LOCUS       sequence-461775-       10368 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bxb1-GT donor plasmid with downstream tandem syntax for all-in-one 
            doxycycline-inducible expression of mouse neurogenin 2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 10368)
  AUTHORS   Blanch-Asensio A, Ploessl DS, Wang NB, Mummery CL, Galloway KE, 
            Davis RP
  TITLE     STRAIGHT-IN Dual: a platform for dual, single-copy integrations of 
            DNA payloads and gene circuits into human induced pluripotent stem 
            cells.
  JOURNAL   bioRxiv [Preprint]. 2024 Oct 17:2024.10.17.616637. doi: 
            10.1101/2024.10.17.616637.
  PUBMED    39464154
REFERENCE   2  (bases 1 to 10368)
  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: "bioRxiv 
            [Preprint]. 2024 Oct 17:2024.10.17.616637. doi: 
            10.1101/2024.10.17.616637."
COMMENT     Sequence Label: Bxb1-GT_AIO-TetOn_mNgn2 _Down-Tandem
FEATURES             Location/Qualifiers
     source          1..10368
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    107..128
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(143..148,149..166,167..173)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    181..197
                     /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     186..208
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     205..221
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     205..221
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        253..463
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     promoter        275..471
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      322..457
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     promoter        558..1736
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          789..1727
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1684..1704
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     regulatory      1744..1753
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    1756..1793
                     /label=Bxb1 attB
                     /bound_moiety="Mycobacteriophage Bxb1 integrase"
                     /note="Minimal attB site for the Mycopbacteriophage Bxb1 
                     integrase (Voutev and Mann, 2017)"
     protein_bind    complement(1802..1835)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    1881..1899
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1917..1935
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1953..1971
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1989..2007
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2025..2043
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2061..2079
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2097..2115
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     2160..2184
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             3097..3123
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             3133..3159
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             3169..3195
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     misc_feature    3206..3794
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3259..3279)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3677..3688)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     enhancer        3851..4230
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     5400..5422
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     5484..5503
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     CDS             5535..6278
                     /codon_start=1
                     /product="modified rtTA protein that binds tightly to 
                     promoters containing the tet operator in the presence of 
                     doxycycline"
                     /label=Tet-On(R) 3G
                     /translation="MSRLDKSKVINSALELLNGVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALPIEMLDRHHTHSCPLEGESWQDFLRNNAKSYRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEEQEHQVAKEERETPTT
                     DSMPPLLKQAIELFDRQGAEPAFLFGLELIICGLEKQLKCESGGPTDALDDFDLDMLPA
                     DALDDFDLDMLPADALDDFDLDMLPG"
     CDS             6279..6332
                     /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             6333..7040
                     /codon_start=1
                     /product="enhanced monomeric blue fluorescent protein 
                     (Subach et al., 2011)"
                     /label=mTagBFP2
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELIKENMHMKLYMEGTVDNHHFKCTSEGEGKPYEGTQTMR
                     IKVVEGGPLPFAFDILATSFLYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGV
                     LTATQDTSLQDGCLIYNVKIRGVNFTSNGPVMQKKTLGWEAFTETLYPADGGLEGRNDM
                     ALKLVGGSHLIANAKTTYRSKKPAKNLKMPGVYYVDYRLERIKEANNETYVEQHEVAVA
                     RYCDLPSKLGHKLN"
     CDS             7065..7085
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             7089..7109
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             7113..7133
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(7198..7215)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    7204..7428
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     protein_bind    complement(7457..7489)
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     primer_bind     complement(7497..7514)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(7497..7513)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(7506..7528)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      7726..8181
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(7813..7832)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     8023..8044
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(8198..8217)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     8317..8339
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(8377..8395)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        8463..8567
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(8568..8636,8637..9428)
                     /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(8786..8805)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      9599..10187
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     10088..10107
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     10341..10358
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 agcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc
       61 acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc
      121 tcactcatta ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa
      181 ttgtgagcgg ataacaattt cacacaggaa acagctatga ccatgattac gccaagctta
      241 attccctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccata
      301 gtcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc ccattctccg
      361 ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc ggcctctgag
      421 ctattccaga agtagtgagg aggctttttt ggaggcctag gcttttgcaa aaagctttgc
      481 aaagatggat aaagttttaa acagagagga atctttgcag ctaatggacc ttctaggtct
      541 tgaaaggagt gggaattggc tccggtgccc gtcagtgggc agagcgcaca tcgcccacag
      601 tccccgagaa gttgggggga ggggtcggca attgaaccgg tgcctagaga aggtggcgcg
      661 gggtaaactg ggaaagtgat gtcgtgtact ggctccgcct ttttcccgag ggtgggggag
      721 aaccgtatat aagtgcagta gtcgccgtga acgttctttt tcgcaacggg tttgccgcca
      781 gaacacaggt aagtgccgtg tgtggttccc gcgggcctgg cctctttacg ggttatggcc
      841 cttgcgtgcc ttgaattact tccacctggc tgcagtacgt gattcttgat cccgagcttc
      901 gggttggaag tgggtgggag agttcgaggc cttgcgctta aggagcccct tcgcctcgtg
      961 cttgagttga ggcctggcct gggcgctggg gccgccgcgt gcgaatctgg tggcaccttc
     1021 gcgcctgtct cgctgctttc gataagtctc tagccattta aaatttttga tgacctgctg
     1081 cgacgctttt tttctggcaa gatagtcttg taaatgcggg ccaagatctg cacactggta
     1141 tttcggtttt tggggccgcg ggcggcgacg gggcccgtgc gtcccagcgc acatgttcgg
     1201 cgaggcgggg cctgcgagcg cggccaccga gaatcggacg ggggtagtct caagctggcc
     1261 ggcctgctct ggtgcctggc ctcgcgccgc cgtgtatcgc cccgccctgg gcggcaaggc
     1321 tggcccggtc ggcaccagtt gcgtgagcgg aaagatggcc gcttcccggc cctgctgcag
     1381 ggagctcaaa atggaggacg cggcgctcgg gagagcgggc gggtgagtca cccacacaaa
     1441 ggaaaagggc ctttccgtcc tcagccgtcg cttcatgtga ctccacggag taccgggcgc
     1501 cgtccaggca cctcgattag ttctcgagct tttggagtac gtcgtcttta ggttgggggg
     1561 aggggtttta tgcgatggag tttccccaca ctgagtgggt ggagactgaa gttaggccag
     1621 cttggcactt gatgtaattc tccttggaat ttgccctttt tgagtttgga tcttggttca
     1681 ttctcaagcc tcagacagtg gttcaaagtt tttttcttcc atttcaggtg tcgtgaggaa
     1741 ttcgccacca tggccggctt gtcgacgacg gcggtctccg tcgtcaggat catccggatc
     1801 cataacttcg tatagcatac attatacgaa gttatcatga tattcggcaa gcaggcatcg
     1861 actagttaat taaggtttac tccctatcag tgatagagaa cgtatgaaga gtttactccc
     1921 tatcagtgat agagaacgta tgcagacttt actccctatc agtgatagag aacgtataag
     1981 gagtttactc cctatcagtg atagagaacg tatgaccagt ttactcccta tcagtgatag
     2041 agaacgtatc tacagtttac tccctatcag tgatagagaa cgtatatcca gtttactccc
     2101 tatcagtgat agagaacgta taagctttag gcgtgtacgg tgggcgccta taaaagcaga
     2161 gctcgtttag tgaaccgtca gatcgcctgg agcaattcca caacactttt gtcttatacc
     2221 aactttccgt accacttcct accctcgtaa acttaagaat tcgccaccat gttcgtcaaa
     2281 tctgagactc tggagttgaa ggaggaagag gaggtactga tgctgctggg ctcggcttcc
     2341 ccggcctcgg cgaccctgac cccgatgtcc tccagcgcgg acgaggagga ggacgaggag
     2401 ctgcgccggc cgggctccgc gcgtgggcag cgtggagcgg aagccgggca gggggtgcag
     2461 ggcagtccgg cgtcgggtgc cgggggttgc cggccagggc ggctgctggg cctgatgcac
     2521 gagtgcaagc gtcgcccgtc gcgctcacgg gccgtctccc gaggtgccaa gacggcggag
     2581 acggtgcagc gcatcaagaa gacccgcagg ctcaaggcca acaaccgcga gcgcaaccgc
     2641 atgcacaacc taaacgccgc gctggacgcg ctgcgcgagg tgctgcccac cttccccgag
     2701 gatgccaagc tcacgaagat cgagacgctg cgcttcgccc acaattacat ctgggcgctc
     2761 accgagactc tgcgcctggc ggaccactgc gccggcgccg gtggcctcca gggggcgctc
     2821 ttcacggagg cggtgctcct gagcccggga gctgcgctcg gcgccagcgg ggacagccct
     2881 tctccacctt cctcctggag ctgcaccaac agcccggcgt catcctccaa ctccacgtcc
     2941 ccatacagct gcactttatc gcccgctagc cccgggtcag acgtggacta ctggcagccc
     3001 ccacctccgg agaagcatcg ttatgcgcct cacctgcccc tcgccaggga ctgtatcggg
     3061 tctggctccg gatcaagtgg tggcagcgga tcaggatatc catacgatgt accagactat
     3121 gcgggatcgg gttaccccta cgacgtcccc gactacgccg gctccggcta cccctacgac
     3181 gtccctgatt atgcttagcg gccgcaatca acctctggat tacaaaattt gtgaaagatt
     3241 gactggtatt cttaactatg ttgctccttt tacgctatgt ggatacgctg ctttaatgcc
     3301 tttgtatcat gctattgctt cccgtatggc tttcattttc tcctccttgt ataaatcctg
     3361 gttgctgtct ctttatgagg agttgtggcc cgttgtcagg caacgtggcg tggtgtgcac
     3421 tgtgtttgct gacgcaaccc ccactggttg gggcattgcc accacctgtc agctcctttc
     3481 cgggactttc gctttccccc tccctattgc cacggcggaa ctcatcgccg cctgccttgc
     3541 ccgctgctgg acaggggctc ggctgttggg cactgacaat tccgtggtgt tgtcggggaa
     3601 gctgacgtcc tttccatggc tgctcgcctg tgttgccacc tggattctgc gcgggacgtc
     3661 cttctgctac gtcccttcgg ccctcaatcc agcggacctt ccttcccgcg gcctgctgcc
     3721 ggctctgcgg cctcttccgc gtcttcgcct tcgccctcag acgagtcgga tctccctttg
     3781 ggccgcctcc ccgccctttg ggccgcctcc ccgcgctagt gctggggatg cggtgggctc
     3841 tatgggtctc gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta
     3901 gttcatagcc catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc
     3961 tgaccgccca acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg
     4021 ccaataggga ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg
     4081 gcagtacatc aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa
     4141 tggcccgcct ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac
     4201 atctacgtat tagtcatcgc tattaccatg gtcgaggtga gccccgcgtt ctgcttcact
     4261 ctccccatct cccccccctc ccccccccca attttgtatt tttttttttt ttaatttttt
     4321 ggggcggggg gggggggggg gggggggggg gcgcgcgcca ggcggggcgg ggcggggcga
     4381 ggggcggggc ggggcgaggc ggagaggtgc ggcggcagcc aatcagagcg gcgcgctccg
     4441 aaagtttcct tttatggcga ggcggcggcg gcggcggccc tataaaaagc gaagcgcgcg
     4501 gcgggcggga gtcgctgcgc gctgccttcg ccccgtgccc cgctccgccg ccgcctcgcg
     4561 ccgcccgccc cggctctgac tgaccgcgtt actcccacag gtgagcgggc gggacggccc
     4621 ttctcctccg ggctgtaatt agcgcttggt ttaatgacgg cttgtttctt ttctgtggct
     4681 gcgtgaaagc cttgaggggc tccgggaggg ccctttgtgc ggggggagcg gctcgggggg
     4741 tgcgtgcgtg tgtgtgtgcg tggggagcgc cgcgtgcggc tccgcgctgc ccggcggctg
     4801 tgagcgctgc gggcgcggcg cggggctttg tgcgctccgc agtgtgcgcg aggggagcgc
     4861 ggccgggggc ggtgccccgc ggtgcggggg gggctgcgag gggaacaaag gctgcgtgcg
     4921 gggtgtgtgc gtgggggggt gagcaggggg tgtgggcgcg tcggtcgggc tgcaaccccc
     4981 cctgcacccc cctccccgag ttgctgagcg cggcccggct tcgggtgcgg ggctccgtac
     5041 ggggcgtggc gcggggctcg ccgtgccggg cggggggtgg cggcaggtgg gggtgccggg
     5101 cggcccccgg agcgccggcg gctgtcgagg cgcggcgagc cgcagccatt gccttttatg
     5161 gtaatcgtgc gagagggcgc agggacttcc tttgtcccaa atctgtgcgg agccgaaatc
     5221 tgggaggcgc cgccgcaccc cctctagcgg gcgcggggcg aagcggtgcg gcgccggcag
     5281 gaaggaaatg ggcggggagg gccttcgtgc gtcgccgcgc cgccgtcccc ttctccctct
     5341 ccagcctcgg ggctgtccgc ggggggacgg ctgccttcgg gggggacggg gcagggcggg
     5401 gttcggcttc tggcgtgtga ccggcggctc tagagcctct gctaaccatg ttcatgcctt
     5461 cttctttttc ctacagctcc tgggcaacgt gctggttatt gtgctgtctc atcattttgg
     5521 caaagaatgt cgacatgagc cggctggaca agagcaaagt gatcaacagc gccctggaac
     5581 tgctgaacgg cgtgggcatc gagggcctga ccacccggaa gctggcccag aaactgggcg
     5641 tggaacagcc caccctgtac tggcacgtga agaacaagcg ggccctgctg gacgccctgc
     5701 ccatcgagat gctggaccgg caccacaccc acagctgccc tctggaaggc gagagctggc
     5761 aggacttcct gcggaacaac gccaagagct acagatgcgc cctgctgagc caccgggacg
     5821 gcgccaaagt gcacctgggc accagaccca ccgagaagca gtacgagaca ctggaaaacc
     5881 agctggcctt cctgtgccag cagggcttca gcctggaaaa cgccctgtac gccctgagcg
     5941 ccgtgggcca cttcaccctg ggctgcgtgc tggaagaaca ggaacaccag gtcgccaaag
     6001 aggaaagaga gacacccacc accgacagca tgccccccct gctgaagcag gccatcgagc
     6061 tgttcgacag acagggcgcc gagcccgcct tcctgttcgg cctggaactg atcatctgcg
     6121 gcctcgagaa gcagctgaag tgcgagagcg gcggacccac cgacgccctg gacgacttcg
     6181 acctggacat gctgcccgcc gatgccctgg atgattttga tctggatatg ctccctgctg
     6241 acgctctcga tgacttcgat ctcgatatgc tgccaggcga gggcagggga agtcttctaa
     6301 catgcgggga cgtggaggaa aatcccggcc ccgtgtctaa gggcgaagag ctgattaagg
     6361 agaacatgca catgaagctg tacatggagg gcaccgtgga caaccatcac ttcaagtgca
     6421 catccgaggg cgaaggcaag ccctacgagg gcacccagac catgagaatc aaggtggtcg
     6481 agggcggccc tctccccttc gccttcgaca tcctggctac tagcttcctc tacggcagca
     6541 agaccttcat caaccacacc cagggcatcc ccgacttctt caagcagtcc ttccctgagg
     6601 gcttcacatg ggagagagtc accacatacg aagacggggg cgtgctgacc gctacccagg
     6661 acaccagcct ccaggacggc tgcctcatct acaacgtcaa gatcagaggg gtgaacttca
     6721 catccaacgg ccctgtgatg cagaagaaaa cactcggctg ggaggccttc accgagacgc
     6781 tgtaccccgc tgacggcggc ctggaaggca gaaacgacat ggccctgaag ctcgtgggcg
     6841 ggagccatct gatcgcaaac gccaagacca catatagatc caagaaaccc gctaagaacc
     6901 tcaagatgcc tggcgtctac tatgtggact acagactgga aagaatcaag gaggccaaca
     6961 acgagaccta cgtcgagcag cacgaggtgg cagtggccag atactgcgac ctccctagca
     7021 aactggggca caagcttaat tccggactca gatctcgagc tgatccaaaa aagaagagaa
     7081 aggtagatcc aaaaaagaag agaaaggtag atccaaaaaa gaagagaaag gtaggatcca
     7141 ccggatctag ataactgatc ataatcagcc ataccacatt tgaaacccgc tgatcagcct
     7201 cgactgtgcc ttctagttgc cagccatctg ttgtttgccc ctcccccgtg ccttccttga
     7261 ccctggaagg tgccactccc actgtccttt cctaataaaa tgaggaaatt gcatcgcatt
     7321 gtctgagtag gtgtcattct attctggggg gtggggtggg gcaggacagc aagggggagg
     7381 attgggaaga caatagcagg catgctgggg atgcggtggg ctctatggct agccgctgca
     7441 agaactcttc ctcacgataa cttcgtatag gagactttat acgaagttaa gcgctcactg
     7501 gccgtcgttt tacaacgtcg tgactgggaa aaccctggcg ttacccaact taatcgcctt
     7561 gcagcacatc cccctttcgc cagctggcgt aatagcgaag aggcccgcac cgatcgccct
     7621 tcccaacagt tgcgcagcct gaatggcgaa tggcgcctga tgcggtattt tctccttacg
     7681 catctgtgcg gtatttcaca ccgcatacgt caaagcaacc atagtacgcg ccctgtagcg
     7741 gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg
     7801 ccttagcgcc cgctcctttc gctttcttcc cttcctttct cgccacgttc gccggctttc
     7861 cccgtcaagc tctaaatcgg gggctccctt tagggttccg atttagtgct ttacggcacc
     7921 tcgaccccaa aaaacttgat ttgggtgatg gttcacgtag tgggccatcg ccctgataga
     7981 cggtttttcg ccctttgacg ttggagtcca cgttctttaa tagtggactc ttgttccaaa
     8041 ctggaacaac actcaactct atctcgggct attcttttga tttataaggg attttgccga
     8101 tttcggtcta ttggttaaaa aatgagctga tttaacaaaa atttaacgcg aattttaaca
     8161 aaatattaac gtttacaatt ttatggtgca ctctcagtac aatctgctct gatgccgcat
     8221 agttaagcca gccccgacac ccgccaacac ccgctgacgc gccctgacgg gcttgtctgc
     8281 tcccggcatc cgcttacaga caagctgtga ccgtctccgg gagctgcatg tgtcagaggt
     8341 tttcaccgtc atcaccgaaa cgcgcgagac gaaagggcct cgtgatacgc ctatttttat
     8401 aggttaatgt catgataata atggtttctt agacgtcagg tggcactttt cggggaaatg
     8461 tgcgcggaac ccctatttgt ttatttttct aaatacattc aaatatgtat ccgctcatga
     8521 gacaataacc ctgataaatg cttcaataat attgaaaaag gaagagtatg agtattcaac
     8581 atttccgtgt cgcccttatt cccttttttg cggcattttg ccttcctgtt tttgctcacc
     8641 cagaaacgct ggtgaaagta aaagatgctg aagatcagtt gggtgcacga gtgggttaca
     8701 tcgaactgga tctcaacagc ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc
     8761 caatgatgag cacttttaaa gttctgctat gtggcgcggt attatcccgt attgacgccg
     8821 ggcaagagca actcggtcgc cgcatacact attctcagaa tgacttggtt gagtactcac
     8881 cagtcacaga aaagcatctt acggatggca tgacagtaag agaattatgc agtgctgcca
     8941 taaccatgag tgataacact gcggccaact tacttctgac aacgatcgga ggaccgaagg
     9001 agctaaccgc ttttttgcac aacatggggg atcatgtaac tcgccttgat cgttgggaac
     9061 cggagctgaa tgaagccata ccaaacgacg agcgtgacac cacgatgcct gtagcaatgg
     9121 caacaacgtt gcgcaaacta ttaactggcg aactacttac tctagcttcc cggcaacaat
     9181 taatagactg gatggaggcg gataaagttg caggaccact tctgcgctcg gcccttccgg
     9241 ctggctggtt tattgctgat aaatctggag ccggtgagcg tgggtctcgc ggtatcattg
     9301 cagcactggg gccagatggt aagccctccc gtatcgtagt tatctacacg acggggagtc
     9361 aggcaactat ggatgaacga aatagacaga tcgctgagat aggtgcctca ctgattaagc
     9421 attggtaact gtcagaccaa gtttactcat atatacttta gattgattta aaacttcatt
     9481 tttaatttaa aaggatctag gtgaagatcc tttttgataa tctcatgacc aaaatccctt
     9541 aacgtgagtt ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt
     9601 gagatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag
     9661 cggtggtttg tttgccggat caagagctac caactctttt tccgaaggta actggcttca
     9721 gcagagcgca gataccaaat actgttcttc tagtgtagcc gtagttaggc caccacttca
     9781 agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg
     9841 ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg
     9901 cgcagcggtc gggctgaacg gggggttcgt gcacacagcc cagcttggag cgaacgacct
     9961 acaccgaact gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga
    10021 gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc
    10081 ttccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg
    10141 agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg
    10201 cggccttttt acggttcctg gccttttgct ggccttttgc tcacatgttc tttcctgcgt
    10261 tatcccctga ttctgtggat aaccgtatta ccgcctttga gtgagctgat accgctcgcc
    10321 gcagccgaac gaccgagcgc agcgagtcag tgagcgagga agcggaag
//
