LOCUS       sequence-332422-        9124 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses programmed gRNA scaffold from U6 promoter. Programmed gRNA
            scaffold for streamlined scRNA-seq in CRISPR screen (Direct-seq). 
            3rd generation lentiviral backbone..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9124)
  AUTHORS   Song Q, Ni K, Liu M, Li Y, Wang L, Wang Y, Liu Y, Yu Z, Qi Y, Lu Z, 
            Ma L
  TITLE     Direct-seq: programmed gRNA scaffold for streamlined scRNA-seq in 
            CRISPR screen.
  JOURNAL   Genome Biol. 2020 Jun 8;21(1):136. doi: 10.1186/s13059-020-02044-w.
  PUBMED    32513233
REFERENCE   2  (bases 1 to 9124)
  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: "Genome 
            Biol."; date: "2020-06-8"; volume: "21(1):136. doi"; pages: " 
            10.1186/s13059-020-02044-w"
COMMENT     Sequence Label: Direct-seq_lentiGuide-Puro-Loop2-8A8G
FEATURES             Location/Qualifiers
     source          1..9124
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        95..1273
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          326..1264
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1221..1241
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     CDS             1286..1882
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(1286..1305)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     1782..1802
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     CDS             1883..1936
                     /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             1937..2632
                     /codon_start=1
                     /product="monomeric far-red fluorescent protein (Shcherbo 
                     et al., 2009)"
                     /label=mKate2
                     /note="mammalian codon-optimized"
                     /translation="VSELIKENMHMKLYMEGTVNNHHFKCTSEGEGKPYEGTQTMRIKA
                     VEGGPLPFAFDILATSFMYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLTA
                     TQDTSLQDGCLIYNVKIRGVNFPSNGPVMQKKTLGWEASTETLYPADGGLEGRADMALK
                     LVGGGHLICNLKTTYRSKKPAKNLKMPGVYYVDRRLERIKEADKETYVEQHEVAVARYC
                     DLPSKLGHR"
     misc_feature    2654..3242
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2707..2727)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3125..3136)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             3314..3547
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    3625..3746
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3662..3681)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3716..3735
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      3786..3921
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(3941..3960)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3942..3960)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3970..3987)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3970..3986)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3979..4001)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      4128..4583
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4215..4234)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4425..4446
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        4609..4713
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4714..4782,4783..5574)
                     /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(4932..4951)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5745..6333
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6234..6253
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6487..6504
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    6621..6642
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(6657..6662,6663..6680,6681..6687)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6695..6711
                     /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     6700..6722
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     6719..6735
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6719..6735
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     6754..6774
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        6756..6774
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        6802..7028
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             7029..7209
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    7256..7381
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    7874..8107
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             8292..8336
                     /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"
     promoter        8514..8762
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     8514..8534
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     8685..8704
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_feature    8963..9080
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
ORIGIN
        1 cgagggggcc cgggtgcaaa gatggataaa gttttaaaca gagaggaatc tttgcagcta
       61 atggaccttc taggtcttga aaggagtggg aattggctcc ggtgcccgtc agtgggcaga
      121 gcgcacatcg cccacagtcc ccgagaagtt ggggggaggg gtcggcaatt gatccggtgc
      181 ctagagaagg tggcgcgggg taaactggga aagtgatgtc gtgtactggc tccgcctttt
      241 tcccgagggt gggggagaac cgtatataag tgcagtagtc gccgtgaacg ttctttttcg
      301 caacgggttt gccgccagaa cacaggtaag tgccgtgtgt ggttcccgcg ggcctggcct
      361 ctttacgggt tatggccctt gcgtgccttg aattacttcc acctggctgc agtacgtgat
      421 tcttgatccc gagcttcggg ttggaagtgg gtgggagagt tcgaggcctt gcgcttaagg
      481 agccccttcg cctcgtgctt gagttgaggc ctggcctggg cgctggggcc gccgcgtgcg
      541 aatctggtgg caccttcgcg cctgtctcgc tgctttcgat aagtctctag ccatttaaaa
      601 tttttgatga cctgctgcga cgcttttttt ctggcaagat agtcttgtaa atgcgggcca
      661 agatctgcac actggtattt cggtttttgg ggccgcgggc ggcgacgggg cccgtgcgtc
      721 ccagcgcaca tgttcggcga ggcggggcct gcgagcgcgg ccaccgagaa tcggacgggg
      781 gtagtctcaa gctggccggc ctgctctggt gcctggcctc gcgccgccgt gtatcgcccc
      841 gccctgggcg gcaaggctgg cccggtcggc accagttgcg tgagcggaaa gatggccgct
      901 tcccggccct gctgcaggga gctcaaaatg gaggacgcgg cgctcgggag agcgggcggg
      961 tgagtcaccc acacaaagga aaagggcctt tccgtcctca gccgtcgctt catgtgactc
     1021 cacggagtac cgggcgccgt ccaggcacct cgattagttc tcgagctttt ggagtacgtc
     1081 gtctttaggt tggggggagg ggttttatgc gatggagttt ccccacactg agtgggtgga
     1141 gactgaagtt aggccagctt ggcacttgat gtaattctcc ttggaatttg ccctttttga
     1201 gtttggatct tggttcattc tcaagcctca gacagtggtt caaagttttt ttcttccatt
     1261 tcaggtgtcg tgacgtacgg ccaccatgac cgagtacaag cccacggtgc gcctcgccac
     1321 ccgcgacgac gtccccaggg ccgtacgcac cctcgccgcc gcgttcgccg actaccccgc
     1381 cacgcgccac accgtcgatc cggaccgcca catcgagcgg gtcaccgagc tgcaagaact
     1441 cttcctcacg cgcgtcgggc tcgacatcgg caaggtgtgg gtcgcggacg acggcgccgc
     1501 cgtggcggtc tggaccacgc cggagagcgt cgaagcgggg gcggtgttcg ccgagatcgg
     1561 cccgcgcatg gccgagttga gcggttcccg gctggccgcg cagcaacaga tggaaggcct
     1621 cctggcgccg caccggccca aggagcccgc gtggttcctg gccaccgtcg gagtctcgcc
     1681 cgaccaccag ggcaagggtc tgggcagcgc cgtcgtgctc cccggagtgg aggcggccga
     1741 gcgcgccggg gtgcccgcct tcctggagac ctccgcgccc cgcaacctcc ccttctacga
     1801 gcggctcggc ttcaccgtca ccgccgacgt cgaggtgccc gaaggaccgc gcacctggtg
     1861 catgacccgc aagcccggtg ccgagggcag gggaagtctt ctaacatgcg gggacgtgga
     1921 ggaaaatccc ggccccgtga gcgagctgat taaggagaac atgcacatga agctgtacat
     1981 ggagggcacc gtgaacaacc accacttcaa gtgcacatcc gagggcgaag gcaagcccta
     2041 cgagggcacc cagaccatga gaatcaaggc ggtcgagggc ggccctctcc ccttcgcctt
     2101 cgacatcctg gctaccagct tcatgtacgg cagcaaaacc ttcatcaacc acacccaggg
     2161 catccccgac ttctttaagc agtccttccc cgagggcttc acatgggaga gagtcaccac
     2221 atacgaagac gggggcgtgc tgaccgctac ccaggacacc agcctccagg acggctgcct
     2281 catctacaac gtcaagatca gaggggtgaa cttcccatcc aacggccctg tgatgcagaa
     2341 gaaaacactc ggctgggagg cctccaccga gaccctgtac cccgctgacg gcggcctgga
     2401 aggcagagcc gacatggccc tgaagctcgt gggcgggggc cacctgatct gcaacttgaa
     2461 gaccacatac agatccaaga aacccgctaa gaacctcaag atgcccggcg tctactatgt
     2521 ggacagaaga ctggaaagaa tcaaggaggc cgacaaagag acctacgtcg agcagcacga
     2581 ggtggctgtg gccagatact gcgacctccc tagcaaactg gggcacagat aaggatccac
     2641 gcgttaagtc gacaatcaac ctctggatta caaaatttgt gaaagattga ctggtattct
     2701 taactatgtt gctcctttta cgctatgtgg atacgctgct ttaatgcctt tgtatcatgc
     2761 tattgcttcc cgtatggctt tcattttctc ctccttgtat aaatcctggt tgctgtctct
     2821 ttatgaggag ttgtggcccg ttgtcaggca acgtggcgtg gtgtgcactg tgtttgctga
     2881 cgcaaccccc actggttggg gcattgccac cacctgtcag ctcctttccg ggactttcgc
     2941 tttccccctc cctattgcca cggcggaact catcgccgcc tgccttgccc gctgctggac
     3001 aggggctcgg ctgttgggca ctgacaattc cgtggtgttg tcggggaaat catcgtcctt
     3061 tccttggctg ctcgcctgtg ttgccacctg gattctgcgc gggacgtcct tctgctacgt
     3121 cccttcggcc ctcaatccag cggaccttcc ttcccgcggc ctgctgccgg ctctgcggcc
     3181 tcttccgcgt cttcgccttc gccctcagac gagtcggatc tccctttggg ccgcctcccc
     3241 gcgtcgactt taagaccaat gacttacaag gcagctgtag atcttagcca ctttttaaaa
     3301 gaaaaggggg gactggaagg gctaattcac tcccaacgaa gacaagatct gctttttgct
     3361 tgtactgggt ctctctggtt agaccagatc tgagcctggg agctctctgg ctaactaggg
     3421 aacccactgc ttaagcctca ataaagcttg ccttgagtgc ttcaagtagt gtgtgcccgt
     3481 ctgttgtgtg actctggtaa ctagagatcc ctcagaccct tttagtcagt gtggaaaatc
     3541 tctagcagta cgtatagtag ttcatgtcat cttattattc agtatttata acttgcaaag
     3601 aaatgaatat cagagagtga gaggaacttg tttattgcag cttataatgg ttacaaataa
     3661 agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt
     3721 ttgtccaaac tcatcaatgt atcttatcat gtctggctct agctatcccg cccctaactc
     3781 cgcccatccc gcccctaact ccgcccagtt ccgcccattc tccgccccat ggctgactaa
     3841 ttttttttat ttatgcagag gccgaggccg cctcggcctc tgagctattc cagaagtagt
     3901 gaggaggctt ttttggaggc ctagggacgt acccaattcg ccctatagtg agtcgtatta
     3961 cgcgcgctca ctggccgtcg ttttacaacg tcgtgactgg gaaaaccctg gcgttaccca
     4021 acttaatcgc cttgcagcac atcccccttt cgccagctgg cgtaatagcg aagaggcccg
     4081 caccgatcgc ccttcccaac agttgcgcag cctgaatggc gaatgggacg cgccctgtag
     4141 cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag
     4201 cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt
     4261 tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg ctttacggca
     4321 cctcgacccc aaaaaacttg attagggtga tggttcacgt agtgggccat cgccctgata
     4381 gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac tcttgttcca
     4441 aactggaaca acactcaacc ctatctcggt ctattctttt gatttataag ggattttgcc
     4501 gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg cgaattttaa
     4561 caaaatatta acgcttacaa tttaggtggc acttttcggg gaaatgtgcg cggaacccct
     4621 atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgagaca ataaccctga
     4681 taaatgcttc aataatattg aaaaaggaag agtatgagta ttcaacattt ccgtgtcgcc
     4741 cttattccct tttttgcggc attttgcctt cctgtttttg ctcacccaga aacgctggtg
     4801 aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg gttacatcga actggatctc
     4861 aacagcggta agatccttga gagttttcgc cccgaagaac gttttccaat gatgagcact
     4921 tttaaagttc tgctatgtgg cgcggtatta tcccgtattg acgccgggca agagcaactc
     4981 ggtcgccgca tacactattc tcagaatgac ttggttgagt actcaccagt cacagaaaag
     5041 catcttacgg atggcatgac agtaagagaa ttatgcagtg ctgccataac catgagtgat
     5101 aacactgcgg ccaacttact tctgacaacg atcggaggac cgaaggagct aaccgctttt
     5161 ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa
     5221 gccataccaa acgacgagcg tgacaccacg atgcctgtag caatggcaac aacgttgcgc
     5281 aaactattaa ctggcgaact acttactcta gcttcccggc aacaattaat agactggatg
     5341 gaggcggata aagttgcagg accacttctg cgctcggccc ttccggctgg ctggtttatt
     5401 gctgataaat ctggagccgg tgagcgtggg tctcgcggta tcattgcagc actggggcca
     5461 gatggtaagc cctcccgtat cgtagttatc tacacgacgg ggagtcaggc aactatggat
     5521 gaacgaaata gacagatcgc tgagataggt gcctcactga ttaagcattg gtaactgtca
     5581 gaccaagttt actcatatat actttagatt gatttaaaac ttcattttta atttaaaagg
     5641 atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg tgagttttcg
     5701 ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga tccttttttt
     5761 ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg
     5821 ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag agcgcagata
     5881 ccaaatactg ttcttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca
     5941 ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag
     6001 tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc
     6061 tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga
     6121 tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg
     6181 tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc agggggaaac
     6241 gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg tcgatttttg
     6301 tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg
     6361 ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc ccctgattct
     6421 gtggataacc gtattaccgc ctttgagtga gctgataccg ctcgccgcag ccgaacgacc
     6481 gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc caatacgcaa accgcctctc
     6541 cccgcgcgtt ggccgattca ttaatgcagc tggcacgaca ggtttcccga ctggaaagcg
     6601 ggcagtgagc gcaacgcaat taatgtgagt tagctcactc attaggcacc ccaggcttta
     6661 cactttatgc ttccggctcg tatgttgtgt ggaattgtga gcggataaca atttcacaca
     6721 ggaaacagct atgaccatga ttacgccaag cgcgcaatta accctcacta aagggaacaa
     6781 aagctggagc tgcaagctta atgtagtctt atgcaatact cttgtagtct tgcaacatgg
     6841 taacgatgag ttagcaacat gccttacaag gagagaaaaa gcaccgtgca tgccgattgg
     6901 tggaagtaag gtggtacgat cgtgccttat taggaaggca acagacgggt ctgacatgga
     6961 ttggacgaac cactgaattg ccgcattgca gagatattgt atttaagtgc ctagctcgat
     7021 acataaacgg gtctctctgg ttagaccaga tctgagcctg ggagctctct ggctaactag
     7081 ggaacccact gcttaagcct caataaagct tgccttgagt gcttcaagta gtgtgtgccc
     7141 gtctgttgtg tgactctggt aactagagat ccctcagacc cttttagtca gtgtggaaaa
     7201 tctctagcag tggcgcccga acagggactt gaaagcgaaa gggaaaccag aggagctctc
     7261 tcgacgcagg actcggcttg ctgaagcgcg cacggcaaga ggcgaggggc ggcgactggt
     7321 gagtacgcca aaaattttga ctagcggagg ctagaaggag agagatgggt gcgagagcgt
     7381 cagtattaag cgggggagaa ttagatcgcg atgggaaaaa attcggttaa ggccaggggg
     7441 aaagaaaaaa tataaattaa aacatatagt atgggcaagc agggagctag aacgattcgc
     7501 agttaatcct ggcctgttag aaacatcaga aggctgtaga caaatactgg gacagctaca
     7561 accatccctt cagacaggat cagaagaact tagatcatta tataatacag tagcaaccct
     7621 ctattgtgtg catcaaagga tagagataaa agacaccaag gaagctttag acaagataga
     7681 ggaagagcaa aacaaaagta agaccaccgc acagcaagcg gccgctgatc ttcagacctg
     7741 gaggaggaga tatgagggac aattggagaa gtgaattata taaatataaa gtagtaaaaa
     7801 ttgaaccatt aggagtagca cccaccaagg caaagagaag agtggtgcag agagaaaaaa
     7861 gagcagtggg aataggagct ttgttccttg ggttcttggg agcagcagga agcactatgg
     7921 gcgcagcgtc aatgacgctg acggtacagg ccagacaatt attgtctggt atagtgcagc
     7981 agcagaacaa tttgctgagg gctattgagg cgcaacagca tctgttgcaa ctcacagtct
     8041 ggggcatcaa gcagctccag gcaagaatcc tggctgtgga aagataccta aaggatcaac
     8101 agctcctggg gatttggggt tgctctggaa aactcatttg caccactgct gtgccttgga
     8161 atgctagttg gagtaataaa tctctggaac agatttggaa tcacacgacc tggatggagt
     8221 gggacagaga aattaacaat tacacaagct taatacactc cttaattgaa gaatcgcaaa
     8281 accagcaaga aaagaatgaa caagaattat tggaattaga taaatgggca agtttgtgga
     8341 attggtttaa cataacaaat tggctgtggt atataaaatt attcataatg atagtaggag
     8401 gcttggtagg tttaagaata gtttttgctg tactttctat agtgaataga gttaggcagg
     8461 gatattcacc attatcgttt cagacccacc tcccaacccc gaggggaccc agagagggcc
     8521 tatttcccat gattccttca tatttgcata tacgatacaa ggctgttaga gagataatta
     8581 gaattaattt gactgtaaac acaaagatat tagtacaaaa tacgtgacgt agaaagtaat
     8641 aatttcttgg gtagtttgca gttttaaaat tatgttttaa aatggactat catatgctta
     8701 ccgtaacttg aaagtatttc gatttcttgg ctttatatat cttgtggaaa ggacgaaaca
     8761 ccggagacga gtgatactcg tctctgtttg agagctatgc tggaaacagc atagcaagtt
     8821 caaataaggc tagtccgtta tcaacttggc ccgaaaaaaa agaaaaaaag aaaaaaagaa
     8881 aaaggccaag tggcaccgag tcggtgcttt tttttgaatt ccgtaactag atcttgagac
     8941 aaatggcagt attcatccac aattttaaaa gaaaaggggg gattgggggg tacagtgcag
     9001 gggaaagaat agtagacata atagcaacag acatacaaac taaagaatta caaaaacaaa
     9061 ttacaaaaat tcaaaatttt cgggtttatt acagggacag cagagatcca ctttggcgcc
     9121 ggct
//
