pDEST-hisMBP-AsCpf1-EC

LOCUS      pDEST-hisMBP-AsCpf1-EC       10644 bp    DNA     circular SYN 14-APR-2025
DEFINITION  Expressing his-MBP tagged AsCpf1 (E.coli codon optimized).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 10644)
  AUTHORS   Hur JK, Kim K, Been KW, Baek G, Ye S, Hur JW, Ryu SM, Lee YS, Kim JS
  TITLE     Targeted mutagenesis in mice by electroporation of Cpf1 
            ribonucleoproteins.
  JOURNAL   Nat Biotechnol. 2016 Jun 6. doi: 10.1038/nbt.3596.
  PUBMED    27272385
REFERENCE   2  (bases 1 to 10644)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.biofeng.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat 
            Biotechnol. 2016 Jun 6. doi: 10.1038/nbt.3596."
COMMENT     Sequence Label: pDEST-hisMBP-AsCpf1-EC
FEATURES             Location/Qualifiers
     source          1..10644
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(1..6,7..22,23..29)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    37..53
                     /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     42..64
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     CDS             132..149
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     regulatory      1098..1107
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     1215..1238
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     protein_bind    1251..1275
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             1290..1310
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1317..5237
                     /codon_start=1
                     /product="CRISPR-associated protein from Acidaminococcus 
                     sp. BV3L6 (Zetsche et al., 2015)"
                     /label=AsCpf1
                     /translation="MTQFEGFTNLYQVSKTLRFELIPQGKTLKHIQEQGFIEEDKARND
                     HYKELKPIIDRIYKTYADQCLQLVQLDWENLSAAIDSYRKEKTEETRNALIEEQATYRN
                     AIHDYFIGRTDNLTDAINKRHAEIYKGLFKAELFNGKVLKQLGTVTTTEHENALLRSFD
                     KFTTYFSGFYENRKNVFSAEDISTAIPHRIVQDNFPKFKENCHIFTRLITAVPSLREHF
                     ENVKKAIGIFVSTSIEEVFSFPFYNQLLTQTQIDLYNQLLGGISREAGTEKIKGLNEVL
                     NLAIQKNDETAHIIASLPHRFIPLFKQILSDRNTLSFILEEFKSDEEVIQSFCKYKTLL
                     RNENVLETAEALFNELNSIDLTHIFISHKKLETISSALCDHWDTLRNALYERRISELTG
                     KITKSAKEKVQRSLKHEDINLQEIISAAGKELSEAFKQKTSEILSHAHAALDQPLPTTL
                     KKQEEKEILKSQLDSLLGLYHLLDWFAVDESNEVDPEFSARLTGIKLEMEPSLSFYNKA
                     RNYATKKPYSVEKFKLNFQMPTLASGWDVNKEKNNGAILFVKNGLYYLGIMPKQKGRYK
                     ALSFEPTEKTSEGFDKMYYDYFPDAAKMIPKCSTQLKAVTAHFQTHTTPILLSNNFIEP
                     LEITKEIYDLNNPEKEPKKFQTAYAKKTGDQKGYREALCKWIDFTRDFLSKYTKTTSID
                     LSSLRPSSQYKDLGEYYAELNPLLYHISFQRIAEKEIMDAVETGKLYLFQIYNKDFAKG
                     HHGKPNLHTLYWTGLFSPENLAKTSIKLNGQAELFYRPKSRMKRMAHRLGEKMLNKKLK
                     DQKTPIPDTLYQELYDYVNHRLSHDLSDEARALLPNVITKEVSHEIIKDRRFTSDKFFF
                     HVPITLNYQAANSPSKFNQRVNAYLKEHPETPIIGIDRGERNLIYITVIDSTGKILEQR
                     SLNTIQQFDYQKKLDNREKERVAARQAWSVVGTIKDLKQGYLSQVIHEIVDLMIHYQAV
                     VVLENLNFGFKSKRTGIAEKAVYQQFEKMLIDKLNCLVLKDYPAEKVGGVLNPYQLTDQ
                     FTSFAKMGTQSGFLFYVPAPYTSKIDPLTGFVDPFVWKTIKNHESRKHFLEGFDFLHYD
                     VKTGDFILHFKMNRNLSFQRGLPGFMPAWDIVFEKNETQFDAKGTPFIAGKRIVPVIEN
                     HRFTGRYRDLYPANELIALLEEKGIVFRDGSNILPKLLENDDSHAIDTMVALIRSVLQM
                     RNSNAATGEDYINSPVRDLNGVCFDSRFQNPEWPMDADANGAYHIALKGQLLLNHLKES
                     KDLKLQNGISNQDWLAYIQELRN"
     CDS             5238..5258
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             5265..5291
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /note="This is a variant of the standard HA tag, which has 
                     sequence YPYDVPDYA. Some anti-HA antibodies may not 
                     recognize this variant."
                     /translation="YPYDVPDYA"
     CDS             5265..5291
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     protein_bind    complement(5296..5320)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     complement(5332..5349)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5332..5348)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5341..5363)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(5527..5544)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      5697..5783
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      5875..5902
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        5921..6012
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6013..6081,6082..6873)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(6231..6250)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      complement(6915..7428)
                     /direction=LEFT
                     /label=M13 ori
                     /note="M13 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(7110..7131)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     7322..7341
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     rep_origin      7539..8127
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8028..8047
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     8281..8298
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    8313..8453
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(8379..8398)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     8539..8561
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(8555..8746)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     promoter        9242..9319
                     /gene="lacI (mutant)"
                     /label=lacIq promoter
                     /note="In the lacIq allele, a single base change in the 
                     promoter boosts expression of the lacI gene about 10-fold."
     CDS             9320..10402
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     complement(9339..9358)
                     /label=LacI-R
                     /note="LacI, reverse primer"
ORIGIN
        1 ttgacaatta atcatcggct cgtataatgt gtggaattgt gagcggataa caatttcaca
       61 caggaaacag ccagtccgtt taggtgtttt cacgagcact tcaccaacaa ggaccataga
      121 ttatgaaaat ccatcaccat caccatcacg aagaaggtaa actggtaatc tggattaacg
      181 gcgataaagg ctataacggt ctcgctgaag tcggtaagaa attcgagaaa gataccggaa
      241 ttaaagtcac cgttgagcat ccggataaac tggaagagaa attcccacag gttgcggcaa
      301 ctggcgatgg ccctgacatt atcttctggg cacacgaccg ctttggtggc tacgctcaat
      361 ctggcctgtt ggctgaaatc accccggaca aagcgttcca ggacaagctg tatccgttta
      421 cctgggatgc cgtacgttac aacggcaagc tgattgctta cccgatcgct gttgaagcgt
      481 tatcgctgat ttataacaaa gatctgctgc cgaacccgcc aaaaacctgg gaagagatcc
      541 cggcgctgga taaagaactg aaagcgaaag gtaagagcgc gctgatgttc aacctgcaag
      601 aaccgtactt cacctggccg ctgattgctg ctgacggggg ttatgcgttc aagtatgaaa
      661 acggcaagta cgacattaaa gacgtgggcg tggataacgc tggcgcgaaa gcgggtctga
      721 ccttcctggt tgacctgatt aaaaacaaac acatgaatgc agacaccgat tactccatcg
      781 cagaagctgc ctttaataaa ggcgaaacag cgatgaccat caacggcccg tgggcatggt
      841 ccaacatcga caccagcaaa gtgaattatg gtgtaacggt actgccgacc ttcaagggtc
      901 aaccatccaa accgttcgtt ggcgtgctga gcgcaggtat taacgccgcc agtccgaaca
      961 aagagctggc aaaagagttc ctcgaaaact atctgctgac tgatgaaggt ctggaagcgg
     1021 ttaataaaga caaaccgctg ggtgccgtag cgctgaagtc ttacgaggaa gagttggcga
     1081 aagatccacg tattgccgcc accatggaaa acgcccagaa aggtgaaatc atgccgaaca
     1141 tcccgcagat gtccgctttc tggtatgccg tgcgtactgc ggtgatcaac gccgccagcg
     1201 gtcgtcagac tgtcgatgaa gccctgaaag acgcgcagac taattcgatc acaagtttgt
     1261 acaaaaaagc aggcttcatg ggtgctcctc caaaaaagaa gagaaaggta ggtggtatga
     1321 cacagtttga aggcttcacc aatctctacc aggtcagcaa gacgctacgt tttgagctta
     1381 tcccgcaggg aaaaaccctg aaacacattc aggaacaggg gttcatagag gaagataagg
     1441 cgcgtaacga ccattataaa gaactgaagc ctataatcga ccgtatttat aaaacgtacg
     1501 cggatcagtg cctgcagctg gttcagctgg attgggagaa tctgtccgcg gctattgata
     1561 gctatcgcaa agagaagacc gaggaaaccc gtaacgcact gattgaagag caggcgacct
     1621 atcggaatgc gatccatgat tacttcatcg gccgcaccga caacctgacc gatgcaatta
     1681 acaaacgtca cgcagagatt tacaaaggtc tgtttaaagc agagttattc aatggcaagg
     1741 ttctgaaaca gctgggtacg gtcaccacca ccgaacacga aaacgcactg ctgaggagct
     1801 ttgataaatt taccacatat ttcagcggtt tctatgaaaa tcgtaagaat gtatttagcg
     1861 ccgaagatat ttccaccgca attcctcatc gtattgtgca ggataatttt ccgaagttta
     1921 aagaaaattg tcatattttt acccgtctga tcaccgcggt accgagcctg cgagagcatt
     1981 ttgaaaacgt taagaaagcc attggaattt ttgtcagtac cagcattgaa gaagtgtttt
     2041 cgttcccgtt ctataaccaa ctgctgaccc agacccagat tgatctgtac aatcagctgc
     2101 tggggggcat aagccgcgag gcaggtaccg aaaagataaa gggactcaat gaggtgctga
     2161 atctggcaat tcagaagaat gatgaaacgg ctcatatcat tgctagcctg ccgcatcgtt
     2221 tcattcccct gtttaagcaa atcctgagcg atcgcaatac actgagcttt atcctcgaag
     2281 agtttaaatc ggacgaagaa gttatccaga gcttttgcaa atacaaaacc ctgctgcgga
     2341 acgaaaatgt gctggagacc gctgaagcac tgtttaatga actgaactcg atcgacctca
     2401 cccatatttt tatatcccac aaaaaactgg aaaccataag cagcgctctg tgtgaccatt
     2461 gggataccct gcgcaacgcc ctgtatgaac ggcgtatcag cgagctgacc gggaaaatca
     2521 ccaaatccgc aaaggaaaaa gttcagcgta gtctgaaaca cgaggacatc aacctgcaag
     2581 aaattattag cgcagcaggt aaagagctga gcgaagcatt caaacagaaa accagcgaaa
     2641 tcctgagcca tgcccatgct gcactggatc agccgctgcc gaccaccctg aaaaaacagg
     2701 aggaaaagga gattctgaaa agccaactgg acagcctgct gggcctgtat cacctgctgg
     2761 actggtttgc agtcgatgag agcaacgagg ttgatcctga gttctccgct cgtctgaccg
     2821 gaatcaagct ggagatggaa ccgagtctgt cgttttacaa taaagcgcgt aattacgcga
     2881 ccaagaaacc gtatagcgtg gaaaaattca aactgaactt tcagatgccg acccttgcaa
     2941 gcggatggga cgttaacaaa gaaaaaaaca atggggcaat tctgtttgtg aaaaatggcc
     3001 tctattatct gggtatcatg ccgaaacaga aagggcgcta caaagccctg tcatttgagc
     3061 cgaccgagaa aacctcagag ggtttcgaca agatgtacta cgattatttc ccggatgcgg
     3121 caaaaatgat acccaaatgt agcacccaac tgaaggcagt tacagcccac tttcagaccc
     3181 ataccacccc gatcctgctg tcgaacaatt ttatagagcc gctggaaatt accaaagaga
     3241 tttatgatct gaataatccg gaaaaggagc ccaagaaatt tcagacggcg tatgcaaaaa
     3301 agaccgggga tcagaaaggt tatcgtgaag cgctgtgcaa atggattgac tttacccgtg
     3361 actttctgtc aaaatatacc aaaacgacga gcattgatct gagcagccta cgtccgagca
     3421 gccaatataa ggatctgggc gaatattacg ccgaactgaa tccgctgctc taccatattt
     3481 ccttccaacg aatcgctgaa aaagaaataa tggacgccgt tgaaaccggc aaactgtatc
     3541 tgtttcaaat ctacaacaaa gatttcgcca aaggccatca cggtaagccg aacctgcata
     3601 ccctgtattg gaccggtctg tttagcccgg agaatctggc caaaaccagc atcaagctga
     3661 acggacaggc agaactgttt taccgcccca aaagccgtat gaaaaggatg gcacaccgcc
     3721 tgggcgaaaa aatgctgaat aagaaactca aagatcagaa aacgccgata ccggataccc
     3781 tttatcagga gctgtatgat tatgttaacc accggctgag ccatgacctg agcgacgaag
     3841 cgcgtgcact gctgccgaac gtgattacca aggaagtctc gcatgaaatt attaaagatc
     3901 ggcgcttcac cagtgataaa tttttcttcc atgtaccgat caccctgaat tatcaagccg
     3961 caaatagccc ttccaaattt aatcaacgcg tgaatgcgta cctgaaagag catccggaga
     4021 ccccaattat tggcatagac cgaggagaac gcaatctcat ttatatcacc gtcattgata
     4081 gcaccggtaa gatcctggaa cagcgtagcc tgaataccat tcagcagttt gactaccaga
     4141 aaaagctgga caacagagaa aaggaacgtg tagccgcccg gcaggcttgg agtgtggtgg
     4201 gtactatcaa ggatctgaag caggggtatc tctcccaagt tatccatgaa attgtcgatc
     4261 taatgattca ctatcaagca gtagtggtac tggaaaatct gaatttcggt ttcaaaagca
     4321 aacgtacagg gatcgctgaa aaagccgttt atcagcagtt cgagaaaatg ctgatagaca
     4381 agctgaattg cctggttctg aaagattatc cggcagagaa ggtgggcggt gtgctgaacc
     4441 cgtaccagct gactgatcaa tttacgagct ttgcaaaaat gggaacgcag agcggtttcc
     4501 tgttctatgt tccggcgcca tataccagca agatagaccc gctgacaggt ttcgtagatc
     4561 cgtttgtctg gaaaaccatt aaaaatcatg aaagtcgcaa acattttctg gagggctttg
     4621 attttctgca ctatgacgtg aaaaccggcg acttcattct gcattttaaa atgaaccgta
     4681 atctgtcctt tcagcgcggc ctgcctggct ttatgccggc gtgggacatt gtttttgaaa
     4741 agaatgagac acagtttgat gccaaaggta ccccctttat tgcggggaaa cgcattgtgc
     4801 ccgttataga aaatcaccgc ttcaccggac ggtataggga cttgtacccg gcaaatgaat
     4861 tgatagcgct gctggaggag aaaggtattg tctttcggga tggatcaaac atcctgccga
     4921 agctgctgga gaacgatgac agccacgcaa tagacaccat ggtagcgctg atccgaagcg
     4981 tgctgcagat gcgtaacagt aatgcggcta cgggggaaga ctacattaat agcccggtcc
     5041 gtgatctgaa cggcgtttgt ttcgatagca gatttcaaaa tccggagtgg ccgatggatg
     5101 ccgatgccaa tggagcttac catatcgctc tcaaaggtca gctcctactg aaccatttga
     5161 aagaatcaaa agatctgaaa ctgcagaacg gcatctcgaa tcaggactgg ctggcctaca
     5221 ttcaagaact gagaaaccca aagaaaaaga gaaaagtagg tggttacccc tacgacgtgc
     5281 ccgactacgc ctaacaccca gctttcttgt acaaagtggt gatagcttgg cactggccgt
     5341 cgttttacaa cgtcgtgact gggaaaaccc tggcgttacc caacttaatc gccttgcagc
     5401 acatccccct ttcgccagct ggcgtaatag cgaagaggcc cgcaccgatc gcccttccca
     5461 acagttgcgc agcctgaatg gcgaatggca gcttggctgt tttggcggat gagataagat
     5521 tttcagcctg atacagatta aatcagaacg cagaagcggt ctgataaaac agaatttgcc
     5581 tggcggcagt agcgcggtgg tcccacctga ccccatgccg aactcagaag tgaaacgccg
     5641 tagcgccgat ggtagtgtgg ggtctcccca tgcgagagta gggaactgcc aggcatcaaa
     5701 taaaacgaaa ggctcagtcg aaagactggg cctttcgttt tatctgttgt ttgtcggtga
     5761 acgctctcct gagtaggaca aatccgccgg gagcggattt gaacgttgcg aagcaacggc
     5821 ccggagggtg gcgggcagga cgcccgccat aaactgccag gcatcaaatt aagcagaagg
     5881 ccatcctgac ggatggcctt tttgcgtttc tacaaactct tttgtttatt tttctaaata
     5941 cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga
     6001 aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca
     6061 ttttgccttc ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat
     6121 cagttgggtg cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag
     6181 agttttcgcc ccgaagaacg tttcccaatg atgagcactt ttaaagttct gctatgtggc
     6241 gcggtattat cccgtgttga cgccgggcaa gagcaactcg gtcgccgcat acactattct
     6301 cagaatgact tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca
     6361 gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt
     6421 ctgacaacga tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat
     6481 gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt
     6541 gacaccacga tgcctgtagc aatggcaaca acgttgcgca aactattaac tggcgaacta
     6601 cttactctag cttcccggca acaattaata gactggatgg aggcggataa agttgcagga
     6661 ccacttctgc gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt
     6721 gagcgtgggt ctcgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc
     6781 gtagttatct acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct
     6841 gagataggtg cctcactgat taagcattgg taactgtcag accaagttta ctcatatata
     6901 ctttagattg atttaccccg gttgataatc agaaaagccc caaaaacagg aagattgtat
     6961 aagcaaatat ttaaattgta aacgttaata ttttgttaaa attcgcgtta aatttttgtt
     7021 aaatcagctc attttttaac caataggccg aaatcggcaa aatcccttat aaatcaaaag
     7081 aatagcccga gatagggttg agtgttgttc cagtttggaa caagagtcca ctattaaaga
     7141 acgtggactc caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg
     7201 aaccatcacc caaatcaagt tttttggggt cgaggtgccg taaagcacta aatcggaacc
     7261 ctaaagggag cccccgattt agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg
     7321 aagggaagaa agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc
     7381 gcgtaaccac cacacccgcc gcgcttaatg cgccgctaca gggcgcgtaa aaggatctag
     7441 gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac
     7501 tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc
     7561 gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat
     7621 caagagctac caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat
     7681 actgtccttc tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct
     7741 acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt
     7801 cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg
     7861 gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta
     7921 cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg
     7981 gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg
     8041 tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc
     8101 tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg
     8161 gccttttgct ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat
     8221 aaccgtatta ccgcctttga gtgagctgat accgctcgcc gcagccgaac gaccgagcgc
     8281 agcgagtcag tgagcgagga agcggaagag cgcctgatgc ggtattttct ccttacgcat
     8341 ctgtgcggta tttcacaccg catatggtgc actctcagta caatctgctc tgatgccgca
     8401 tagttaagcc agtatacact ccgctatcgc tacgtgactg ggtcatggct gcgccccgac
     8461 acccgccaac acccgctgac gcgccctgac gggcttgtct gctcccggca tccgcttaca
     8521 gacaagctgt gaccgtctcc gggagctgca tgtgtcagag gttttcaccg tcatcaccga
     8581 aacgcgcgag gcagctgcgg taaagctcat cagcgtggtc gtgcagcgat tcacagatgt
     8641 ctgcctgttc atccgcgtcc agctcgttga gtttctccag aagcgttaat gtctggcttc
     8701 tgataaagcg ggccatgtta agggcggttt tttcctgttt ggtcactgat gcctccgtgt
     8761 aagggggatt tctgttcatg ggggtaatga taccgatgaa acgagagagg atgctcacga
     8821 tacgggttac tgatgatgaa catgcccggt tactggaacg ttgtgagggt aaacaactgg
     8881 cggtatggat gcggcgggac cagagaaaaa tcactcaggg tcaatgccag cgcttcgtta
     8941 atacagatgt aggtgttcca cagggtagcc agcagcatcc tgcgatgcag atccggaaca
     9001 taatggtgca gggcgctgac ttccgcgttt ccagacttta cgaaacacgg aaaccgaaga
     9061 ccattcatgt tgttgctcag gtcgcagacg ttttgcagca gcagtcgctt cacgttcgct
     9121 cgcgtatcgg tgattcattc tgctaaccag taaggcaacc ccgccagcct agccgggtcc
     9181 tcaacgacag gagcacgatc atgcgcaccc gtggccagga cccaacgctg cccgaaattc
     9241 cgacaccatc gaatggtgca aaacctttcg cggtatggca tgatagcgcc cggaagagag
     9301 tcaattcagg gtggtgaatg tgaaaccagt aacgttatac gatgtcgcag agtatgccgg
     9361 tgtctcttat cagaccgttt cccgcgtggt gaaccaggcc agccacgttt ctgcgaaaac
     9421 gcgggaaaaa gtggaagcgg cgatggcgga gctgaattac attcccaacc gcgtggcaca
     9481 acaactggcg ggcaaacagt cgttgctgat tggcgttgcc acctccagtc tggccctgca
     9541 cgcgccgtcg caaattgtcg cggcgattaa atctcgcgcc gatcaactgg gtgccagcgt
     9601 ggtggtgtcg atggtagaac gaagcggcgt cgaagcctgt aaagcggcgg tgcacaatct
     9661 tctcgcgcaa cgcgtcagtg ggctgatcat taactatccg ctggatgacc aggatgccat
     9721 tgctgtggaa gctgcctgca ctaatgttcc ggcgttattt cttgatgtct ctgaccagac
     9781 acccatcaac agtattattt tctcccatga agacggtacg cgactgggcg tggagcatct
     9841 ggtcgcattg ggtcaccagc aaatcgcgct gttagcgggc ccattaagtt ctgtctcggc
     9901 gcgtctgcgt ctggctggct ggcataaata tctcactcgc aatcaaattc agccgatagc
     9961 ggaacgggaa ggcgactgga gtgccatgtc cggttttcaa caaaccatgc aaatgctgaa
    10021 tgagggcatc gttcccactg cgatgctggt tgccaacgat cagatggcgc tgggcgcaat
    10081 gcgcgccatt accgagtccg ggctgcgcgt tggtgcggat atctcggtag tgggatacga
    10141 cgataccgaa gacagctcat gttatatccc gccgttaacc accatcaaac aggattttcg
    10201 cctgctgggg caaaccagcg tggaccgctt gctgcaactc tctcagggcc aggcggtgaa
    10261 gggcaatcag ctgttgcccg tctcactggt gaaaagaaaa accaccctgg cgcccaatac
    10321 gcaaaccgcc tctccccgcg cgttggccga ttcattaatg cagctggcac gacaggtttc
    10381 ccgactggaa agcgggcagt gagcgcaacg caattaatgt aagttagctc actcattagg
    10441 cacaattctc atgtttgaca gcttatcatc gactgcacgg tgcaccaatg cttctggcgt
    10501 caggcagcca tcggaagctg tggtatggct gtgcaggtcg taaatcactg cataattcgt
    10561 gtcgctcaag gcgcactccc gttctggata atgttttttg cgccgacatc ataacggttc
    10621 tggcaaatat tctgaaatga gctg
//