacgc ctatggcgac
     9481 gtcgacgcag gcagcgattg aaaaacgcag ttaactggca ttcaacattc accagccact
     9541 ttcagtcggt ttattccagt cattcctttc aaaccgtgcg gtcgcttagc tcagcctcgc
     9601 cacttgcgtt tacagtagtt ttcacgcctt gaatttgtta aatcgaacaa aaaggtaaag
     9661 tttaactagc tttgaaaagt ttcgtggctc ttaattgtta aattttctag agtgcgttta
     9721 gtgttttttt ttttattttg taatgttaat ttcgggttcc aattcgagtt ttaggcagcc
     9781 gccattttaa gggcgcatac acacaggcaa ctgtgctctc tttgcggctt tcttttgcac
     9841 cggcattcgt taagtgtcgt ctagaagctt ctcccctccc ttttcggcat attcgtattg
     9901 tggttttaat ttttcggggc ggggccttct attttgtaac tgttctttta atttcttatt
     9961 acaattcgat cgcaagtgaa aatcagtttt caatcggaaa agtatttttt tatgaaattt
    10021 ttttttgtcc aagattaaaa ttttgtacta aaaaaacgta cattgcattg agtgattttt
    10081 aattgtacac gaaaaacaag ttagtttgtt atgacaattg tattttggta aatcagcgca
    10141 gtccaaggaa accacgcaaa ttctcagttt ttttttttgc catttctaca ttaccaaata
    10201 aggtaaccaa aaactaatgg gaaatccgca ttctttccat tgcagcttac aatggcctcc
    10261 tccgagaacg tcatcaccga gttcatgcgc ttcaaggtgc gcatggaggg caccgtgaac
    10321 ggccacgagt tcgagatcga gggcgagggc gagggccgcc cctacgaggg ccacaacacc
    10381 gtgaagctga aggtgaccaa gggcggcccc ctgcccttcg cctgggacat cctgtccccc
    10441 cagttccagt acggctccaa ggtgtacgtg aagcaccccg ccgacatccc cgactacaag
    10501 aagctgtcct tccccgaggg cttcaagtgg gagcgcgtga tgaacttcga ggacggcggc
    10561 gtggcgaccg tgacccagga ctcctccctg caggacggct gcttcatcta caaggtgaag
    10621 ttcatcggcg tgaacttccc ctccgacggc cccgtgatgc agaagaagac catgggctgg
    10681 gaggcctcca ccgagcgcct gtacccccgc gacggcgtgc tgaagggcga gacccacaag
    10741 gccctgaagc tgaaggacgg cggccactac ctggtggagt tcaagtccat ctacatggcc
    10801 aagaagcccg tgcagctgcc cggctactac tacgtggacg ccaagctgga catcacctcc
    10861 cacaacgagg actacaccat cgtggagcag tacgagcgca ccgagggccg ccaccacctg
    10921 ttcctgtagc ggccgcgact ctagatcata atcagccata ccacatttgt agaggtttta
    10981 cttgctttaa aaaacctccc acacctcccc ctgaacctga aacataaaat gaatgcaatt
    11041 gttgttgtta acttgtttat tgcagcttat aatggttaca aataaagcaa tagcatcaca
    11101 aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc caaactatcg
    11161 aggtcgacgg tatcgataag cttgatatct ataacaagaa aatatatata taataagtta
    11221 tcacgtaagt agaacatgaa ataacaatat aattatcgta tgagttaaat cttaaaagtc
    11281 acgtaaaaga taatcatgcg tcattttgac tcacgcggtc gttatagttc aaaatcagtg
    11341 acacttaccg cattgacaag cacgcctcac gggagctcca agcggcgact gagatgtcct
    11401 aaatgcacag cgacggattc gcgctattta gaaagagaga gcaatatttc aagaatgcat
    11461 gcgtcaattt tacgcagact atctttctag ggttaatcta gctgcatcag gatcatatcg
    11521 tcgggtcttt tttccggctc agtcatcgcc caagctggcg ctatctgggc atcggggagg
    11581 aagaagcccg tgccttttcc cgcgaggttg aagcggcatg gaaagagttt gccgaggatg
    11641 actgctgctg cattgacgtt gagcgaaaac gcacgtttac catgatgatt cgggaaggtg
    11701 tggccatgca cgcctttaac ggtgaactgt tcgttcaggc cacctgggat accagttcgt
    11761 cgcggctttt ccggacacag ttccggatgg tcagcccgaa gcgcatcagc aacccgaaca
    11821 ataccggcga cagccggaac tgccgtgccg gtgtgcagat taatgacagc ggtgcggcgc
    11881 tgggatatta cgtcagcgag gacgggtatc ctggctggat gccgcagaaa tggacatgga
    11941 taccccgtga gttacccggc gggcgcgctt ggcgtaatca tggtcatagc tgtttcctgt
    12001 gtgaaattgt tatccgctca caattccaca caacatacga gccggaagca taaagtgtaa
    12061 agcctggggt gcctaatgag tgagctaact cacattaatt gcgttgcgct cactgcccgc
    12121 tttccagtcg ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag
    12181 aggcggtttg cgtattgggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt
    12241 cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga
    12301 atcaggggat aacgcaggaa agaacatgcg atgtagaacg cgagcaaatt cttttccttc
    12361 catgacagca gcagccacag tgggaagcct aacgtcagac gtgtttgaca tgccgagctg
    12421 ggcgggaaaa ttaccgcgtg cgcattgttt tcaagcaaat cacaactcgc tgcaaacaaa
    12481 accgttgaga aattgattgt tttataattt gtattttatt ttatttgtta taataaacta
    12541 aaaagacata ctttttgcat tttttataca taaaaacata catacagcat tataaaacac
    12601 atataagccc tccctgtaga gtcccgtgtc gaaatcttcc atccaagttg cacagtacga
    12661 cggacgagta ggccgtgtgg gtgcaaaatc cagcattttg ctgtcttttg ctcggagtac
    12721 tctcggcgag tcggaggttt ctgctgacgt gcttagcgct aaattagcca attgcttttg
    12781 caagtgaaat aaccagccga atagtacttc aaaactcagg taagtgaact agttttatag
    12841 aacaaatgtt tgtttgttag aagttagtga agtgtttgtg aaaaaaatct ctcatttcgg
    12901 caaaactaac gtaactgatt tcaaattgaa ttattgtttt gtgatgttat attatttcat
    12961 ccagttgatt agtattttct tagttatgtt caaaatacag ttaaattaaa tttcatttca
    13021 tttactcata aaataatctc ttggcttatt taatttttct cgaatttgct tgtattgttc
    13081 agtagcacgc gccattcgcc ctttgttaca ttttgtacct gctcccacta acacactggc
    13141 agtgcgaaac aaaagccttc gcacgcgttg ctggtattag agtgtgtgcg tgtgtgtgtt
    13201 gagcgctctg tcaaaatcgg ctgttgccgc cggtaccgaa attgcctgtt cgcacgctgt
    13261 tcgtaaacat tccgtggtgt gtatcgtgtg ttgtgcatat tgcgcgcctc ccccccttat
    13321 gatagcaaaa ggttgccgtg gtgtgtggcg cagttgagtt tttggattaa ttttctaagg
    13381 aaatggcacg agaagagcgg tggcagtgtg ttggtttgct ctgtcccttc ctttctgtgt
    13441 gaagtgttct tacagcacag cacgtatcca ccaccgcaca cagagcaggc aaggaagtgg
    13501 aagtgaacaa gtgtgctgcg catgcatgtg tgtggggggc attttagctg agatcgtcgt
    13561 tatttgagaa gcggtatagg ggccagtcgg tgtcgacgta cggaagcgat ttagttttaa
    13621 tccaagcgta tcccgtcgtg gagtggttgt gtggctctgt gtgctctcat atcagttcct
    13681 gagtgaggtt agtagaatca cagtccttgg cctttttcgt tacaagatat ccagaaggat
    13741 ggcgttattt ccacagctta ccatggtgct cttgtgtgct cgaatcaggg gtgaaaatcc
    13801 gttttgtgtt tcatgaaccg cagttggcac tggagcggat tcataagtct tcgatatgca
    13861 atagataaga gagacgttgg ggcatagttg ggaagctttc cgagatgtgg agtttccgag
    13921 aggagaaatg gtgctttcgt gcacgttccg ggacagcggg ccccgcgaag agcatctcgt
    13981 tgtcgttcat ccggcaataa ttgatgcgaa aagcgcgcgc gccactggct tagcgcagtg
    14041 tacacagtga tattcaccta cacacacaga ggcacacgcc ttcacacgcg cgcgtgcttc
    14101 aaaggctact tcggtagcgg tgtgtgaggt cgcttgcaat ggacaatgaa aatttcgctg
    14161 gaaaatacca tcgtcgcttt aggttgctat gagtgcgggt agagcggtgg tcgtcgatag
    14221 tggtggtgta gtgtgtgtgt gtgcaacggc aattattttt tataatattt cgaccatctt
    14281 tctttctctc tctccacgtg ctgctgctgc tgcattgcat gttccactat tcctctcggt
    14341 ttgtgcctgc ggacgccatt gctagtcgaa agagagtcgc cgttagtcgc gcttcgagca
    14401 acggacacgt tttttggttg aaaccaacag cttttttcat cttcgggaga cacacagatc
    14461 tcgaatcgta cattcccaaa ggagaattgt catcttccgg tgaataaaga aaggaaacaa
    14521 tatgggctct agcctggacg acgagcacat cctgagcgcc ctgctgcaga gcgacgacga
    14581 actggtgggc gaggacagcg acagcgaggt cagcgaccac gtgtccgagg acgacgtgca
    14641 gtccgacacc gaggaagcct tcatcgacga ggtgcacgaa gtgcagccta ccagcagcgg
    14701 ctccgagatc ctggacgagc agaacgtgat cgagcagcct ggcagctccc tggccagcaa
    14761 cagaatcctg accctgcccc agagaaccat cagaggcaag aacaagcact gctggtccac
    14821 ctccaagagc accaggcgga gcagagtgtc cgccctgaac atcgtgcgga gccagagggg
    14881 ccccaccaga atgtgcagaa acatctacga ccccctgctg tgcttcaagc tgttcttcac
    14941 cgacgagatc atcagcgaga tcgtgaagtg gaccaacgcc gagatcagcc tgaagaggcg
    15001 ggagagcatg accagcgcca ccttcagaga caccaacgag gacgagatct acgccttctt
    15061 cggcatcctg gtgatgaccg ccgtgagaaa ggacaaccac atgagcaccg acgacctgtt
    15121 cgacagatcc ctgagcatgg tgtacgtgtc cgtgatgagc agagacagat tcgacttcct
    15181 gatcagatgc ctgagaatgg acgacaagag catcagaccc accctgcggg agaacgacgt
    15241 gttcaccccc gtgcggaaga tctgggacct gttcatccac cagtgcatcc agaactacac
    15301 ccctggcgcc cacctgacca tcgatgagca gctgctgggc ttcagaggca gatgcccctt
    15361 cagagtgtac atccccaaca agcccagcaa gtacggcatc aagatcctga tgatgtgcga
    15421 cagcggcacc aagtacatga tcaacggcat gccctacctg ggcagaggca cccagacaaa
    15481 cggcgtgccc ctgggcgagt actacgtgaa agaactgagc aagcctgtgc atggcagctg
    15541 caggaacatc acctgcgaca actggttcac cagcatcccc ctggccaaga acctgctgca
    15601 ggaaccctac aagctgacca tcgtgggcac cgtgcggagc aacaagcggg agatcccaga
    15661 ggtgctgaag aacagcagat ccagacctgt gggaacaagc atgttctgct tcgacggccc
    15721 cctgaccctg gtgtcctaca agcccaagcc cgccaagatg gtgtacctgc tgtccagctg
    15781 cgacgaggac gccagcatca acgagagcac cggcaagccc cagatggtga tgtactacaa
    15841 ccagaccaag ggcggcgtgg acaccctgga ccagatgtgc agcgtgatga cctgcagcag
    15901 aaagaccaac agatggccca tggccctgct gtacggcatg atcaatatcg cctgcatcaa
    15961 cagcttcatc atctacagcc acaacgtgtc cagcaagggc gagaaggtgc agagccggaa
    16021 gaaattcatg cggaacctgt acatgagcct gacctccagc ttcatgagaa agagactgga
    16081 agcccccacc ctgaagagat acctgcggga caacatcagc aacatcctgc ccaaggaagt
    16141 gccaggaaca agcgacgaca gcaccgagga acccgtgatg aagaagagga cctactgcac
    16201 ctactgtccc agcaagatca gaagaaaggc caacgccagc tgcaagaaat gcaaaaaagt
    16261 gatctgccgg gagcacaaca tcgacatgtg ccagagctgt ttctgacatg tgagcaaaag
    16321 gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc
    16381 gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag
    16441 gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga
    16501 ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc
    16561 atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg
    16621 tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt
    16681 ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca
    16741 gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca
    16801 ctagaagaac agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag
    16861 ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca
    16921 agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg
    16981 ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa
    17041 aaaggatctt cacctagatc cctgtttcct ggcagctctg gcccgtgtct caaaatctct
    17101 gatgttacat tgcacaagat aaaaatatat catcatgaac aataaaactg tctgcttaca
    17161 taaacagtaa tacaaggggt gttatgagcc atattcaacg ggaaacgtcg aggccgcgat
    17221 taaattccaa catggatgct gatttatatg ggtataaatg ggctcgcgat aatgtcgggc
    17281 aatcaggtgc gacaatctat cgcttgtatg ggaagcccga tgcgccagag ttgtttctga
    17341 aacatggcaa aggtagcgtt gccaatgatg ttacagatga gatggtcaga ctaaactggc
    17401 tgacggaatt tatgcctctt ccgaccatca agcattttat ccgtactcct gatgatgcat
    17461 ggttactcac cactgcgatc cccggaaaaa cagcattcca ggtattagaa gaatatcctg
    17521 attcaggtga aaatattgtt gatgcgctgg cagtgttcct gcgccggttg cattcgattc
    17581 ctgtttgtaa ttgtcctttt aacagcgatc gcgtatttcg tctcgctcag gcgcaatcac
    17641 gaatgaataa cggtttggtt gatgcgagtg attttgatga cgagcgtaat ggctggcctg
    17701 ttgaacaagt ctggaaagaa atgcataaac ttttgccatt ctcaccggat tcagtcgtca
    17761 ctcatggtga tttctcactt gataacctta tttttgacga ggggaaatta ataggttgta
    17821 ttgatgttgg acgagtcgga atcgcagacc gataccagga tcttgccatc ctatggaact
    17881 gcctcggtga gttttctcct tcattacaga aacggctttt tcaaaaatat ggtattgata
    17941 atcctgatat gaataaattg cagtttcatt tgatgctcga tgagtttttc taatcagaat
    18001 tggttaattg gttgtaacac tggccatttc cccgaaaagt gccac
//
