LOCUS       sequence-472753-       11929 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Lentiviral expression of ComMAND closed-loop circuit regulating 
            EGFP-FMRP (therapeutically relevant gene).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 11929)
  AUTHORS   Love KS, Johnstone CP, Peterman EL, Gaglione S, Birnbaum ME, 
            Galloway KE
  TITLE     Model-guided design of microRNA-based gene circuits supports precise
            dosage of transgenic cargoes into diverse primary cells.
  JOURNAL   Cell Syst. 2025 Apr 22:101269. doi: 10.1016/j.cels.2025.101269.
  PUBMED    40300600
REFERENCE   2  (bases 1 to 11929)
  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: "Cell Syst. 
            2025 Apr 22:101269. doi: 10.1016/j.cels.2025.101269."
COMMENT     Sequence Label: 
            pLentiX1-[TRE3G-EGFP(miRE.FF4)-FMRP-TS.FF4x1-bGH]-EFS-rtTA-P2A-mRuby
            2-WPRE
FEATURES             Location/Qualifiers
     source          1..11929
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    36..161
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    654..887
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1072..1116
                     /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    1383..1499
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     polyA_signal    1526..1750
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     CDS             complement(3643..3666)
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     primer_bind     complement(3751..3772)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     ncRNA           complement(4133..4245)
                     /label=3' miR-30a
                     /note="sequence downstream of the 71-nt precursor of the 
                     human miR-30a microRNA (Zeng et al., 2002)"
                     /ncRNA_class="miRNA"
     ncRNA           complement(4302..4316)
                     /label=miR-30a loop
                     /note="loop from the the 71-nt precursor of the human 
                     miR-30a microRNA (Zeng et al., 2002)"
                     /ncRNA_class="miRNA"
     primer_bind     4517..4536
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4776..4797
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      4839..4848
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(4903..5281)
                     /label=TRE3G promoter
                     /note="3rd-generation Tet-responsive promoter that can be 
                     activated by binding of Tet-On(R) 3G"
     protein_bind    5039..5057
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    5075..5093
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    5111..5129
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    5147..5165
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    5183..5201
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    5219..5237
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    5255..5273
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     complement(5462..5481)
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     promoter        5591..5802
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     CDS             5815..6558
                     /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             6568..6624
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             6625..7335
                     /codon_start=1
                     /product="monomeric red fluorescent protein derived from 
                     mRuby, with improved photophysical properties (Lam et al., 
                     2012)"
                     /label=mRuby2
                     /translation="MVSKGEELIKENMRMKVVMEGSVNGHQFKCTGEGEGNPYMGTQTM
                     RIKVIEGGPLPFAFDILATSFMYGSRTFIKYPKGIPDFFKQSFPEGFTWERVTRYEDGG
                     VVTVMQDTSLEDGCLVYHVQVRGVNFPSNGPVMQKKTKGWEPNTEMMYPADGGLRGYTH
                     MALKVDGGGHLSCSFVTTYRSKKTVGNIKMPGIHAVDHRLERLEESDNEMFVVQREHAV
                     AKFAGLGGGMDELYK"
     misc_feature    7349..7937
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(7402..7422)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(7820..7831)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             8029..8262
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    8334..8455
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(8371..8390)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     8425..8444
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      8495..8630
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(8650..8669)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(8651..8669)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(8679..8696)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(8679..8695)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(8688..8710)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      8837..9292
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(8924..8943)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     9134..9155
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        9318..9422
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(9423..9491,9492..10283)
                     /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(9641..9660)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      10454..11042
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     10943..10962
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     11196..11213
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    11330..11351
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(11366..11371,11372..11389,11390..11396)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    11404..11420
                     /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     11409..11431
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     11428..11444
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     11428..11444
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     11463..11483
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        11465..11483
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        11511..11737
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             11738..11918
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
ORIGIN
        1 acagggactt gaaagcgaaa gggaaaccag aggagctctc tcgacgcagg actcggcttg
       61 ctgaagcgcg cacggcaaga ggcgaggggc ggcgactggt gagtacgcca aaaattttga
      121 ctagcggagg ctagaaggag agagatgggt gcgagagcgt cagtattaag cgggggagaa
      181 ttagatcgcg atgggaaaaa attcggttaa ggccaggggg aaagaaaaaa tataaattaa
      241 aacatatagt atgggcaagc agggagctag aacgattcgc agttaatcct ggcctgttag
      301 aaacatcaga aggctgtaga caaatactgg gacagctaca accatccctt cagacaggat
      361 cagaagaact tagatcatta tataatacag tagcaaccct ctattgtgtg catcaaagga
      421 tagagataaa agacaccaag gaagctttag acaagataga ggaagagcaa aacaaaagta
      481 agaccaccgc acagcaagcg gccgctgatc ttcagacctg gaggaggaga tatgagggac
      541 aattggagaa gtgaattata taaatataaa gtagtaaaaa ttgaaccatt aggagtagca
      601 cccaccaagg caaagagaag agtggtgcag agagaaaaaa gagcagtggg aataggagct
      661 ttgttccttg ggttcttggg agcagcagga agcactatgg gcgcagcgtc aatgacgctg
      721 acggtacagg ccagacaatt attgtctggt atagtgcagc agcagaacaa tttgctgagg
      781 gctattgagg cgcaacagca tctgttgcaa ctcacagtct ggggcatcaa gcagctccag
      841 gcaagaatcc tggctgtgga aagataccta aaggatcaac agctcctggg gatttggggt
      901 tgctctggaa aactcatttg caccactgct gtgccttgga atgctagttg gagtaataaa
      961 tctctggaac agatttggaa tcacacgacc tggatggagt gggacagaga aattaacaat
     1021 tacacaagct taatacactc cttaattgaa gaatcgcaaa accagcaaga aaagaatgaa
     1081 caagaattat tggaattaga taaatgggca agtttgtgga attggtttaa cataacaaat
     1141 tggctgtggt atataaaatt attcataatg atagtaggag gcttggtagg tttaagaata
     1201 gtttttgctg tactttctat agtgaataga gttaggcagg gatattcacc attatcgttt
     1261 cagacccacc tcccaacccc gaggggaccc gacaggcccg aaggaataga agaagaaggt
     1321 ggagagagag acagagacag atccattcga ttagtgaacg gatctcgacg gtatcggtta
     1381 acttttaaaa gaaaaggggg gattgggggg tacagtgcag gggaaagaat agtagacata
     1441 atagcaacag acatacaaac taaagaatta caaaaacaaa ttacaaaatt caaaatttta
     1501 tcgtccagtg tggtggctac tagcgccata gagcccaccg catccccagc atgcctgcta
     1561 ttgtcttccc aatcctcccc cttgctgtcc tgccccaccc caccccccag aatagaatga
     1621 cacctactca gacaatgcga tgcaatttcc tcattttatt aggaaaggac agtgggagtg
     1681 gcaccttcca gggtcaagga aggcacgggg gaggggcaaa caacagatgg ctggcaacta
     1741 gaaggcacag gtgcaactcc tcctaggttt aattaaagac ttcaagcggc cacaagcttg
     1801 ttgttagggt actccattca ccagcggttg cagcccatct acgctgtctg gcttttcctt
     1861 cttctggtta cgatctttac ccgtgcgcag ccggctccct tcactggagg cactctgcag
     1921 ggatccatca gctgttcttc ctttagcctc tcttggatta cgtggacgat tatctgttcg
     1981 ggaatgatcg tcgtttcctt tgaagcctcc tcctcttcct cttcctcctt gtcctcttcc
     2041 tcctcctcca cgccgccgtc cgtctcctct gcgcaggaag ctctccctct cttcctctgt
     2101 tggagctaat gaccaatcac tgagttcgtc tctgtggtca gattctgttt cagaagcatt
     2161 tgatgcttca gaattagttc ctgaagtata tccaggaccg cgtctgccgt gccccctatt
     2221 tctgtaaggt ctactacctc gacccattcc ttgaccatca tcagtcacat agcctttttc
     2281 cttatctgta cgatttggtg gtggtctaga actagctcca atttgtcgca actgctcatc
     2341 aatttgtaat ctctccaaac gcaactggtc tacttccttt aaatagttca ggtgataatc
     2401 caaaagaaca gtggcattag cgatgctgtc ttttgttccc acaaaaacaa atggtaccat
     2461 accctgccaa gccttgggtt caggtttctg gggtccccct gctacaattg atgaaaccac
     2521 taacaccctc tggacttttg tactgttagg ttgagaaaaa tgggtgtttt cctttgtatc
     2581 taaatgtttc ttttcttcag aggagttagg tccaaccctt gaattattgg aaggtaggga
     2641 acttggtggc ataatttcct cttcttgtgg gacacttttc tcattttcag cctcaatcct
     2701 caccctcaca actcctgact tgtccacgat ctcttgaatc agctttccat tttttcctat
     2761 tactttgcct actaagtttc gtggaacctg tatgacatct tcagcaaatt ccagaaagct
     2821 tctagccttt ttcactgcat cttgatcctc tccataaata tgaaatgtgc aggtatcctc
     2881 atctaaatca atagcagtga cgccaggcac ttttctagct tgctgaatat tagcaccatg
     2941 agtaccaata gctaaaccca tcagatcttc tcgtacgata aactgttcat gaaatcttga
     3001 ggcaagctgc cttgaactct ccagttgttt actggcttct tcatttctca gtataagaga
     3061 caacttggtg cgcagacttc gaaagtgcat gtcaatcaac atgtgggctc gctttgaggt
     3121 gacttcattg atggacaaaa ttaccagctg ataattttct ggatcataag ttacagagaa
     3181 ggcaccaact gcctttttaa aatccttatg tgctgattct ttggcacaca tttgtcgtaa
     3241 atcttctggc acctccagct tgatcttatg gaaagtatct tttgtagcag gtttgttggg
     3301 attaacagat cgtagacgct caattgtgac aatttcatta tacgtagcat cacatgctgc
     3361 atattctatc acataaaact caccctttat catcctcact ttagctaacc accaacagca
     3421 aggctctttt tcatttgctc tggaataaac ctcaacttca tcactttcgt ttatatcttt
     3481 attataacct acaggtggtg ggaatctcac atcatggaat ggaatctgtc tctctggttg
     3541 ccagttgttt tcaaaagcaa ctgttataga atcttcatgg acatctttta caaatgcctt
     3601 gtagaaagcg ccattggagc cccgcacttc caccaccagc tccttgtcat cgtcgtcctt
     3661 gtagtcctcc atcgaattcg aagcttgagc tcgagatctg agtccggact tgtacagctc
     3721 gtccatgccg agagtgatcc cggcggcggt cacgaactcc agcaggacca tgtgatcgcg
     3781 cttctcgttg gggtctttgc tcagggcgga ctgggtgctc aggtagtggt tgtcgggcag
     3841 cagcacgggg ccgtcgccga tgggggtgtt ctgctggtag tggtcggcga gctgcacgct
     3901 gccgtcctcg atgttgtggc ggatcttgaa gttcaccttg atgccgttct tctgcttgtc
     3961 ggccatgata tagacgttgt ggctgttgta gttgtactcc agcttgtgcc ccaggatgtt
     4021 gccgtcctcc ttgaagtcga tgcccttcag ctcgatgcgg ttcaccaggg tgtcgccctc
     4081 gaacttcacc tgtgaaaaaa aaaaagaaga ctcgaagtta gtacaattga aaaaagtgat
     4141 ttaatttata ccattttaat tcagctttgt aaaaatgtat caaagagata gcaaggtatt
     4201 cagttttagt aaacaagata attgctccta aagtagcccc ttgaattcta gccccttgaa
     4261 gtccgaggca gtaggcaccg cttgaagtct ttaattaaat acatctgtgg cttcactatt
     4321 taattaaaga cttcaagcgg cgctcactgt caacagcaat ataccttctc gagccttctg
     4381 ttgggttaag aagtccctga agaagtaatc ccagcaagtg tttccaagat gtgcaggcaa
     4441 cgattctgta aagtactgaa gcctcattca acatagtata tgtgctgccg aagcgagcac
     4501 acttacctcg gcgcgggtct tgtagttgcc gtcgtccttg aagaagatgg tgcgctcctg
     4561 gacgtagcct tcgggcatgg cggacttgaa gaagtcgtgc tgcttcatgt ggtcggggta
     4621 gcggctgaag cactgcacgc cgtaggtcag ggtggtcacg agggtgggcc agggcacggg
     4681 cagcttgccg gtggtgcaga tgaacttcag ggtcagcttg ccgtaggtgg catcgccctc
     4741 gccctcgccg gacacgctga acttgtggcc gtttacgtcg ccgtccagct cgaccaggat
     4801 gggcaccacc ccggtgaaca gctcctcgcc cttgctcacc atggtggcca ttacagcgtg
     4861 gatggcgtct ccaggcgatc tgacggttca ctaaacgagc tctttacgag ggtaggaagt
     4921 ggtacggaaa gttggtataa gacaaaagtg ttgtggaatt gctccaggcg atctgacggt
     4981 tcactaaacg agctctgctt ttataggcgc ccaccgtaca cgcctaaagc ttatacgttc
     5041 tctatcactg atagggagta aactggatat acgttctcta tcactgatag ggagtaaact
     5101 gtagatacgt tctctatcac tgatagggag taaactggtc atacgttctc tatcactgat
     5161 agggagtaaa ctccttatac gttctctatc actgataggg agtaaagtct gcatacgttc
     5221 tctatcactg atagggagta aactcttcat acgttctcta tcactgatag ggagtaaact
     5281 cgaggaatag tacgaacatg gtagctagtg gatccttcgt cgacctgcag actggctgtg
     5341 tataagggag cctgacattt atattcccca gaacatcagg ttaatggcgt ttttgatgtc
     5401 attttcgcgg tggctgagat cagccacttc ttccccgata acggagaccg gcacactggc
     5461 gcatatcggt ggtcatcatg cgccagcttt catccccgat atgcaccacc gggtaaagtt
     5521 cacgcatatg aagctggaca tcacctccca caacctcgag gtggaacagt acgctgatat
     5581 ctcttatact gggcagagcg cacatcgccc acagtccccg agaagttggg gggaggggtc
     5641 ggcaattgaa ccggtgccta gagaaggtgg cgcggggtaa actgggaaag tgatgtcgtg
     5701 tactggctcc gcctttttcc cgagggtggg ggagaaccgt atataagtgc agtagtcgcc
     5761 gtgaacgttc tttttcgcaa cgggtttgcc gccagaacac agtaatgtgc caccatgagc
     5821 cggctggaca agagcaaagt gatcaacagc gccctggaac tgctgaacgg cgtgggcatc
     5881 gagggcctga ccacccggaa gctggcccag aaactgggcg tggaacagcc caccctgtac
     5941 tggcacgtga agaacaagcg ggccctgctg gacgccctgc ccatcgagat gctggaccgg
     6001 caccacaccc acagctgccc tctggaaggc gagagctggc aggacttcct gcggaacaac
     6061 gccaagagct acagatgcgc cctgctgagc caccgggacg gcgccaaagt gcacctgggc
     6121 accagaccca ccgagaagca gtacgagaca ctggaaaacc agctggcctt cctgtgccag
     6181 cagggcttca gcctggaaaa cgccctgtac gccctgagcg ccgtgggcca cttcaccctg
     6241 ggctgcgtgc tggaagaaca ggaacaccag gtcgccaaag aggaaagaga gacacccacc
     6301 accgacagca tgccccccct gctgaagcag gccatcgagc tgttcgacag acagggcgcc
     6361 gagcccgcct tcctgttcgg cctggaactg atcatctgcg gcctcgagaa gcagctgaag
     6421 tgcgagagcg gcggacccac cgacgccctg gacgacttcg acctggacat gctgcccgcc
     6481 gatgccctgg atgattttga tctggatatg ctccctgctg acgctctcga tgacttcgat
     6541 ctcgatatgc tgccaggcgg aagcggagct actaacttca gcctgctgaa gcaggctgga
     6601 gacgtggagg agaaccctgg acctatggtg tctaagggcg aagagctgat caaggaaaat
     6661 atgcgtatga aggtggtcat ggaaggttcg gtcaacggcc accaattcaa atgcacaggt
     6721 gaaggagaag gcaatccgta catgggaact caaaccatga ggatcaaagt catcgaggga
     6781 ggacccctgc catttgcctt tgacattctt gccacgtcgt tcatgtatgg cagccgtact
     6841 tttatcaagt acccgaaagg cattcctgat ttctttaaac agtcctttcc tgagggtttt
     6901 acttgggaaa gagttacgag atacgaagat ggtggagtcg tcaccgtcat gcaggacacc
     6961 agccttgagg atggctgtct cgtttaccac gtccaagtca gaggggtaaa ctttccctcc
     7021 aatggtcccg tgatgcagaa gaagaccaag ggttgggagc ctaatacaga gatgatgtat
     7081 ccagcagatg gtggtctgag gggatacact catatggcac tgaaagttga tggtggtggc
     7141 catctgtctt gctctttcgt aacaacttac aggtcaaaaa agaccgtcgg gaacatcaag
     7201 atgcccggta tccatgccgt tgatcaccgc ctggaaaggt tagaggaaag tgacaatgaa
     7261 atgttcgtag tacaacgcga acacgcagtt gccaagttcg ccgggcttgg tggtgggatg
     7321 gacgagctgt acaagtaaca acgccgataa tcaacctctg gattacaaaa tttgtgaaag
     7381 attgactggt attcttaact atgttgctcc ttttacgcta tgtggatacg ctgctttaat
     7441 gcctttgtat catgctattg cttcccgtat ggctttcatt ttctcctcct tgtataaatc
     7501 ctggttgctg tctctttatg aggagttgtg gcccgttgtc aggcaacgtg gcgtggtgtg
     7561 cactgtgttt gctgacgcaa cccccactgg ttggggcatt gccaccacct gtcagctcct
     7621 ttccgggact ttcgctttcc ccctccctat tgccacggcg gaactcatcg ccgcctgcct
     7681 tgcccgctgc tggacagggg ctcggctgtt gggcactgac aattccgtgg tgttgtcggg
     7741 gaaatcatcg tcctttcctt ggctgctcgc ctgtgttgcc acctggattc tgcgcgggac
     7801 gtccttctgc tacgtccctt cggccctcaa tccagcggac cttccttccc gcggcctgct
     7861 gccggctctg cggcctcttc cgcgtcttcg ccttcgccct cagacgagtc ggatctccct
     7921 ttgggccgcc tccccgccgc tcgctgatat agggcccggt acctttaaga ccaatgactt
     7981 acaaggcagc tgtagatctt agccactttt taaaagaaaa ggggggactg gaagggctag
     8041 ctcactccca acgaagacaa gatctgcttt ttgcttgtac tgggtctctc tggttagacc
     8101 agatctgagc ctgggagctc tctggctaac tagggaaccc actgcttaag cctcaataaa
     8161 gcttgccttg agtgcttcaa gtagtgtgtg cccgtctgtt gtgtgactct ggtaactaga
     8221 gatccctcag acccttttag tcagtgtgga aaatctctag cagtagtagt tcatgtcatc
     8281 ttattattca gtatttataa cttgcaaaga aatgaatatc agagagtgag aggaacttgt
     8341 ttattgcagc ttataatggt tacaaataaa gcaatagcat cacaaatttc acaaataaag
     8401 catttttttc actgcattct agttgtggtt tgtccaaact catcaatgta tcttatcatg
     8461 tctggctcta gctatcccgc ccctaactcc gcccatcccg cccctaactc cgcccagttc
     8521 cgcccattct ccgccccatg gctgactaat tttttttatt tatgcagagg ccgaggccgc
     8581 ctcggcctct gagctattcc agaagtagtg aggaggcttt tttggaggcc tagggacgta
     8641 cccaattcgc cctatagtga gtcgtattac gcgcgctcac tggccgtcgt tttacaacgt
     8701 cgtgactggg aaaaccctgg cgttacccaa cttaatcgcc ttgcagcaca tccccctttc
     8761 gccagctggc gtaatagcga agaggcccgc accgatcgcc cttcccaaca gttgcgcagc
     8821 ctgaatggcg aatgggacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt
     8881 acgcgcagcg tgaccgctac acttgccagc gccctagcgc ccgctccttt cgctttcttc
     8941 ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct
     9001 ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga ttagggtgat
     9061 ggttcacgta gtgggccatc gccctgatag acggtttttc gccctttgac gttggagtcc
     9121 acgttcttta atagtggact cttgttccaa actggaacaa cactcaaccc tatctcggtc
     9181 tattcttttg atttataagg gattttgccg atttcggcct attggttaaa aaatgagctg
     9241 atttaacaaa aatttaacgc gaattttaac aaaatattaa cgcttacaat ttaggtggca
     9301 cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata
     9361 tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga
     9421 gtatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca ttttgccttc
     9481 ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat cagttgggtg
     9541 cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag agttttcgcc
     9601 ccgaagaacg ttttccaatg atgagcactt ttaaagttct gctatgtggc gcggtattat
     9661 cccgtattga cgccgggcaa gagcaactcg gtcgccgcat acactattct cagaatgact
     9721 tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca gtaagagaat
     9781 tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt ctgacaacga
     9841 tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat gtaactcgcc
     9901 ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt gacaccacga
     9961 tgcctgtagc aatggcaaca acgttgcgca aactattaac tggcgaacta cttactctag
    10021 cttcccggca acaattaata gactggatgg aggcggataa agttgcagga ccacttctgc
    10081 gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt gagcgtgggt
    10141 ctcgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc gtagttatct
    10201 acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct gagataggtg
    10261 cctcactgat taagcattgg taactgtcag accaagttta ctcatatata ctttagattg
    10321 atttaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttttt gataatctca
    10381 tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc gtagaaaaga
    10441 tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa
    10501 aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga
    10561 aggtaactgg cttcagcaga gcgcagatac caaatactgt tcttctagtg tagccgtagt
    10621 taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt
    10681 taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat
    10741 agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct
    10801 tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca
    10861 cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag
    10921 agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc
    10981 gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga
    11041 aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca
    11101 tgttctttcc tgcgttatcc cctgattctg tggataaccg tattaccgcc tttgagtgag
    11161 ctgataccgc tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc gaggaagcgg
    11221 aagagcgccc aatacgcaaa ccgcctctcc ccgcgcgttg gccgattcat taatgcagct
    11281 ggcacgacag gtttcccgac tggaaagcgg gcagtgagcg caacgcaatt aatgtgagtt
    11341 agctcactca ttaggcaccc caggctttac actttatgct tccggctcgt atgttgtgtg
    11401 gaattgtgag cggataacaa tttcacacag gaaacagcta tgaccatgat tacgccaagc
    11461 gcgcaattaa ccctcactaa agggaacaaa agctggagct gcaagcttaa tgtagtctta
    11521 tgcaatactc ttgtagtctt gcaacatggt aacgatgagt tagcaacatg ccttacaagg
    11581 agagaaaaag caccgtgcat gccgattggt ggaagtaagg tggtacgatc gtgccttatt
    11641 aggaaggcaa cagacgggtc tgacatggat tggacgaacc actgaattgc cgcattgcag
    11701 agatattgta tttaagtgcc tagctcgata cataaacggg tctctctggt tagaccagat
    11761 ctgagcctgg gagctctctg gctaactagg gaacccactg cttaagcctc aataaagctt
    11821 gccttgagtg cttcaagtag tgtgtgcccg tctgttgtgt gactctggta actagagatc
    11881 cctcagaccc ttttagtcag tgtggaaaat ctctagcagt ggcgcccga
//
