pRetroQ-AcGFP1-C1

pRetroQ-AcGFP1-C1

载体基本信息

出品公司:	Ybscience
载体名称:	pRetroQ-AcGFP1-C1
质粒类型:	逆病毒载体；荧光报告载体
高拷贝/低拷贝:	低拷贝
克隆方法:	限制性内切酶，多克隆位点
启动子:	CMV IE
载体大小:	7614 BP
5' 测序引物及序列:	--
3' 测序引物及序列:	--
载体标签:	AcGFP1(N-端）
载体抗性:	氨苄青霉素
筛选标记:	嘌呤霉素（Puromycin）
克隆菌株:	DH5α
宿主细胞（系）:	常规细胞系（293、CV-1、CHO等）；原代细胞
备注:	pRetroQ-AcGFP1-C1载体是自我失活型逆病毒载体，表达N端AcGFP1融合蛋白； pRetroQ-AcGFP1-C1载体能够高效递送目的基因到靶细胞中，尤其适用于原代细胞和其它难转染的细胞系。
稳定性:	稳表达
组成型/诱导型:	组成型
病毒/非病毒:	逆转录病毒

 

载体质粒图谱和多克隆位点信息

载体简介

 

pRetroQ-AcGFP1-C1载体描述

pRetroQ-AcGFP1-C1 is a high-titer, self-inactivating retroviral vector that facilitates efficient delivery and expression of AcGFP1, as well as C-terminal fusions of AcGFP1, to target cells. AcGFP1 is the green fluorescent protein from Aequorea coerulescens (AcGFP1; Excitation maximum = 475 nm; emission maximum = 505 nm).
The AcGFP1 coding sequence contains silent base changes, which correspond to human codon-usage preferences (1). The multiple cloning site (MCS) in pRetroQAcGFP1-C1 follows the AcGFP1 coding sequence. Genes cloned into the MCS are expressed as fusions to the C-terminus of AcGFP1 when they are in the same reading frame as AcGFP1 and there are no intervening stop codons.

The RetroQ retroviral vector backbone incorporates several unique features. This vector contains a puromycin resistance cassette (Puror) driven by the PGK promoter (PPGK) for selection of positively-infected cells (2).The hybrid 5’ long terminal repeats (LTR) consists of the CMV type I enhancer and the murine sarcoma virus (MSV) promoter. This vector demonstrates high levels of transcription in HEK 293-based packaging cell lines due, in part, to the presence of adenoviral E1A (3–6) in these cells. The self-inactivating feature of the vector is provided by a deletion in the 3’ LTR enhancer region (U3). During reverse transcription of the retroviral RNA, the inactivated 3’ LTR is copied and replaces the 5’ LTR CMV enhancer sequences. This can reduce the phenomenon known as promoter interference (7) and allow more efficient expression.

Additionally, the viral genomic transcript contains the necessary viral RNA processing elements, including the LTRs, packaging signal (ψ+), and tRNA primer-binding site. pRetroQ-AcGFP1-C1 contains a bacterial origin of replication, an E. coli Ampr gene for propagation and selection in bacteria, and an SV40 origin for replication in mammalian cells expressing the SV40 large T antigen.

pRetroQ-AcGFP1-C1 is designed to efficiently deliver and express fusions to the C terminus of AcGFP1 into primary cells or cells that are difficult to transfect. Fusions to the C terminus of AcGFP1 retain the fluorescent properties of the native protein, allowing the in vivo localization of the fusion protein. The target gene should be cloned into pRetroQ- AcGFP1-C1 so that it is in frame with the AcGFP1 coding sequences, with no intervening in-frame stop codons. The inserted gene should include the initiating ATG codon. The recombinant AcGFP1 vector can be infected or transfected into mammalian cells. If required, stable transformants can be selected using Puromycin. pRetroQ-AcGFP1-C1 can also be used simply to express AcGFP1 in a cell line of interest (e.g., as an infection marker).

Once pRetroQ-AcGFP1-C1 is transfected into a packaging cell line (such as the RetroPack PT67 Cell line, AmphoPack-293 , EcoPack2-293, Pantropic Expression System, or Retro-X Universal Packaging System, RNA from the vector is packaged into non-infectious, replication-incompetent retroviral particles, since pRetroQ-AcGFP1-C1 lacks the structural genes (gag, pol, and env) necessary for particle formation and replication. These genes, however, are stably integrated as part of the packaging cell genome. Once a high-titer supernatant is produced, these retroviral particles can infect target cells and transmit the gene of interest but cannot replicate within these cells due to the absence of viral structural genes. The separate introduction and integration of the structural genes into the packaging cell line minimizes the chances of producing replication-competent virus due to recombination events during cell proliferation.

Propagation in E. coli
 Suitable host strains: DH5α, Fusion Blue, and other general purpose strains.
 Selectable marker: plasmid confers resistance to ampicillin (100 μg/ml) in E. coli hosts.
 E. coli replication origin: ColE1
 Copy number: low

 

载体序列

LOCUS       pRetroQ-AcGFP1-C1	7614 bp 	DNA	SYN
DEFINITION  pRetroQ-AcGFP1-C1
ACCESSION   
KEYWORDS    
SOURCE      
  ORGANISM  other sequences; artificial sequences; vectors.
FEATURES             Location/Qualifiers
     source          1..7614
                     /organism="pRetroQ-AcGFP1-C1"
                     /mol_type="other DNA"
     promoter        2..506
                     /label="CMV_immearly_promoter"
     misc_feature    6..688
                     /label="5_LTR"
     misc_feature    9..296
                     /label="CAG_enhancer"
     misc_feature    463..483
                     /label="CMV_fwd_primer"
     misc_feature    758..1567
                     /label="psi_plus_pack"
     misc_feature    1154..1561
                     /label="gag"
     misc_feature    1497..1519
                     /label="MSCV_primer"
     misc_feature    1497..1519
                     /label="pLXSN_5_primer"
     misc_feature    1535..1551
                     /label="pBABE_5_primer"
     promoter        1591..2143
                     /label="CMV_immearly_promoter"
     misc_feature    1646..1933
                     /label="CAG_enhancer"
     misc_feature    2100..2120
                     /label="CMV_fwd_primer"
     promoter        2101..2170
                     /label="CMV_promoter"
     CDS             2194..2991
                     /label="ORF frame 1"
                     /translation="MVSKGAELFTGIVPILIELNGDVNGHKFSVSGEGEGDATYGKLT
                     LKFICTTGKLPVPWPTLVTTLSYGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFE
                     DDGNYKSRAEVKFEGDTLVNRIELTGTDFKEDGNILGNKMEYNYNAHNVYIMTDKAKN
                     GIKVNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDH
                     MIYFGFVTAAAITHGMDELYKSGLRSRAQASNSAVDGTAGPGSTGSR*"
     gene            2197..2910
                     /label="AcGFP1"
                     /gene="AcGFP1"
                     /translation="MVSKGAELFTGIVPILIELNGDVNGHKFSVSGEGEGDATYGKLT
                     LKFICTTGKLPVPWPTLVTTLSYGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFE
                     DDGNYKSRAEVKFEGDTLVNRIELTGTDFKEDGNILGNKMEYNYNAHNVYIMTDKAKN
                     GIKVNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDH
                     MIYFGFVTAAAITHGMDELYKSGLRSRAQASNSAVDGTAGPGSTGSR*"
     misc_feature    complement(3044..3067)
                     /label="MSCV_rev_primer"
                     /translation="MVSKGAELFTGIVPILIELNGDVNGHKFSVSGEGEGDATYGKLT
                     LKFICTTGKLPVPWPTLVTTLSYGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFE
                     DDGNYKSRAEVKFEGDTLVNRIELTGTDFKEDGNILGNKMEYNYNAHNVYIMTDKAKN
                     GIKVNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDH
                     MIYFGFVTAAAITHGMDELYKSGLRSRAQASNSAVDGTAGPGSTGSR*"
     CDS             3289..4128
                     /label="ORF frame 1"
                     /translation="MEAGRPLGQRPIAALLLRFLGSEAGKGWVRGRAQGRAQGRGGRP
                     KVLRRPGILHASKAHVCRAVLLFLISGPFDLQPKLTMTEYKPTVRLATRDDVPRAVRT
                     LAAAFADYPATRHTVDPDRHIERVTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPE
                     SVEAGAVFAEIGPRMAELSGSRLAAQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKG
                     LGSAVVLPGVEAAERAGVPAFLETSAPRNLPFYERLGFTVTADVEVPEGPRTWCMTRK
                     PGA*"
     misc_feature    3441..3460
                     /label="mPGK_F_primer"
                     /translation="MEAGRPLGQRPIAALLLRFLGSEAGKGWVRGRAQGRAQGRGGRP
                     KVLRRPGILHASKAHVCRAVLLFLISGPFDLQPKLTMTEYKPTVRLATRDDVPRAVRT
                     LAAAFADYPATRHTVDPDRHIERVTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPE
                     SVEAGAVFAEIGPRMAELSGSRLAAQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKG
                     LGSAVVLPGVEAAERAGVPAFLETSAPRNLPFYERLGFTVTADVEVPEGPRTWCMTRK
                     PGA*"
     CDS             complement(3454..4179)
                     /label="ORF frame 1"
                     /translation="MGSCAPFGRALRVVGRASGTGLAGHAPGARSFGHLDVGGDGEAE
                     PLVEGEVAGRGGLQEGGHPGALGRLHSGEHDGAAQTLALVVGRDADGGQEPRGLLGPV
                     RRQEAFHLLLRGQPGTAQLGHARADLGEHRPRFDALRRGPDRHRGAVVRDPHLADVEP
                     DAREEEFLQLGDPLDVAVRIDGVARGGVVGERGGEGAYGPGDVVAGGEAHRGLVLGHG
                     KLGLQVERPGDEEEENSAADVRF*"
     gene            3529..4128
                     /label="puro (variant)"
                     /gene="puro (variant)"
                     /translation="MGSCAPFGRALRVVGRASGTGLAGHAPGARSFGHLDVGGDGEAE
                     PLVEGEVAGRGGLQEGGHPGALGRLHSGEHDGAAQTLALVVGRDADGGQEPRGLLGPV
                     RRQEAFHLLLRGQPGTAQLGHARADLGEHRPRFDALRRGPDRHRGAVVRDPHLADVEP
                     DAREEEFLQLGDPLDVAVRIDGVARGGVVGERGGEGAYGPGDVVAGGEAHRGLVLGHG
                     KLGLQVERPGDEEEENSAADVRF*"
     misc_feature    complement(4848..4870)
                     /label="pGEX_3_primer"
                     /translation="MGSCAPFGRALRVVGRASGTGLAGHAPGARSFGHLDVGGDGEAE
                     PLVEGEVAGRGGLQEGGHPGALGRLHSGEHDGAAQTLALVVGRDADGGQEPRGLLGPV
                     RRQEAFHLLLRGQPGTAQLGHARADLGEHRPRFDALRRGPDRHRGAVVRDPHLADVEP
                     DAREEEFLQLGDPLDVAVRIDGVARGGVVGERGGEGAYGPGDVVAGGEAHRGLVLGHG
                     KLGLQVERPGDEEEENSAADVRF*"
     misc_feature    complement(5006..5026)
                     /label="pBABE_3_primer"
                     /translation="MGSCAPFGRALRVVGRASGTGLAGHAPGARSFGHLDVGGDGEAE
                     PLVEGEVAGRGGLQEGGHPGALGRLHSGEHDGAAQTLALVVGRDADGGQEPRGLLGPV
                     RRQEAFHLLLRGQPGTAQLGHARADLGEHRPRFDALRRGPDRHRGAVVRDPHLADVEP
                     DAREEEFLQLGDPLDVAVRIDGVARGGVVGERGGEGAYGPGDVVAGGEAHRGLVLGHG
                     KLGLQVERPGDEEEENSAADVRF*"
     misc_feature    complement(5012..5227)
                     /label="SV40_enhancer"
                     /translation="MGSCAPFGRALRVVGRASGTGLAGHAPGARSFGHLDVGGDGEAE
                     PLVEGEVAGRGGLQEGGHPGALGRLHSGEHDGAAQTLALVVGRDADGGQEPRGLLGPV
                     RRQEAFHLLLRGQPGTAQLGHARADLGEHRPRFDALRRGPDRHRGAVVRDPHLADVEP
                     DAREEEFLQLGDPLDVAVRIDGVARGGVVGERGGEGAYGPGDVVAGGEAHRGLVLGHG
                     KLGLQVERPGDEEEENSAADVRF*"
     promoter        5024..5292
                     /label="SV40_promoter"
                     /translation="MGSCAPFGRALRVVGRASGTGLAGHAPGARSFGHLDVGGDGEAE
                     PLVEGEVAGRGGLQEGGHPGALGRLHSGEHDGAAQTLALVVGRDADGGQEPRGLLGPV
                     RRQEAFHLLLRGQPGTAQLGHARADLGEHRPRFDALRRGPDRHRGAVVRDPHLADVEP
                     DAREEEFLQLGDPLDVAVRIDGVARGGVVGERGGEGAYGPGDVVAGGEAHRGLVLGHG
                     KLGLQVERPGDEEEENSAADVRF*"
     rep_origin      5191..5268
                     /label="SV40_origin"
                     /translation="MGSCAPFGRALRVVGRASGTGLAGHAPGARSFGHLDVGGDGEAE
                     PLVEGEVAGRGGLQEGGHPGALGRLHSGEHDGAAQTLALVVGRDADGGQEPRGLLGPV
                     RRQEAFHLLLRGQPGTAQLGHARADLGEHRPRFDALRRGPDRHRGAVVRDPHLADVEP
                     DAREEEFLQLGDPLDVAVRIDGVARGGVVGERGGEGAYGPGDVVAGGEAHRGLVLGHG
                     KLGLQVERPGDEEEENSAADVRF*"
     misc_feature    5253..5272
                     /label="SV40pro_F_primer"
                     /translation="MGSCAPFGRALRVVGRASGTGLAGHAPGARSFGHLDVGGDGEAE
                     PLVEGEVAGRGGLQEGGHPGALGRLHSGEHDGAAQTLALVVGRDADGGQEPRGLLGPV
                     RRQEAFHLLLRGQPGTAQLGHARADLGEHRPRFDALRRGPDRHRGAVVRDPHLADVEP
                     DAREEEFLQLGDPLDVAVRIDGVARGGVVGERGGEGAYGPGDVVAGGEAHRGLVLGHG
                     KLGLQVERPGDEEEENSAADVRF*"
     gene            complement(6392..7252)
                     /label="Ampicillin"
                     /gene="Ampicillin"
                     /translation="MGSCAPFGRALRVVGRASGTGLAGHAPGARSFGHLDVGGDGEAE
                     PLVEGEVAGRGGLQEGGHPGALGRLHSGEHDGAAQTLALVVGRDADGGQEPRGLLGPV
                     RRQEAFHLLLRGQPGTAQLGHARADLGEHRPRFDALRRGPDRHRGAVVRDPHLADVEP
                     DAREEEFLQLGDPLDVAVRIDGVARGGVVGERGGEGAYGPGDVVAGGEAHRGLVLGHG
                     KLGLQVERPGDEEEENSAADVRF*"
     CDS             complement(6392..7252)
                     /label="ORF frame 3"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGY
                     IELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVE
                     YSPVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRL
                     DRWEPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPL
                     LRSALPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIA
                     EIGASLIKHW*"
     promoter        complement(7294..7322)
                     /label="AmpR_promoter"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGY
                     IELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVE
                     YSPVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRL
                     DRWEPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPL
                     LRSALPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIA
                     EIGASLIKHW*"
     promoter        7544..506
                     /label="CMV_immearly_promoter"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGY
                     IELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVE
                     YSPVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRL
                     DRWEPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPL
                     LRSALPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIA
                     EIGASLIKHW*"
     misc_feature    7608..688
                     /label="5_LTR"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGY
                     IELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVE
                     YSPVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRL
                     DRWEPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPL
                     LRSALPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIA
                     EIGASLIKHW*"
ORIGIN
    1 TCCGCGTTAC ATAACTTACG GTAAATGGCC CGCCTGGCTG ACCGCCCAAC GACCCCCGCC
   61 CATTGACGTC AATAATGACG TATGTTCCCA TAGTAACGCC AATAGGGACT TTCCATTGAC
  121 GTCAATGGGT GGAGTATTTA CGGTAAACTG CCCACTTGGC AGTACATCAA GTGTATCATA
  181 TGCCAAGTAC GCCCCCTATT GACGTCAATG ACGGTAAATG GCCCGCCTGG CATTATGCCC
  241 AGTACATGAC CTTATGGGAC TTTCCTACTT GGCAGTACAT CTACGTATTA GTCATCGCTA
  301 TTACCATGGT GATGCGGTTT TGGCAGTACA TCAATGGGCG TGGATAGCGG TTTGACTCAC
  361 GGGGATTTCC AAGTCTCCAC CCCATTGACG TCAATGGGAG TTTGTTTTGG CACCAAAATC
  421 AACGGGACTT TCCAAAATGT CGTAACAACT CCGCCCCATT GACGCAAATG GGCGGTAGGC
  481 GTGTACGGTG GGAGGTCTAT ATAAGCAGAG CTCAATAAAA GAGCCCACAA CCCCTCACTC
  541 GGCGCGCCAG TCTTCCGATA GACTGCGTCG CCCGGGTACC CGTATTCCCA ATAAAGCCTC
  601 TTGCTGTTTG CATCCGAATC GTGGTCTCGC TGTTCCTTGG GAGGGTCTCC TCTGAGTGAT
  661 TGACTACCCA CGACGGGGGT CTTTCATTTG GGGGCTCGTC CGGGATTTGG AGACCCCTGC
  721 CCAGGGACCA CCGACCCACC ACCGGGAGGT AAGCTGGCCA GCAACTTATC TGTGTCTGTC
  781 CGATTGTCTA GTGTCTATGT TTGATGTTAT GCGCCTGCGT CTGTACTAGT TAGCTAACTA
  841 GCTCTGTATC TGGCGGACCC GTGGTGGAAC TGACGAGTTC TGAACACCCG GCCGCAACCC
  901 TGGGAGACGT CCCAGGGACT TTGGGGGCCG TTTTTGTGGC CCGACCTGAG GAAGGGAGTC
  961 GATGTGGAAT CCGACCCCGT CAGGATATGT GGTTCTGGTA GGAGACGAGA ACCTAAAACA
 1021 GTTCCCGCCT CCGTCTGAAT TTTTGCTTTC GGTTTGGAAC CGAAGCCGCG CGTCTTGTCT
 1081 GCTGCAGCGC TGCAGCATCG TTCTGTGTTG TCTCTGTCTG ACTGTGTTTC TGTATTTGTC
 1141 TGAAAATTAG GGCCAGACTG TTACCACTCC CTTAAGTTTG ACCTTAGGTC ACTGGAAAGA
 1201 TGTCGAGCGG ATCGCTCACA ACCAGTCGGT AGATGTCAAG AAGAGACGTT GGGTTACCTT
 1261 CTGCTCTGCA GAATGGCCAA CCTTTAACGT CGGATGGCCG CGAGACGGCA CCTTTAACCG
 1321 AGACCTCATC ACCCAGGTTA AGATCAAGGT CTTTTCACCT GGCCCGCATG GACACCCAGA
 1381 CCAGGTCCCC TACATCGTGA CCTGGGAAGC CTTGGCTTTT GACCCCCCTC CCTGGGTCAA
 1441 GCCCTTTGTA CACCCTAAGC CTCCGCCTCC TCTTCCTCCA TCCGCCCCGT CTCTCCCCCT
 1501 TGAACCTCCT CGTTCGACCC CGCCTCGATC CTCCCTTTAT CCAGCCCTCA CTCCTTCTCT
 1561 AGGCGCCGGA ATTGAAGATC ATAGTTATTA ATAGTAATCA ATTACGGGGT CATTAGTTCA
 1621 TAGCCCATAT ATGGAGTTCC GCGTTACATA ACTTACGGTA AATGGCCCGC CTGGCTGACC
 1681 GCCCAACGAC CCCCGCCCAT TGACGTCAAT AATGACGTAT GTTCCCATAG TAACGCCAAT
 1741 AGGGACTTTC CATTGACGTC AATGGGTGGA GTATTTACGG TAAACTGCCC ACTTGGCAGT
 1801 ACATCAAGTG TATCATATGC CAAGTACGCC CCCTATTGAC GTCAATGACG GTAAATGGCC
 1861 CGCCTGGCAT TATGCCCAGT ACATGACCTT ATGGGACTTT CCTACTTGGC AGTACATCTA
 1921 CGTATTAGTC ATCGCTATTA CCATGGTGAT GCGGTTTTGG CAGTACATCA ATGGGCGTGG
 1981 ATAGCGGTTT GACTCACGGG GATTTCCAAG TCTCCACCCC ATTGACGTCA ATGGGAGTTT
 2041 GTTTTGGCAC CAAAATCAAC GGGACTTTCC AAAATGTCGT AACAACTCCG CCCCATTGAC
 2101 GCAAATGGGC GGTAGGCGTG TACGGTGGGA GGTCTATATA AGCAGAGCTG GTTTAGTGAA
 2161 CCGTCAGATC CGCTAGCGCT ACCGGTCGCC ACCATGGTGA GCAAGGGCGC CGAGCTGTTC
 2221 ACCGGCATCG TGCCCATCCT GATCGAGCTG AATGGCGATG TGAATGGCCA CAAGTTCAGC
 2281 GTGAGCGGCG AGGGCGAGGG CGATGCCACC TACGGCAAGC TGACCCTGAA GTTCATCTGC
 2341 ACCACCGGCA AGCTGCCTGT GCCCTGGCCC ACCCTGGTGA CCACCCTGAG CTACGGCGTG
 2401 CAGTGCTTCT CACGCTACCC CGATCACATG AAGCAGCACG ACTTCTTCAA GAGCGCCATG
 2461 CCTGAGGGCT ACATCCAGGA GCGCACCATC TTCTTCGAGG ATGACGGCAA CTACAAGTCG
 2521 CGCGCCGAGG TGAAGTTCGA GGGCGATACC CTGGTGAATC GCATCGAGCT GACCGGCACC
 2581 GATTTCAAGG AGGATGGCAA CATCCTGGGC AATAAGATGG AGTACAACTA CAACGCCCAC
 2641 AATGTGTACA TCATGACCGA CAAGGCCAAG AATGGCATCA AGGTGAACTT CAAGATCCGC
 2701 CACAACATCG AGGATGGCAG CGTGCAGCTG GCCGACCACT ACCAGCAGAA TACCCCCATC
 2761 GGCGATGGCC CTGTGCTGCT GCCCGATAAC CACTACCTGT CCACCCAGAG CGCCCTGTCC
 2821 AAGGACCCCA ACGAGAAGCG CGATCACATG ATCTACTTCG GCTTCGTGAC CGCCGCCGCC
 2881 ATCACCCACG GCATGGATGA GCTGTACAAG TCCGGACTCA GATCTCGAGC TCAAGCTTCG
 2941 AATTCTGCAG TCGACGGTAC CGCGGGCCCG GGATCCACCG GATCTAGATA ACTGATCATA
 3001 ATTCTACCGG GTAGGGGAGG CGCTTTTCCC AAGGCAGTCT GGAGCATGCG CTTTAGCAGC
 3061 CCCGCTGGGC ACTTGGCGCT ACACAAGTGG CCTCTGGCCT CGCACACATT CCACATCCAC
 3121 CGGTAGGCGC CAACCGGCTC CGTTCTTTGG TGGCCCCTTC GCGCCACCTT CTACTCCTCC
 3181 CCTAGTCAGG AAGTTCCCCC CCGCCCCGCA GCTCGCGTCG TGCAGGACGT GACAAATGGA
 3241 AGTAGCACGT CTCACTAGTC TCGTGCAGAT GGACAGCACC GCTGAGCAAT GGAAGCGGGT
 3301 AGGCCTTTGG GGCAGCGGCC AATAGCAGCT TTGCTCCTTC GCTTTCTGGG CTCAGAGGCT
 3361 GGGAAGGGGT GGGTCCGGGG GCGGGCTCAG GGGCGGGCTC AGGGGCGGGG CGGGCGCCCG
 3421 AAGGTCCTCC GGAGGCCCGG CATTCTGCAC GCTTCAAAAG CGCACGTCTG CCGCGCTGTT
 3481 CTCCTCTTCC TCATCTCCGG GCCTTTCGAC CTGCAGCCCA AGCTTACCAT GACCGAGTAC
 3541 AAGCCCACGG TGCGCCTCGC CACCCGCGAC GACGTCCCCA GGGCCGTACG CACCCTCGCC
 3601 GCCGCGTTCG CCGACTACCC CGCCACGCGC CACACCGTCG ATCCGGACCG CCACATCGAG
 3661 CGGGTCACCG AGCTGCAAGA ACTCTTCCTC ACGCGCGTCG GGCTCGACAT CGGCAAGGTG
 3721 TGGGTCGCGG ACGACGGCGC CGCGGTGGCG GTCTGGACCA CGCCGGAGAG CGTCGAAGCG
 3781 GGGGCGGTGT TCGCCGAGAT CGGCCCGCGC ATGGCCGAGT TGAGCGGTTC CCGGCTGGCC
 3841 GCGCAGCAAC AGATGGAAGG CCTCCTGGCG CCGCACCGGC CCAAGGAGCC CGCGTGGTTC
 3901 CTGGCCACCG TCGGCGTCTC GCCCGACCAC CAGGGCAAGG GTCTGGGCAG CGCCGTCGTG
 3961 CTCCCCGGAG TGGAGGCGGC CGAGCGCGCC GGGGTGCCCG CCTTCCTGGA GACCTCCGCG
 4021 CCCCGCAACC TCCCCTTCTA CGAGCGGCTC GGCTTCACCG TCACCGCCGA CGTCGAGGTG
 4081 CCCGAAGGAC CGCGCACCTG GTGCATGACC CGCAAGCCCG GTGCCTGACG CCCGCCCCAC
 4141 GACCCGCAGC GCCCGACCGA AAGGAGCGCA CGACCCCATG CATCGGCGTC TCGATCGAGA
 4201 TATCAGTGGT CCAGGCTCTA GTTTTGACTC AACAATATCA CCAGCTGAAG CCTATAGAGT
 4261 ACGAGCCATA GATAAAATAA AAGATTTTAT TTAGTCTCCA GAAAAAGGGG GGAATGAAAG
 4321 ACCCCACCTG TAGGTTTGGC AAGCTACTAG CTTAAGTAAC GCCATTTTGC AAGGCATGGA
 4381 AAAATACATA ACTGAGAATA GAGAAGTTCA GATCAAGGTC AGGAACAGAT GGAACAGGGT
 4441 CGATCGACCC TAGAGAACCA TCAGATGTTT CCAGGGTGCC CCAAGGACCT GAAATGACCC
 4501 TGTGCCTTAT TTGAACTAAC CAATCAGTTC GCTTCTCGCT TCTGTTCGCG CGCTTCTGCT
 4561 CCCCGAGCTC AATAAAAGAG CCCACAACCC CTCACTCGGG GCGCCAGTCC TCCGATTGAC
 4621 TGAGTCGCCC GGGTACCCGT GTATCCAATA AACCCTCTTG CAGTTGCATC CGACTTGTGG
 4681 TCTCGCTGTT CCTTGGGAGG GTCTCCTCTG AGTGATTGAC TACCCGTCAG CGGGGGTCTT
 4741 TCATTTGGGG GCTCGTCCGG GATCGGGAGA CCCCTGCCCA GGGACCACCG ACCCACCACC
 4801 GGGAGGTAAG CTGGCTGCCT CGCGCGTTTC GGTGATGACG GTGAAAACCT CTGACACATG
 4861 CAGCTCCCGG AGACGGTCAC AGCTTGTCTG TAAGCGGATG CCGGGAGCAG ACAAGCCCGT
 4921 CAGGGCGCGT CAGCGGGTGT TGGCGGGTGT CGGGGCGCAG CCATGACCCA GTCACGTAGC
 4981 GATAGCGGAG TGTAGATCCG GCTGTGGAAT GTGTGTCAGT TAGGGTGTGG AAAGTCCCCA
 5041 GGCTCCCCAG CAGGCAGAAG TATGCAAAGC ATGCATCTCA ATTAGTCAGC AACCAGGTGT
 5101 GGAAAGTCCC CAGGCTCCCC AGCAGGCAGA AGTATGCAAA GCATGCATCT CAATTAGTCA
 5161 GCAACCATAG TCCCGCCCCT AACTCCGCCC ATCCCGCCCC TAACTCCGCC CAGTTCCGCC
 5221 CATTCTCCGC CCCATGGCTG ACTAATTTTT TTTATTTATG CAGAGGCCGA GGCCGCCTCG
 5281 GCCTCTGAGC TATTCCAGAA GTAGTGAGGA GGCTTTTTTG GAGGCCTAGG CTTTTGCAAA
 5341 AAGCTTACTG GCTTAACTAT GCGGCATCAG AGCAGATTGT ACTGAGAGTG CACCATATGC
 5401 GGTGTGAAAT ACCGCACAGA TGCGTAAGGA GAAAATACCG CATCAGGCGC TCTTCCGCTT
 5461 CCTCGCTCAC TGACTCGCTG CGCTCGGTCG TTCGGCTGCG GCGAGCGGTA TCAGCTCACT
 5521 CAAAGGCGGT AATACGGTTA TCCACAGAAT CAGGGGATAA CGCAGGAAAG AACATGTGAG
 5581 CAAAAGGCCA GCAAAAGGCC AGGAACCGTA AAAAGGCCGC GTTGCTGGCG TTTTTCCATA
 5641 GGCTCCGCCC CCCTGACGAG CATCACAAAA ATCGACGCTC AAGTCAGAGG TGGCGAAACC
 5701 CGACAGGACT ATAAAGATAC CAGGCGTTTC CCCCTGGAAG CTCCCTCGTG CGCTCTCCTG
 5761 TTCCGACCCT GCCGCTTACC GGATACCTGT CCGCCTTTCT CCCTTCGGGA AGCGTGGCGC
 5821 TTTCTCATAG CTCACGCTGT AGGTATCTCA GTTCGGTGTA GGTCGTTCGC TCCAAGCTGG
 5881 GCTGTGTGCA CGAACCCCCC GTTCAGCCCG ACCGCTGCGC CTTATCCGGT AACTATCGTC
 5941 TTGAGTCCAA CCCGGTAAGA CACGACTTAT CGCCACTGGC AGCAGCCACT GGTAACAGGA
 6001 TTAGCAGAGC GAGGTATGTA GGCGGTGCTA CAGAGTTCTT GAAGTGGTGG CCTAACTACG
 6061 GCTACACTAG AAGGACAGTA TTTGGTATCT GCGCTCTGCT GAAGCCAGTT ACCTTCGGAA
 6121 AAAGAGTTGG TAGCTCTTGA TCCGGCAAAC AAACCACCGC TGGTAGCGGT GGTTTTTTTG
 6181 TTTGCAAGCA GCAGATTACG CGCAGAAAAA AAGGATCTCA AGAAGATCCT TTGATCTTTT
 6241 CTACGGGGTC TGACGCTCAG TGGAACGAAA ACTCACGTTA AGGGATTTTG GTCATGAGAT
 6301 TATCAAAAAG GATCTTCACC TAGATCCTTT TAAATTAAAA ATGAAGTTTT AAATCAATCT
 6361 AAAGTATATA TGAGTAAACT TGGTCTGACA GTTACCAATG CTTAATCAGT GAGGCACCTA
 6421 TCTCAGCGAT CTGTCTATTT CGTTCATCCA TAGTTGCCTG ACTCCCCGTC GTGTAGATAA
 6481 CTACGATACG GGAGGGCTTA CCATCTGGCC CCAGTGCTGC AATGATACCG CGAGACCCAC
 6541 GCTCACCGGC TCCAGATTTA TCAGCAATAA ACCAGCCAGC CGGAAGGGCC GAGCGCAGAA
 6601 GTGGTCCTGC AACTTTATCC GCCTCCATCC AGTCTATTAA TTGTTGCCGG GAAGCTAGAG
 6661 TAAGTAGTTC GCCAGTTAAT AGTTTGCGCA ACGTTGTTGC CATTGCTGCA GGCATCGTGG
 6721 TGTCACGCTC GTCGTTTGGT ATGGCTTCAT TCAGCTCCGG TTCCCAACGA TCAAGGCGAG
 6781 TTACATGATC CCCCATGTTG TGCAAAAAAG CGGTTAGCTC CTTCGGTCCT CCGATCGTTG
 6841 TCAGAAGTAA GTTGGCCGCA GTGTTATCAC TCATGGTTAT GGCAGCACTG CATAATTCTC
 6901 TTACTGTCAT GCCATCCGTA AGATGCTTTT CTGTGACTGG TGAGTACTCA ACCAAGTCAT
 6961 TCTGAGAATA GTGTATGCGG CGACCGAGTT GCTCTTGCCC GGCGTCAACA CGGGATAATA
 7021 CCGCGCCACA TAGCAGAACT TTAAAAGTGC TCATCATTGG AAAACGTTCT TCGGGGCGAA
 7081 AACTCTCAAG GATCTTACCG CTGTTGAGAT CCAGTTCGAT GTAACCCACT CGTGCACCCA
 7141 ACTGATCTTC AGCATCTTTT ACTTTCACCA GCGTTTCTGG GTGAGCAAAA ACAGGAAGGC
 7201 AAAATGCCGC AAAAAAGGGA ATAAGGGCGA CACGGAAATG TTGAATACTC ATACTCTTCC
 7261 TTTTTCAATA TTATTGAAGC ATTTATCAGG GTTATTGTCT CATGAGCGGA TACATATTTG
 7321 AATGTATTTA GAAAAATAAA CAAATAGGGG TTCCGCGCAC ATTTCCCCGA AAAGTGCCAC
 7381 CTGACGTCTA AGAAACCATT ATTATCATGA CATTAACCTA TAAAAATAGG CGTATCACGA
 7441 GGCCCTTTCG TCTTCAAGAA TTAGCTTGGC CATTGCATAC GTTGTATCCA TATCATAATA
 7501 TGTACATTTA TATTGGCTCA TGTCCAACAT TACCGCCATG TTGACATTGA TTATTGACTA
 7561 GTTATTAATA GTAATCAATT ACGGGGTCAT TAGTTCATAG CCCATATATG GAGT