A mitochondrial-based drug delivery system and its use

Abstract

A mitochondrial-based drug delivery system, comprising: mitochondria, and a drug loaded on the mitochondria, wherein the drug is loaded on the mitochondria by non-free diffusion. The drug delivery system can effectively deliver proteins, nucleic acids or other drugs into cells, can be used to treat single gene genetic diseases, multi-gene genetic diseases or other diseases, and can also be used for healthy individuals to achieve the purpose of delaying aging, enhancing body functions or preventing diseases.

Description

Technical Field

[0001] The present invention relates to a mitochondria-based drug delivery system, a method for preparing the drug delivery system, use of the drug delivery system for delivering drugs, and use of the drug delivery system in preparing drugs and / or preparations for treating and / or preventing diseases, beautifying, losing weight, resisting fatigue, promoting growth and development, enhancing body functions or delaying aging. Background Art

[0002] The absence or defect of specific biomacromolecules in human cells can cause a variety of major diseases, such as cancer, hemophilia, and albinism. A common treatment method is to introduce exogenous biomolecule drugs into target cells to replace, compensate, block or correct defective molecules in patient cells, thereby achieving the purpose of preventing and treating diseases.

[0003] However, biomacromolecule drugs such as proteins and nucleic acids must be loaded on certain carriers to successfully enter target cells due to their strong polarity, large molecular weight, and easy degradation. The natural excellent transmembrane transport ability of viruses makes them a common drug delivery carrier. Although viral vectors have achieved preliminary therapeutic effects in clinical practice, their limitations cannot be ignored, such as potential carcinogenicity, cytotoxicity, strong immunogenicity, limited loading capacity, and difficult quality control. At the same time, the high preparation cost of viral vectors further limits their widespread use. For example, patients with lipoprotein lipase deficiency can spend up to one million euros on the gene therapy drug Glybera. This extremely high medical cost far exceeds the economic affordability of ordinary patients, and the drug was eventually withdrawn from the market.

[0004] Another common biomacromolecule carrier is a non-viral carrier. Currently, such carriers mainly include cationic polymers, liposomes or other nanoparticles. However, non-viral carriers still face many challenges, mainly in terms of low drug loading, low transfection efficiency, and high cytotoxicity. These shortcomings also limit its wide clinical application. Therefore, in the field of drug delivery, it is still necessary to develop non-viral biomacromolecule carriers with high transfection efficiency and low toxicity. Summary of the invention

[0005] Mitochondria are important organelles responsible for energy synthesis, cell differentiation, information transmission and apoptosis of eukaryotic cells. Mitochondria evolved from ancient bacteria 1.5 billion years ago, and still retain some characteristics of bacteria, with "infectivity" similar to intracellular bacteria. For example, mitochondria are separated from cells and co-cultured with other cells, and mitochondria can quickly and efficiently enter these cells (Clark, MA and JW Shay (1982). "Mitochondrial transformation of mammalian cells." Nature 295 (5850): 605-607.). Inspired by this, the inventors of the present invention creatively realized that mitochondria can be used as biomacromolecule carriers to achieve safe and effective intracellular delivery of target drugs.

[0006] So far, there are only studies on loading small molecule anticancer drugs or quantum dots into mitochondria by free diffusion or electroporation to achieve their intracellular delivery (Li, W.-Q., Wang, Z., Hao, S., Sun, L., Nisic, M., Cheng, G., … Zheng, S.-Y. (2018). Mitochondria-based aircraft carrier enhances in vivo imaging of carbon quantum dots and delivery of anticancer drug. Nanoscale, 10 (8), 3744–3752). However, after entering the cell, these drugs need to pass through the two layers of the inner and outer mitochondrial membranes to be released, which undoubtedly reduces their delivery efficiency. At the same time, the drugs that can enter the mitochondria by diffusion are very limited, and the electroporation transfection method is generally believed to be able to destroy the cell membrane and thus lead to a high cell lethality. Therefore, the purpose of the present invention is to provide a new, widely applicable and safe mitochondrial loading method, so as to achieve efficient transmembrane delivery of multiple drugs, especially biomacromolecules such as proteins and nucleic acids.

[0007] In one aspect, the present invention provides a mitochondrial-based drug delivery system, the drug delivery system comprising:

[0008] (1) Mitochondria, and

[0009] (2) a drug loaded into the mitochondria,

[0010] The drug is loaded into the mitochondria by non-free diffusion.

[0011] Wherein, the source of mitochondria can be autologous, allogeneic or xenogeneic and their combinations. The drug can be a protein drug such as a polypeptide drug, an antibody drug, a cytokine, an enzyme, a protein vaccine, a polypeptide hormone, a protein translated by a normal gene, a protein derivative, a nucleic acid drug such as DNA, RNA or a nucleic acid analog, or any other substance that cannot or is difficult to penetrate the cell membrane, and their combinations. Preferably, the drug is a protein drug or a nucleic acid drug and their combinations. The drug can be reversibly or irreversibly loaded on the mitochondrial outer membrane, inner membrane, membrane gap or mitochondrial matrix and their combinations in the form of covalent bonds or non-covalent bonds.

[0012] The drug delivery system according to the present invention can achieve the delivery of drugs into cells, wherein the cells may include the inner surface of the cell membrane, the cytoplasm, the nucleus and a combination thereof. After the drug is delivered into the cell, it can exist in a state of being combined with or separated from the mitochondria.

[0013] In the drug delivery system according to the present invention, the drug loaded on the mitochondria can be a protein drug. The protein drug can be bound to the mitochondria by any method, including but not limited to: (1) the protein drug forms a fusion protein with a mitochondrial localization sequence, such as a mitochondrial outer membrane localization sequence such as FIS1.MTS, a mitochondrial matrix localization sequence such as COX8.MTS, and binds to the mitochondria through the localization sequence; (2) the protein drug is fused with other proteins that can specifically bind to mitochondria, such as antibodies, variable regions of antibody-binding antigens, nanobodies, or other non-antibodies but amino acid sequences that can bind to mitochondria to form a fusion protein, thereby binding to the mitochondria; (3) it is bound to the mitochondria by chemical bonds such as covalent bonds, ionic bonds, secondary bonds such as van der Waals forces, hydrogen bonds, aromatic ring stacking effects, hydrophobic forces and halogen bonds, and combinations thereof.

[0014] Protein drugs loaded into mitochondria can be a single protein or a combination of multiple proteins. Protein drugs loaded into mitochondria can be complete proteins, such as GFP, or each part of a protein divided into several parts, such as GFP1-11 and GFP12 of Split-GFP. Protein drugs loaded into mitochondria can be labeled with fluorescent proteins or other protein tags (such as His-Tag, etc.), or they can be unlabeled.

[0015] In the drug delivery system according to the present invention, the medicine loaded in mitochondria can be a nucleic acid drug. Nucleic acid drugs can be bound to mitochondria by the nucleic acid binding domain of nucleic acid binding protein or nucleic acid binding protein, and can also be bound to mitochondria by chemical bonds such as covalent bonds, ionic bonds, secondary bonds such as van der Waals forces, hydrogen bonds, aromatic ring stacking effects, hydrophobic forces and halogen bonds, and combinations thereof. For example, DNA is negatively charged, and if protein X is positively charged, X-MOM (Mitochondrial outer membrane, mitochondrial outer membrane localization sequence) is anchored to the outer membrane of mitochondria, then DNA can be combined with the protein X, and then loaded on the outer membrane of mitochondria. The nucleic acid drugs loaded in mitochondria can be a gene or multiple genes.

[0016] Nucleic acid drugs can be DNA, RNA, analogs of DNA and RNA, and combinations thereof. DNA can be circular or linear, single-stranded or double-stranded, including but not limited to plasmids, PCR products, genomic DNA, etc.; RNA includes but is not limited to mRNA, miRNA, shRNA, siRNA or aptomer. DNA or RNA analogs refer to substances that can transcribe, replicate, or bind to DNA or RNA through base complementarity, such as morpholino oligos, etc.

[0017] In yet another aspect, the present invention also provides use of the above-mentioned drug delivery system in delivering drugs, such as protein drugs, nucleic acid drugs or other macromolecular drugs.

[0018] In yet another aspect, the present invention also provides the above-mentioned drug delivery system in the preparation of a preparation and / or composition for treating and / or preventing disease, beauty, weight loss, anti-fatigue, promoting growth and development, enhancing body function or delaying aging. The preparation or composition prepared using the drug delivery system of the present invention can be used for treating and / or preventing diseases including nervous system diseases, immune system diseases, respiratory system diseases, circulatory system diseases, urinary system diseases, reproductive system diseases, motor system diseases, endocrine system diseases, digestive system diseases, single gene genetic diseases, polygenic genetic diseases, infectious diseases, tumors, diabetes, neurodegenerative diseases or cardiovascular and cerebrovascular diseases and combinations thereof. BRIEF DESCRIPTION OF THE DRAWINGS

[0019] The above and other objects, features and other advantages of the present invention will be more clearly understood through the following detailed description taken in conjunction with the accompanying drawings, in which:

[0020] Figure 1It is shown that the target protein NeoR in Example 1 of the present invention is loaded on the outer membrane of the mitochondria through the mitochondrial outer membrane targeting sequence and then introduced into the cell. Among them, (A) CellMask Deep-red marks the cell membrane. (B) The red fluorescent protein mScarlet-i shows the target protein NeoR. (C) The cell membrane signal overlaps with the target protein signal, indicating that the target protein NeoR is delivered into the cell.

[0021] Figure 2 It is shown that the target protein NeoR in Example 2 of the present invention is loaded on the outer membrane of the mitochondria by the optogenetic protein pair and then introduced into the cell. Among them, (A) CellMask Deep-red marks the cell membrane. (B) The red fluorescent protein mScarlet-i shows the target protein NeoR. (C) The cell membrane signal overlaps with the target protein signal, indicating that the target protein NeoR is delivered into the cell.

[0022] Figure 3 It is shown that according to Example 3 of the present invention, mitochondria simultaneously deliver two target proteins into the cell. Among them, (A) CellMask Deep-red marks the cell membrane. (B) Red fluorescent protein mScarlet-i shows target protein 1 (Catalase). (C) Green fluorescent protein GFP shows target protein 2 (IL10). (D) The cell membrane signal overlaps with the target protein signal, showing that the target proteins Catalase and IL10 are delivered into the cell.

[0023] Figure 4 The target protein is introduced into the cell by binding to the antibody of the mitochondrial outer membrane protein according to Example 4 of the present invention. Among them, (A) CellMask Deep-red staining marks the cell membrane. (B) GFP marks the mitochondrial outer membrane and simulates the mitochondrial outer membrane protein. (C) The target protein NeoR connected to the anti-GFP nanobody. (D) After overlapping, it shows that the target protein NeoR has been introduced into the cell.

[0024] Figure 5 It shows that according to Example 5 of the present invention, plasmid DNA was introduced into cells using mitochondria as a vector and successfully expressed. Wherein, (A) bright field, showing the cell outline. (B) shows GFP marking the cell nucleus (bright signal in the middle) and mitochondria (weaker signal around). (C) after overlapping, it shows that some cells successfully express GFP marking mitochondria and cell nuclei, indicating that the plasmid DNA has been successfully introduced into the cell and expressed.

[0025] Figure 6It shows that according to Example 6 of the present invention, mRNA is introduced into cells using mitochondria as carriers and successfully expressed. Among them, (A) CellMask Deep-red staining marks the cell membrane. (B) Red fluorescent protein marking mitochondria indicates that Mito-mScarlet-i has been introduced into cells and translated. (C) Overlapping (A) and (B) shows that the translated protein is located in the cell, indicating that the mRNA is successfully delivered into the cell and expressed. DETAILED DESCRIPTION

[0026] The present invention will now be described in more detail with reference to the following examples. These examples are provided only to illustrate the present invention and should not be construed as limiting the scope and spirit of the present invention.

[0027] In one embodiment of the present invention, the drug can be loaded on the outer membrane of the mitochondria by fusing with the FIS1.MTS protein, wherein the FIS1.MTS protein is a targeting polypeptide (MTS, mitochondrial targeting sequence) of the mitochondrial outer membrane protein FIS1. Since the drug contains the targeting portion of FIS1 after being fused with the FIS1.MTS protein, it can be loaded on the outer membrane of the mitochondria.

[0028] In one embodiment of the present invention, the drug can be fused with a fluorescent protein and a FIS1.MTS protein, and then loaded on the surface of the mitochondria. Wherein, the role of the fluorescent protein is to label the drug, which is only for tracking and monitoring the delivery of the drug, and is not a necessary means to achieve drug delivery. Fluorescent proteins include but are not limited to green fluorescent protein GFP, red fluorescent protein RFP, enhanced green fluorescent protein EGFR, red fluorescent protein mScarlet-I, etc.

[0029] In one embodiment of the present invention, the drug is loaded onto the outer membrane of the mitochondria by optogenetics, for example, by pMag and nMag, Cry2-CIB or any other optogenetic protein pair. Specifically, in one embodiment of the present invention, for example, the drug forms a fusion protein with pMag, and nMag forms a fusion protein with a mitochondrial outer membrane localization sequence such as FIS1.MTS protein, and the pMag and nMag protein pairs are paired and combined under the action of blue light, so that the drug is loaded onto the outer membrane of the mitochondria.

[0030] In some embodiments of the present invention, in addition to optogenetic protein pairs, the drug delivery system according to the present invention may also use protein pairs bound under the action of chemical substances, such as FKBP12 and FRB regulated by rapamycin or any other reversibly or irreversibly bound protein pairs. In some embodiments of the present invention, a pair of protein pairs may be used between the drug and the mitochondrial outer membrane localization sequence, or multiple pairs of protein pairs may be used, and one type of protein pair may be used, or multiple types of protein pairs may be used.

[0031] In one embodiment of the present invention, the drug can be fused with the fluorescent protein and the above protein pair, and then loaded on the outer membrane of the mitochondria. Among them, the fluorescent protein plays the role of marking the drug, which is only for tracking and monitoring the delivery of the drug, and is not necessary for achieving drug delivery. Fluorescent proteins include but are not limited to green fluorescent protein GFP, red fluorescent protein RFP, enhanced green fluorescent protein EGFR, red fluorescent protein mScarlet-I, etc.

[0032] In one embodiment of the present invention, the drug delivery system according to the present invention is prepared by a method comprising the following steps:

[0033] (1) Designing an XY-FIS1.MTS protein, wherein the protein comprises three parts: X is any target drug; Y is a fluorescent protein used to label the target drug; and FIS1.MTS is a targeting polypeptide of the mitochondrial outer membrane protein FIS1;

[0034] (2) constructing a plasmid of XY-FIS1.MTS protein using pcDNA3.1 vector and transfecting HEK 293T cells; and

[0035] (3) After successful expression, XY-FIS1.MTS-labeled mitochondria were extracted from the cells and then co-cultured with unlabeled cells.

[0036] In one embodiment of the present invention, the drug delivery system according to the present invention is prepared by a method comprising the following steps:

[0037] (1) Design X-Y1-pMag protein and nMagHigh1-Y2-FIS1.MTS protein respectively;

[0038] (2) constructing plasmids of the two proteins using pcDNA3.1 vector and transfecting HEK 293T cells to simultaneously express the two proteins;

[0039] (3) 24 hours later, irradiating the cells with blue light for 10 minutes to allow the pMag and nMagHigh1 protein pairs to bind; and

[0040] (4) Y1- and Y2-labeled mitochondria were extracted from the cells and then co-cultured with unlabeled cells.

[0041] Among them, X is any target drug, Y1 and Y2 are fluorescent proteins for labeling, pMag and nMagHigh1 are a pair of optogenetic proteins, and FIS1.MTS is a targeting peptide of the mitochondrial outer membrane protein FIS1.

[0042] Furthermore, the fluorescent protein may be GFP, EGFP, mScarlet-i, YFP, CFP or other fluorescent proteins.

[0043] The plasmid vector may be pcDNA3.1, pEGFP, pCMVp-NEO-BAN, pSV2pUC19 DNA, pUC18 DNA, PQE30 or any other commonly used cloning vector.

[0044] The transfected cells can be HEK 293T, CHO-K1, COS-7, HeLa, NIH-3T3, PC-3, A549, MCF-7, HuH-7, U-2OS, HepG2 or any other transfectable cells.

[0045] Transfection reagents can be Lipofectamine, X-tremeGENE 9, Escort TM III. Escort TM IV. NeuroPorter TM DOTAP methosulfate, Calcium Phosphate or any other transfection reagent.

[0046] The transfection method can be liposome transfection, calcium phosphate transfection, electroporation transfection or any other transfection method.

[0047] The co-culture time of mitochondria and cells can be 0.5-120 hours, preferably 6-24 hours.

[0048] <Example 1>

[0049] The protein NeoR-mScarlet-i-FIS1.MTS was designed. NeoR is a neomycin-resistant protein, which is the target drug; mScarlet-i is a red fluorescent protein, which is used to label the target drug; FIS1.MTS is the targeting sequence of the mitochondrial outer membrane protein FIS1, so that the NeoR-mScarlet-i-FIS1.MTS protein can be targeted to the outer membrane of the mitochondria. The NeoR-mScarlet-i-FIS1.MTS plasmid (see sequence table, SEQ ID NO: 1) was constructed using the pcDNA3.1 vector and transfected into HEK293T cells. After 24 hours of successful expression, the mitochondria labeled with NeoR-mScarlet-i-FIS1.MTS in the HEK 293T cells, i.e., the mitochondria loaded with the NeoR-mScarlet-i-FIS1.MTS fusion protein, were extracted and then co-cultured with unlabeled 293A cells for 6 hours. The cell membrane was stained with the cell membrane dye CellMask Deep-red, and then a laser confocal microscope was used to observe that some mitochondria in 293A cells were mScarlet-i positive, indicating that the target drug NeoR had been successfully delivered into 293A cells. Figure 1 shown.

[0050] <Example 2>

[0051] The proteins NeoR-mScarlet-i-pMag and nMagHigh1-GFP-FIS1.MTS were designed. NeoR is the target drug, and mScarlet-i is a red fluorescent protein used to label the target drug. The nMagHigh1-GFP-FIS1.MTS fusion protein can bind to the outer membrane of mitochondria through the mitochondrial outer membrane targeting sequence FIS1.MTS. pMag and nMagHigh1 are a pair of optogenetic proteins that can bind to each other under blue light irradiation. Therefore, under the action of blue light, the fusion protein NeoR-mScarlet-i-pMag and the fusion protein nMagHigh1-GFP-FIS1.MTS bind to each other, so that the target drug NeoR is loaded on the outer membrane of mitochondria. Specifically, the NeoR-mScarlet-i-pMag plasmid (see the sequence list, SEQ ID NO: 2) and the nMagHigh1-GFP-FIS1.MTS plasmid (see the sequence list, SEQ ID NO: 3) were constructed using the pcDNA3.1 vector, and pcDNA3.1-NeoR-mScarlet-i-pMag and pcDNA3.1-nMagHigh1-GFP-FIS1.MTS were co-transfected into 293T cells. After 24 hours, the cells were irradiated with strong light for 10 minutes to allow the NeoR-mScarlet-i-pMag fusion protein to bind to the nMagHigh1-GFP-FIS1.MTS fusion protein, thereby loading the target protein NeoR onto the outer membrane of the mitochondria. Then, mitochondria loaded with NeoR protein (mScarlet-i marker) were extracted and diluted with DMEM complete medium, and then added to the cultured unmarked 293A cells. After incubation at 37°C for 6-8 hours, the cell membrane was stained with the cell membrane dye CellMask Deep-red, and then imaged with a laser confocal microscope. It was found that part of the target protein NeoR marked with mScarlet-i had entered the 293A cells. The test results are as follows Figure 2 shown.

[0052] <Example 3>

[0053] Plasmids pcDNA3.1-NeoR-mScarlet-i-NB.GFP (see sequence table, SEQ ID NO: 4) and pcDNA3.1-GFP-FIS1.MTS (see sequence table, SEQ ID NO: 5) were constructed. NeoR is the target drug, mScarlet-i is a red fluorescent protein used to mark the target drug, and NB.GFP is a nanobody that can specifically bind to GFP. The role of GFP is to mark mitochondria and simulate mitochondrial outer membrane proteins, and FIS1.MTS is the targeting sequence of mitochondrial outer membrane protein FIS1. Two 10 cm culture dishes of 293T cells were transfected with the two plasmids, and cells expressing NeoR-mScarlet-i-NB.GFP were harvested 24 hours later. The cells were swollen with distilled water for 5 minutes, and then 1 / 9 volume of 10X PBS was added, and the cells were broken by a glass homogenizer, centrifuged at 10000g for 10 minutes, and the supernatant containing NeoR-mScarlet-i-NB.GFP was taken for use. Cells expressing GFP-FIS1.MTS were harvested, mitochondria were extracted, and resuspended with supernatant containing NeoR-mScarlet-i-NB.GFP. After being placed at room temperature for 10 minutes, they were co-cultured with unlabeled 293A cells. After incubation at 37°C for 6-8 hours, the cell membrane was stained with the cell membrane dye CellMask Deep-red, and then imaged with a laser confocal microscope. It was found that some GFP-labeled mitochondria had successfully transferred the mScarlet-i-labeled NeoR into the cells. The test results are as follows: Figure 3 shown.

[0054] <Example 4>

[0055] Designed proteins COX8.MTS-hCatalase-mScarlet-i, IL10-mNeonGreen-FIS1.MTS. In the protein COX8.MTS-hCatalase-mScarlet-i, hCatalase is human Catalase, which is the target drug, mScarlet-i is a red fluorescent protein used to label the target drug, COX8.MTS is the targeting sequence of the mitochondrial matrix protein COX8, and the protein COX8.MTS-hCatalase-mScarlet-i can be localized to the matrix of mitochondria through the targeting sequence COX8.MTS. In the protein IL10-mNeonGreen-FIS1.MTS, IL10 is the target drug, mNeonGreen is a high-brightness green fluorescent protein used to label the target drug, and the protein IL10-mNeonGreen-FIS1.MTS can bind to the outer membrane of mitochondria through the targeting sequence FIS1.MTS. Specifically, the pcDNA3.1 vector was used to construct the COX8.MTS-hCatalase-mScarlet-i plasmid (see the sequence list, SEQ ID NO: 6) and the IL10-mNeonGreen-FIS1.MTS plasmid (see the sequence list, SEQ ID NO: 7), and the two plasmids were co-transfected into 293T cells. After 24 hours, the mitochondria loaded with hCatalase (mScarlet-i labeled) and / or IL10 (mNeonGreen labeled) were extracted, and the extracted mitochondria were diluted with DMEM complete medium and added to the cultured unlabeled 293A cells. After incubation at 37°C for 6-8 hours, the cell membrane was stained with the cell membrane dye CellMask Deep-red, and then imaged and detected using a laser confocal microscope, and it was found that some of the hCatalase labeled with mNeonGreen and the IL10 labeled with mScarlet-i had entered the cells. The test results are as follows: Figure 4 shown.

[0056] <Example 5>

[0057] Plasmid pcDNA3.1-hTFAM1-CRT-mScarlet-i-FIS1.MTS (see sequence table, SEQ ID NO: 8) was constructed, wherein hTFAM1 is the DNA binding domain of human TFAM1, and CRT is the DNA binding domain of TopII alpha, both of which can bind to DNA. mScarlet-i is a red fluorescent protein used to mark mitochondria carrying fusion proteins; FIS1.MTS is a signal peptide targeting the outer membrane of mitochondria. Mouse 3T3 cells were transfected with pcDNA3.1-hTFAM1-CRT-mScarlet-i-FIS1.MTS, and the fusion protein was expressed 24 hours later. Ten 7 Mitochondria were extracted from each cell, resuspended in 100 μL DMEM complete medium, and 200 ng of plasmid pcDNA3.1-Mito-EGFP-2A-n.EGFP (see sequence table, SEQ ID NO: 9) was added. The cells were incubated at room temperature for 1 hour. 2A can translate one mRNA into two proteins, Mito-EGFP marks mitochondria, and nEGFP marks the nucleus. The mitochondria were then co-cultured with unlabeled 293A cells. After 24 hours, fluorescence microscopy imaging showed that some cells expressed GFP marking mitochondria and nuclei, indicating that the plasmid had been successfully transferred into the cells and expressed. The test results are shown in Figure 2. Figure 5 shown.

[0058] <Example 6>

[0059] The plasmid pcDNA3.1-UP1-FIS1.MTS (see sequence list, SEQ ID NO: 10) was constructed, wherein UP1 is the nonspecific RNA binding domain of human hnRNP A1 protein, and FIS1.MTS is a signal peptide targeting the outer membrane of mitochondria. 293T cells were transfected with pcDNA3.1-UP1-FIS1.MTS, and 10 7 Mitochondria were extracted from cells and resuspended in 100 μL DMEM complete medium. 200 ng of Mito-mScarlet-i mRNA (transcribed in vitro from plasmid pcDNA3.1-Mito-mScarlet-I (see sequence table, SEQ ID NO: 11)) was added and incubated at room temperature for 0.5 hours. The mitochondria were then co-cultured with unlabeled 293A cells. After 24 hours, the cell membrane was stained with the cell membrane dye CellMask Deep-red. Laser confocal microscopy imaging showed red signals expressing labeled mitochondria in some cells, indicating that the mRNA had been successfully transferred into the cells and translated. The test results are shown in Figure 2. Figure 6 shown. SEQUENCE LISTING <110> Mai Tang Biological <120> A mitochondrial-based drug delivery system and its use <130> Mito0002 <160> 11 <170> PatentIn version 3.5 <210> 1 <211> 7135 <212> DNA <213> Artificial sequence <400> 1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg 60 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg 120 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc 180 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt 240 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata 300 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc 360 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc 420 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt 480 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt 540 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca 600 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg 660 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc 720 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg 780 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca 840 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagaccacaa gctggctagc 900 atgattgaac aagatggatt gcagcaggt tctccggccg cttgggtgga gaggctattc 960 ggctatgact gggcacaaca gacaatcggc tgctctgatg ccgccgtgtt ccggctgtca 1020 gcgcaggggc gcccggttct ttttgtcaag accgacctgt ccggtgccct gaatgaactg 1080 caggacgagg cagcgcggct atcgtggctg gccacgacgg gcgttccttg cgcagctgtg 1140 ctcgacgttg tcactgaagc gggaagggac tggctgctat tgggcgaagt gccggggcag 1200 gatctcctgt catctcacct tgctcctgcc gagaagtat ccatcatggc tgatgcaatg 1260 cggcggctgc atacgcttga tccggctacc tgcccattcg accaccaagc gaaacatcgc 1320 atcgagcgag cacgtactcg gatggaagcc ggtcttgtcg atcaggatga tctggacgaa 1380 gagcatcagg ggctcgcgcc agccgaactg ttcgccaggc tcaaggcgcg catgcccgac 1440 ggcgaggatc tcgtcgtgac ccatggcgat gcctgcttgc cgaatatcat ggtggaaaat 1500 ggccgctttt ctggattcat cgactgtggc cggctgggtg tggcggaccg ctatcaggac 1560 atagcgttgg ctacccgtga tattgctgaa gagcttggcg gcgaatgggc tgaccgcttc 1620 ctcgtgcttt acggtatcgc cgctcccgat tcgcagcgca tcgccttcta tcgccttctt 1680 gacgagttct tctcgggatc cgaaaatctc tacttccaga gtggaagttc tggttcaagt 1740 ggttccggct cggcaagtgg ttccggcatg gtgagcaagg gcgaggcagt gatcaaggag 1800 ttcatgcggt tcaaggtgca catggagggc tccatgaacg gccacgagtt cgagatcgag 1860 ggcgagggcg agggccgccc ctacgagggc acccagaccg ccaagctgaa ggtgaccaag 1920 ggtggccccc tgcccttctc ctgggacatc ctgtcccctc agttcatgta cggctccagg 1980 gccttcatca agcaccccgc cgacatcccc gactactata agcagtcctt cccgaggc 2040 ttcaagtggg agcgcgtgat gaacttcgag gacggcggcg ccgtgaccgt gacccaggac 2100 acctccctgg aggacggcac cctgatctac aggtgaagc tccgaggcac caactccct 2160 cctgacggcc ccgtaatgca gagagaca atgggctggg aagcgtccac cgagcggttg 2220 taccccgagg acggcgtgct gaagggcgac attaagatgg ccctgcgcct gaggacgga 2280 ggccgctacc tggcggactt caaccacc tacaagcca agagcccgt gcagatgccc 2340 ggcgcctaca acgtcgaccg caagttggac atcaccccc acacgagga ctacaccgtg 2400 gtggaacagt acgaacgctc cgaggggccgc cactccaccg gcggcatgga cgagctatac 2460 aagtccagct cgggctccag tggaggtca ggctcgtcta gtagctccgg agggtcaagc 2520 ggtaccggaa tggaaccgca gaacaaccag gcgaagaac tggaacgcct gattgataaa 2580 gcgatgaaaa aagatggcct gtgggcatg gcgattgtgg gcggcatggc gctggggcgtg 2640 gcggggcctgg cgggcctgat tggcctggcg gtgagcaaaa gcaaagcta gtctagatag 2700 gggcccgttt aaacccgctg atcagcctcg actgtgcctt ctagttgcca gccatctgtt 2760 gttgcccct cccccgtgcc ttccttgacc ctggaaggtg ccactcccac tgtccttttcc 2820 tataaaatg aggaaattgc atcgcattgt ctgagtaggt gtcattctat tctggggggt 2880 ggggtggggc aggacagcaa gggggaggat tgggaagaca atagcaggca tgctggggat 2940 gcggtgggct ctatggcttc tgaggcggaa agaaccagct ggggctctag ggggtatccc 3000 cacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc 3060 gctacacttg cagcgccct agcgcccgct cctttcgctt tctccttc cttctcgcc 3120 acgtcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt 3180 agtgctttac ggcacctcga ccccaaaaaa cttgattagg gtgatggttc acgtagtggg 3240 ccatcgccct gatagacggt tttcgccct ttgacgttgg agtccacgtt cttaatagt 3300 ggactcttgt tccaaactgg aacaacactc aaccctatct cggtctattc ttttgattta 3360 taagggattt tgccgatttc ggcctattgg ttaaaaaatg agctgattta acaaaaattt 3420 aacgcgaatt aattctgtgg aatgtgtgtc agttagggtg tggaagtcc ccaggctccc 3480 cagcaggcag aagtatgcaa to catchcatc tcaattagtc to catch tgtggaagt ccccaggctc cccagcaggc agagtatgc aaagcatgca tctcaattag tcagcaacca 3660. 3660. 3660. 3660. tagtcccgcc cgccccatgg ctgactaatt ttttttattt atgcagaggc cgaggccgcc tctgcctctg 3720 agctattcca gaagtagtga ggaggctttt ttggaggcct aggcttttgc aaaaagctcc 3780 cgggagcttg tatatccatt ttcggatctg atcaagagac aggatgagga tcgtttcgca 3840. tgattgaaca agatggattg cacgcaggtt ctccggccgc ttgggtggag aggctattcg gctatgactg ggcacaacag acaatcggct gctctgatgc cgccgtgttc cggctgtcag cgcaggggcg cccggttctt tttgtcaaga ccgacctgtc cggtgccctg aatgaactgc aggacgaggc agcgcggcta tcgtggctgg ccacgacggg cgttccttgc gcagctgtgc 4080 tcgacgttgt cactgaagcg ggaagggact ggctgctatt gggcgaagtg ccggggcagg 4140 atctcctgtc atctcacctt gctcctgccg agaaagtatc catcatggct gatgcaatgc 4200 ggcggctgca tacgcttgat ccggctacct gcccattcga ccaccaagcg aaacatcgca 4260 tcgagcgagc acgtactcgg atggaagccg gtcttgtcga tcaggatgat ctggacgaag 4320 agcatcaggg gctcgcgcca gccgaactgt tcgccaggct caaggcgcgc atgcccgacg 4380 gcgaggatct cgtcgtgacc catggcgatg cctgcttgcc gaatatcatg gtggaaaatg 4440 gccgcttttc tggattcatc gactgtggcc ggctgggtgt ggcggaccgc tatcaggaca 4500 tagcgttggc tacccgtgat attgctgaag agcttggcgg cgaatgggct gaccgcttcc 4560 tcgtgcttta cggtatcgcc gctcccgatt cgcagcgcat cgccttctat cgccttcttg 4620 acgagttctt ctgagcggga ctctggggtt cgaaatgacc gaccaagcga cgcccaacct 4680 gccatcacga gatttcgatt ccaccgccgc cttctatgaa aggttgggct tcggaatcgt 4740 tttccgggac gccggctgga tgatcctcca gcgcggggat ctcatgctgg agttcttcgc 4800 ccaccccaac ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa 4860 tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa 4920 tgtatcttat catgtctgta taccgtcgac ctctagctag agcttggcgt aatcatggtc 4980 atagctgttt cctgtgtgaa attgttatcc gctcacaatt ccacacaaca tacgagccgg 5040 aagcataaag tgtaaagcct ggggtgccta atgagtgagc taactcacat taattgcgtt 5100 gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg 5160 ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga 5220 ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat 5280 acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca 5340 aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc 5400 tgacgagcat cacaaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata 5460 aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc 5520 gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc 5580 acgctgtagg tatctcagtt cggtgtaggt cgtcgctcc aagctgggct gtgtgcacga 5640 accccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc 5700 ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag 5760 gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag 5820 aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag 5880 ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca 5940 gattacgc agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga 6000 cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat 6060 cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga 6120 gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg 6180 tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga 6240 gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc 6300 agatttatca gcaataacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc 6480. gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc 6540. catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtagtt ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc 6660. atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca tcattggaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc 6900 atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa 7080 aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg acgtc 7135 <210> 2 <211> 7426 <212> DNA <213> Artificial sequence <400> 2 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg 60 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg 120 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc 180 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt 240 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata 300 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc 360 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc 420 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt 480 atcatatgcc aagtacgcccc cctattgacg tcaatgacgg taaatggccc gcctggcatt 540 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca 600 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg 660 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc 720 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg 780 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca 840 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagaccacaa gctggctagc 900 atgattgaac aagatggatt gcagcaggt tctccggccg cttgggtgga gaggctattc 960 ggctatgact gggcacaaca gacaatcggc tgctctgatg ccgccgtgtt ccggctgtca 1020 gcgcaggggc gcccggttct ttttgtcaag accgacctgt ccggtgccct gaatgaactg 1080 caggacgagg cagcgcggct atcgtggctg gccacgacgg gcgttccttg cgcagctgtg 1140 ctcgacgttg tcactgaagc gggaagggac tggctgctat tgggcgaagt gccggggcag 1200 gatctcctgt catctcacct tgctcctgcc gagaaagtat ccatcatggc tgatgcaatg 1260 cggcggctgc atacgcttga tccggctacc tgcccattcg accaccaagc gaaacatcgc 1320 atcgagcgag cacgtactcg gatggaagcc ggtcttgtcg atcaggatga tctggacgaa 1380 gagcatcagg ggctcgcgcc agccgaactg ttcgccaggc tcaaggcgcg catgcccgac 1440 ggcgaggatc tcgtcgtgac ccatggcgat gcctgcttgc cgaatatcat ggtggaaaat 1500 ggccgctttt ctggattcat cgactgtggc cggctgggtg tggcggaccg ctatcaggac 1560 atagcgttgg ctacccgtga tattgctgaa gagcttggcg gcgaatgggc tgaccgcttc 1620 ctcgtgcttt acggtatcgc cgctcccgat tcgcagcgca tcgccttcta tcgccttctt 1680 gacgagttct tctcgggatc cgaaaatctc tacttccaga gtggaagttc tggttcaagt 1740 ggttccggct cggcaagtgg ttccggcatg gtgagcaagg gcgaggcagt gatcaaggag 1800 ttcatgcggt tcaaggtgca catggagggc tccatgaacg gccacgagtt cgagatcgag 1860 ggcgagggcg agggccgccc ctacgagggc acccagaccg ccaagctgaa ggtgaccaag 1920 ggtggcccccc tgcccttctc ctgggacatc ctgtcccctc agttcatgta cggctccagg 1980 gccttcatca agcaccccgc cgacatcccc gactactata agcagtcctt ccccgagggc 2040 ttcaagtggg agcgcgtgat gaacttcgag gacggcggcg ccgtgaccgt gacccaggac 2100 acctccctgg aggacggcac cctgatctac aaggtgaagc tccgaggcac caacttccct 2160 cctgacggcc ccgtaatgca gaagagaca atgggctggg aagcgtccac cgagcggttt 2220 taccccgagg acggcgtgct gaagggcgac attaagatgg ccctgcgcct gaaggacgga 2280 ggccgctacc tggcggactt caagaccacc tacaaggcca agaagcccgt gcagatgccc 2340 ggcgcctaca acgtcgaccg caagttggac atcacctccc aacaggag ctacaccgtg 2400 gtggaacagt acgaacgctc cgagggccgc cactccaccg gcggcatgga cgagctatac 2460 aagtccagct cgggctccag tggagggtca ggctcgtcta gtagctccgg agggtcaagc 2520 ggtaccggac atactcttta tgccccccggt ggatatgaca ttatgggata cctgaggcag 2580 atcaggaacc ggccaaaccc gcaggtggaa ctgggacccg tggatacatc ctgcgccttg 2640 attctttgtg acctgaaaca gaaagacacc ccgatagttt acgcgagtga agccttctc 2700 tacatgacag gttacagcaa cgcagaggtg ctgggccgga attgccggtt tctgcaaagc 2760 cctgacggca tggtgaagcc caagagcacc cggaagtacg tggatagtaa cacaatcaat 2820 actatgcgca aggcaatcga caggaatgcc gaggtgcagg ttgaagtagt caattttaaa 2880 aagaatggac agcgatttgt taatttcctg actatgatac ctgttaggga cgaaacaggc 2940 gagtatcgat actctatggg attccagtgc gaaacagaat agtctagat ggggcccgtt 3000 taaacccgct gatcagcctc gactgtgcct tctagttgcc agccatctgt tgtttgcccc 3060 tccccccgtgc cttccttgac cctggaaggt gccactccca ctgtcctttc ctaataaaat 3120 gaggaaattg catcgcattg tctgagtagg tgtcattcta ttctgggggg tggggtgggg 3180 caggacagca aggggagga ttgggaagac aatagcaggc atgctggggga tgcggtgggc 3240 tctatggctt ctgaggcgga aagaaccagc tggggctcta gggggtatcc ccacgcgcc 3300 tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt 3360 gccagcgccc tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc 3420. ggctttcccc gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta 3480 cggcacctcg accccaaaaa acttgattag ggtgatggtt cacgtagtgg gccatcgccc 3540 tgatagcgg tttttcgccc tttgacgttg gagtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact caaccctatc tcggtctatt cttttgattt ataagggatt ttgccgattt cggcctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat taattctgtg gaatgtgtgt cagttagggt gtggaagtc cccaggctcc ccagcaggca gagtatgca aagcatgcat ctcaattagt cagcaaccag gtgtggaag tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc atagtcccgc ccctaactcc gcccatcccg cccctaactc cgcccagttc cgcccattct ccgccccatg 3960 gctgactaat tttttttatt tatgcagagg ccgaggccgc ctctgcctct gagctattcc agagtagtg aggaggcttt tttggaggcc tagctttttg caaaagctc ccggggagctt 4080. gtatatccat tttcggatct gatcaagaga caggatgagg atcgtttcgc atgattgaac 4140 aagatggatt gcacgcaggt tctccggccg cttgggtgga gaggctattc ggctatgact 4200 gggcacaaca gacaatcggc tgctctgatg ccgccgtgtt ccggctgtca gcgcaggggc 4260 gcccggttct ttttgtcaag accgacctgt ccggtgccct gaatgaactg caggacgagg 4320 cagcgcggct atcgtggctg gccacgacgg gcgttccttg cgcagctgtg ctcgacgttg 4380 tcactgaagc gggaagggac tggctgctat tgggcgaagt gccggggcag gatctcctgt 4440 catctcacct tgctcctgcc gagaaagtat ccatcatggc tgatgcaatg cggcggctgc 4500 atacgcttga tccggctacc tgcccattcg accaccaagc gaaacatcgc atcgagcgag 4560 cacgtactcg gatggaagcc ggtcttgtcg atcaggatga tctggacgaa gagcatcagg 4620 ggctcgcgcc agccgaactg ttcgccaggc tcaaggcgcg catgcccgac ggcgaggatc 4680 tcgtcgtgac ccatggcgat gcctgcttgc cgaatatcat ggtggaaaat ggccgctttt 4740 ctggattcat cgactgtggc cggctgggtg tggcggaccg ctatcaggac atagcgttgg 4800 ctacccgtga tattgctgaa gagcttggcg gcgaatgggc tgaccgcttc ctcgtgcttt 4860 acggtatcgc cgctcccgat tcgcagcgca tcgccttcta tcgccttctt gacgagttct 4920 tctgagcggg actctggggt tcgaaatgac cgaccaagcg acgcccaacc tgccatcacg 4980 agatttcgat tccaccgccg ccttctatga aaggttgggc ttcggaatcg ttttccggga 5040 cgccggctgg atgatcctcc agcgcgggga tctcatgctg gagttcttcg cccaccccaa 5100 cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa atttcacaaa 5160 taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca atgtatctta 5220 tcatgtctgt ataccgtcga cctctagcta gagcttggcg taatcatggt catagctgtt 5280 tcctgtgtga aattgttatc cgctcacaat tccacacaac atacgagccg gaagcataaa 5340 gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca ttaattgcgt tgcgctcact 5400 gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc 5460 ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg 5520 ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc 5580 cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag 5640 gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca 5700 tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca 5760 ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg 5820 atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag 5880 gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt 5940 tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca 6000 cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg 6060 cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa gaacagtatt 6120 tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc 6180 cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg 6240 cagaaaaaaa ggatctcaag aagatccttt gatctttct acggggtctg acgctcagtg 6300. gacgaaac tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat gagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc 6540 atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc cagatttatc 6600 agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga agctaggc agtagttcgc cagttaatag 6720. tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat 6780 ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg 6840. caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt 6900 gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg 7020 accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt 7080 aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct 7140 gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac 7200 tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat 7260 aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat 7320 ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca 7380 aataggggtt ccgcgcacat ttccccgaaa agtgccacct gacgtc 7426 <210> 3 <211> 6801 <212> DNA <213> Artificial Sequence <400> 3 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg 60 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg 120 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc 180 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt 240 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata 300 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc 360 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc 420 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt 480 atcatatgcc aagtacgcccc cctattgacg tcaatgacgg taaatggccc gcctggcatt 540 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca 600 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg 660 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc 720 aaaatcaacg ggactttcca aaatgcgta acaactccgc cccattgacg caaatgggcg 780 gtaggcgtgt acggtgggag gtctataataa gcagagctct ctggctaact agagaaccca 840 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc 900 ctagcatgca tactctttat gcccccggtg gatatgacat tatgggttat ctggaccaga 960 tcggcaaccg gccaaacccg caggtggaac tgggtcccgt ggatacatcc tgcgccttga 1020 ttctttgtga cctgaaacag aaagacaccc cgatagttta cgcgagtgaa gccttcctct 1080 acatgacagg ttacagcaac gcagaggtgc tgggccggaa ttgccggttt ctgcaaagcc 1140 ctgacggcat ggtgaagccc aagagcaccc ggaagtacgt ggatagtaac acaatcaata 1200 ctatccgcaa ggcaatcgac aggaatgccg aggtgcaggt tgaagtagtc aattttaaaa 1260 agaatggaca gcgatttgtt aatttcctga ctatcatacc tgttagggac gaaacaggcg 1320 agtatcgata ctctatggga ttccagtgcg aaacagaatc gggatccgga agttctggtt 1380 caagtggttc cggctcggca agtggttccg gcatggtgag caagggcgag gagctgttca 1440 ccggggtggt gcccatcctg gtcgagctgg acggcgacgt aaacggccac aagttcagcg 1500 tgtccggcga gggcgagggc gatgccacct acggcaagct gaccctgaag ttcatctgca 1560 ccaccggcaa gctgcccgtg ccctggccca ccctcgtgac caccctgacc tacggcgtgc 1620 agtgcttcag ccgctacccc gaccacatga agcagcacga cttcttcaag tccgccatgc 1680 ccgaaggcta cgtccaggag cgcaccatct tcttcaagga cgacggcaac tacaagaccc 1740 gcgccgaggt gaagttcgag ggcgacaccc tggtgaaccg catcgagctg aagggcatcg 1800 acttcaagga ggacggcaac atcctggggc acaagctgga gtacaactac aacagccaca 1860 acgtctatat catggccgac aagcagaaga acggcatcaa ggtgaacttc aagatccgcc 1920 acaacatcga ggacggcagc gtgcagctcg ccgaccacta ccagcagaac acccccatcg 1980 gcgacggccc cgtgctgctg cccgacaacc actacctgag cacccagtcc gccctgagca 2040 aagaccccaa cgagaagcgc gatcacatgg tcctgctgga gttcgtgacc gccgccggga 2100 tcactctcgg catggacgag ctatacaagt ccagctcggg ctccagtgga gggtcaggct 2160 cgtctagtag ctccggaggg tcaagcggta ccggaatgga accgcagaac aaccaggcga 2220 aagaactgga acgcctgatt gataaagcga tgaaaaaaga tggcctggtg ggcatggcga 2280 <h2 style=";text-align:left;direction:ltr">ttgtgggcgg catggcgctg ggcgtggcgg gcctggcggg cctgattggc ctggcggtga 2340<h2 style=";text-align:left;direction:ltr"> <h2 style=";text-align:left;direction:ltr"> gcaaaagcaa aagctagtct agataggggc ccgtttaaac ccgctgatca gcctcgactg 2400<h2 style=";text-align:left;direction:ltr"> <h2 style=";text-align:left;direction:ltr"> tgccttctag ttgccagcca tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg 2460<h2 style=";text-align:left;direction:ltr"> <h2 style=";text-align:left;direction:ltr"> aaggtgccac tcccactgtc ctttcctaat aaaatgagga aattgcatcg cattgtctga 2520<h2 style=";text-align:left;direction:ltr"> <h2 style=";text-align:left;direction:ltr"> gtaggtgtca ttctattctg gggggtgggg tggggcagga cagcaagggg gaggattggg 2580<h2 style=";text-align:left;direction:ltr"> <h2 style=";text-align:left;direction:ltr"> aagacaatag caggcatgct ggggatgcgg tgggctctat ggcttctgag gcggaaagaa 2640<h2 style=";text-align:left;direction:ltr"> <h2 style=";text-align:left;direction:ltr"> ccagctgggg ctctaggggg tatccccacg cgccctgtag cggcgcatta agcgcggcgg 2700<h2 style=";text-align:left;direction:ltr"> <h2 style=";text-align:left;direction:ltr"> gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccctagcg cccgctcctt 2760<h2 style=";text-align:left;direction:ltr"> <h2 style=";text-align:left;direction:ltr"> tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc 2820<h2 style=";text-align:left;direction:ltr"> <h2 style=";text-align:left;direction:ltr"> gggggctccc tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg 2880<h2 style=";text-align:left;direction:ltr"> <h2 style=";text-align:left;direction:ltr"> attagggtga tggttcacgt agtggggccat cgccctgata gacggttttt cgccctttga 2940<h2 style=";text-align:left;direction:ltr"> <h2 style=";text-align:left;direction:ltr"> cgttggagtc cacgttcttt aatagtggac tcttgttcca aactggaaca acactcaacc 3000<h2 style=";text-align:left;direction:ltr"> ctatctcggt ctattctttt gatttataag ggattttgcc gatttcggcc tattggttaa 3060 aaaatgagct gatttaacaa aaatttaacg cgaattaatt ctgtggaatg tgtgtcagtt 3120 agggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa 3180 ttagtcagca accaggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag 3240 catgcatctc aattagtcag caaccatagt cccgccccta actccgccca tcccgcccct 3300 aactccgccc agttccgccc attctccgcc ccatggctga ctaatttttt ttatttatgc 3360 agaggccgag gccgcctctg cctctgagct attccagaag tagtgaggag gcttttttgg 3420 aggcctaggc ttttgcaaaa agctcccggg agcttgtata tccattttcg gatctgatca 3480 agagacagga tgaggatcgt ttcgcatgat tgaacaagat ggattgcacg caggttctcc 3540 ggccgcttgg gtggagaggc tattcggcta tgactgggca caacagacaa tcggctgctc 3600 tgatgccgcc gtgttccggc tgtcagcgca ggggcgcccg gttctttttg tcaagaccga 3660 cctgtccggt gccctgaatg aactgcagga cgaggcagcg cggctatcgt ggctggccac 3720 gacgggcgtt ccttgcgcag ctgtgctcga cgttgtcact gagcggga gggactggct 3780 gctattgggc gaagtgccgg ggcaggatct cctgtcatct caccttgctc ctgccgagaa 3840 agtatccatc atggctgatg caatgcggcg gctgcatacg cttgatccgg ctacctgccc 3900 attcgaccac caagcgaaac atcgcatcga gcgagcacgt actcggatgg aagccggtct 4020. tgtcgatcag gatgatctgg acgaagagca tcaggggctc gcgccagccg aactgttcgc caggctcaag gcgcgcatgc ccgacggcga ggatctcgtc gtgacccatg gcgatgcctg 4080 4140. cttgccgaat atcatggtgg aaaatggccg cttttctgga ttcatcgact gtggccggct gggtgtggcg gaccgctatc aggacatagc gttggctacc cgtgatattg ctgaagagct 4200 tggcggcgaa tgggctgacc gcttcctcgt gctttacggt atcgccgctc ccgattcgca 4260. gcgcatcgcc ttctatcgcc ttcttgacga gttcttctga gcgggactct ggggttcga 4320 atgaccgacc aagcgacgcc caacctgcca tcacgagatt tcgattccac cgccgccttc tatgaaaggt tgggcttcgg aatcgttttc cgggacgccg gctggatgat cctccagcgc 4440 ggggatctca tgctggagtt cttcgcccac cccaacttgt ttattgcagc ttataatggt 4500 tacaaataaa gcaatagcat cacaaatttc acaaataaag catttttttc actgcattct 4560 agttgtggtt tgtccaaact catcaatgta tcttatcatg tctgtatacc gtcgacctct 4620 agctagagct tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc 4680 acaattccac acaacatacg agccggaagc ataaagtgta aagcctgggg tgcctaatga 4740 gtgagctaac tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg 4800 tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg 4860 cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg 4920 gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga 4980 aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg 5040 gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag 5100 aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc 5160 gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg 5220 ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt 5280 cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc 5340 ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc 5400 actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg 5460 tggcctaact acggctcac tagagaaca gtatttggta tctgcgctct gctgaagcca 5520 gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc 5580 ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat 5640 cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt 5700 ttggtcatga gattatcaaa aggatcttc acctagatcc ttttaaatta aaaatgaagt 5760 tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc 5820 agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc 5880 gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata 5940 ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg 6000 gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc 6060 cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct 6120 acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa 6180 cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt 6240 cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca 6300 ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac 6360 tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca 6420 atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt 6480 tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc 6540 actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca 6600 aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata 6660 ctcatactct tcctttttca atattattga agcatttatc agggttattg tctcatgagc 6720 ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc 6780 cgaaaagtgc cacctgacgt c 6801 <210> 4 <211> 7327 <212> DNA <213> Artificial sequence <400> 4 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg 60 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg 120 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc 180 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt 240 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata 300 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc 360 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc 420 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt 480 atcatatgcc aagtacgcccc cctattgacg tcaatgacgg taaatggccc gcctggcatt 540 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca 600 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg 660 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc 720 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg 780 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca 840 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagaccacaa gctggctagc 900 atgattgaac aagatggatt gcagcaggt tctccggccg cttgggtgga gaggctattc 960 ggctatgact gggcacaaca gacaatcggc tgctctgatg ccgccgtgtt ccggctgtca 1020 gcgcaggggc gcccggttct ttttgtcaag accgacctgt ccggtgccct gaatgaactg 1080 caggacgagg cagcgcggct atcgtggctg gccacgacgg gcgttccttg cgcagctgtg 1140 ctcgacgttg tcactgaagc gggaagggac tggctgctat tgggcgaagt gccggggcag 1200 gatctcctgt catctcacct tgctcctgcc gagaaagtat ccatcatggc tgatgcaatg 1260 cggcggctgc atacgcttga tccggctacc tgcccattcg accaccaagc gaaacatcgc 1320 atcgagcgag cacgtactcg gatggaagcc ggtcttgtcg atcaggatga tctggacgaa 1380 gagcatcagg ggctcgcgcc agccgaactg ttcgccaggc tcaaggcgcg catgcccgac 1440 ggcgaggatc tcgtcgtgac ccatggcgat gcctgcttgc cgaatatcat ggtggaaaat 1500 ggccgctttt ctggattcat cgactgtggc cggctgggtg tggcggaccg ctatcaggac 1560 atagcgttgg ctacccgtga tattgctgaa gagcttggcg gcgaatgggc tgaccgcttc 1620 ctcgtgcttt acggtatcgc cgctcccgat tcgcagcgca tcgccttcta tcgccttctt 1680 gacgagttct tctcgggatc cgaaaatctc tacttccaga gtggaagttc tggttcaagt 1740 ggttccggct cggcaagtgg ttccggcatg gtgagcaagg gcgaggcagt gatcaaggag 1800 ttcatgcggt tcaaggtgca catggagggc tccatgaacg gccacgagtt cgagatcgag 1860 ggcgagggcg agggccgccc ctacgagggc acccagaccg ccaagctgaa ggtgaccaag 1920 ggtggcccccc tgcccttctc ctgggacatc ctgtcccctc agttcatgta cggctccagg 1980 gccttcatca agcaccccgc cgacatcccc gactactata agcagtcctt ccccgagggc 2040 ttcaagtggg agcgcgtgat gaacttcgag gacggcggcg ccgtgaccgt gacccaggac 2100 acctccctgg aggacggcac cctgatctac aaggtgaagc tccgaggcac caacttccct 2160 cctgacggcc ccgtaatgca gaagagaca atgggctggg aagcgtccac cgagcggttt 2220 taccccgagg acggcgtgct gaagggcgac attaagatgg ccctgcgcct gaaggacgga 2280 ggccgctacc tggcggactt caagaccacc tacaaggcca agaagcccgt gcagatgccc 2340 ggcgcctaca acgtcgaccg caagttggac atcacctccc aacaggag ctacaccgtg 2400 gtggaacagt acgaacgctc cgagggccgc cactccaccg gcggcatgga cgagctatac 2460 aagtccagct cgggctccag tggagggtca ggctcgtcta gtagctccgg agggtcaagc 2520 ggtaccggaa tggaccaggt gcagctggtg gagtccggag gcgcactggt gcagccaggc 2580 ggttccctgc gactgtcttg cgcagcttcc ggattcccag taaaccgta ctccatgcga 2640 tggtatcgcc aggcacccgg taggaacgc gagtgggtgg caggcatgtc atccgcaggc 2700 gaccgctcct cctacgagga ctccgtgaag ggccgcttca ccatctcccg cgacgacgcc 2760 cgcaacaccg tgtacctgca gatgaactcc ctgaagcccg aggacaccgc cgtgtactac 2820 tgcaacgtga acgtgggctt cgagtactgg ggccagggca cccaggtgac cgtgtcctcc 2880 tagtctagat aggggcccgt ttaaacccgc tgatcagcct cgactgtgcc ttctagttgc 2940 cagccatctg ttgtttgccc ctccccccgtg ccttccttga ccctggaagg tgccactccc 3000 actgtccttt cctaataaaa tgaggaaatt gcatcgcatt gtctgagtag gtgtcattct 3060 attctggggg gtggggtggg gcaggacagc aagggggagg attgggaaga caatagcagg 3120 catgctgggg atgcggtggg ctctatggct tctgaggcgg aaaaaccag ctggggctct 3180 aggggtatc cccacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg 3240 cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct 3300 tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta 3360 gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt 3420 tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg 3480 ttctttaata gtggactctt gttccaaact ggaacaacac tcaaccctat ctcggtctat 3540 tcttttgatt tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt 3600 taacaaaaat ttaacgcgaa ttaattctgt ggaatgtgtg tcagttaggg tgtggaaagt 3660 ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca 3720 ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt 3780 agtcagcaac catagtcccg cccctaactc cgcccatccc gcccctaact ccgcccagtt 3840 ccgcccattc tccgccccat ggctgactaa ttttttttat ttatgcagag gccgaggccg 3900 cctctgcctc tgagctattc cagaagtagt gaggaggctt ttttggaggc ctaggctttt 3960 gcaaaaagct cccgggagct tgtatatcca ttttcggatc tgatcaagag acaggatgag 4020 gatcgtttcg catgattgaa caagatggat tgcacgcagg ttctccggcc gcttgggtgg 4080 agaggctatt cggctatgac tgggcacaac agacaatcgg ctgctctgat gccgccgtgt 4140 tccggctgtc agcgcagggg cgcccggttc tttttgtcaa gaccgacctg tccggtgccc 4200 tgaatgaact gcaggacgag gcagcgcggc tatcgtggct ggccacgacg ggcgttcctt 4260 gcgcagctgt gctcgacgtt gtcactgaag cgggaaggga ctggctgcta ttgggcgaag 4320 tgccggggca ggatctcctg tcatctcacc ttgctcctgc cgagaaagta tccatcatgg 4380 ctgatgcaat gcggcggctg catacgcttg atccggctac ctgcccattc gaccaccaag 4440 cgaaacatcg catcgagcga gcacgtactc ggatggaagc cggtcttgtc gatcaggatg 4500 atctggacga agagcatcag gggctcgcgc cagccgaact gttcgccagg ctcaaggcgc 4560 gcatgcccga cggcgaggat ctcgtcgtga cccatggcga tgcctgcttg ccgaatatca 4620 tggtggaaaa tggccgcttt tctggattca tcgactgtgg ccggctgggt gtggcggacc 4680 gctatcagga catagcgttg gctacccgtg atattgctga agagcttggc ggcgaatggg 4740 ctgaccgctt cctcgtgctt tacggtatcg ccgctcccga ttcgcagcgc atcgccttct 4800 atcgccttct tgacgagttc ttctgagcgg gactctgggg ttcgaaatga ccgaccaagc 4860 gacgcccaac ctgccatcac gagatttcga ttccaccgcc gccttctatg aaaggttggg 4920 cttcggaatc gttttccggg acgccggctg gatgatcctc cagcgcgggg atctcatgct 4980 ggagttcttc gcccacccca acttgtttat tgcagcttat aatggttaca aataaagcaa 5040 tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc 5100 caaactcatc aatgtatctt atcatgtctg tataccgtcg acctctagct agagcttggc 5160 gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa 5220 catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac 5280 attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca 5340 ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc 5400 ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc 5460 aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc 5520 aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag 5580 gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc 5640 gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt 5700 tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct 5760 ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg 5820 ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct 5880 tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat 5940 tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg 6000 ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa 6060 aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt 6120 ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc 6180 tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt 6240 atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta 6300 aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat 6360 ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac 6420 tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg 6480 ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag 6540 tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt 6600 aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt 6660 gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt 6720 tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt 6780 cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct 6840 tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt 6900 ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac 6960 cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa 7020 actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa 7080 ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca 7140 aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct 7200 ttttcaatat tattgaagca tttatcaggg ttatgtctc atgagcggat acatatttga 7260 atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc 7320 tgacgtc 7327 <210> 5 <211> 6294 <212> DNA <213> artificial sequence <400> 5 gacggatcgg gagatctccc gatcccctat ggtgcactct footcaatc tgctctgatg 60 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg 120 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc 180 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt 240 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata 300 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc 360 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc 420 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt 480 atcatatgcc aagtacgcccc cctattgacg tcaatgacgg taaatggccc gcctggcatt 540 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca 600 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg 660 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc 720 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg 780 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca 840 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagaccacaa gctggctagc 900 ctagcatggt gagcaagggc gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc 960 tggacggcga cgtaaacggc cacaagttca gcgtgtccgg cgagggcgag ggcgatgcca 1020 cctacggcaa gctgaccctg aagttcatct gcaccaccgg caagctgccc gtgccctggc 1080 ccaccctcgt gaccaccctg acctacggcg tgcagtgctt cagccgctac cccgaccaca 1140 tgaagcagca cgacttcttc aagtccgcca tgcccgaagg ctacgtccag gagcgcacca 1200 tcttcttcaa ggacgacggc aactacaaga cccgcgccga ggtgaagttc gagggcgaca 1260 ccctggtgaa ccgcatcgag ctgaagggca tcgacttcaa ggaggacggc aacatcctgg 1320 ggcacaagct ggagtacaac tacaacagcc acaacgtcta tatcatggcc gacaagcaga 1380 agaacggcat caaggtgaac ttcaagatcc gccacaacat cgaggacggc agcgtgcagc 1440 tcgccgacca ctaccagcag aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca 1500 accactacct gagcacccag tccgccctga gcaaagaccc caacgagaag cgcgatcaca 1560 tggtcctgct ggagttcgtg accgccgccg ggatcactct cggcatggac gagctataca 1620 agtccagctc gggctccagt ggagggtcag gctcgtctag tagctccgga gggtcaagcg 1680 gtaccggaat ggaaccgcag aacaaccagg cgaaagaact ggaacgcctg attgataaag 1740 cgatgaaaaa agatggcctg gtgggcatgg cgattgtggg cggcatggcg ctgggcgtgg 1800 cgggcctggc gggcctgatt ggcctggcgg tgagcaaaag caaaagctag tctagatagg 1860 ggcccgttta aacccgctga tcagcctcga ctgtgccttc tagttgccag ccatctgttg 1920 tttgcccctc ccccgtgcct tccttgaccc tggaaggtgc cactcccact gtcctttcct 1980 aataaaatga ggaaattgca tcgcattgtc tgagtaggtg tcattctatt ctggggggtg 2040 gggtggggca ggacagcaag ggggaggatt gggaagacaa tagcaggcat gctggggatg 2100 cggtgggctc tatggcttct gaggcggaaa gaaccagctg gggctctagg gggtatcccc 2160 acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg 2220 ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca 2280 cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta 2340 gtgctttacg gcacctcgac cccaaaaaac ttgattaggg tgatggttca cgtagtgggc 2400 catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg 2460 gactcttgtt ccaaactgga acaacactca accctatctc ggtctattct tttgatttat 2520 aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa caaaaattta 2580 acgcgaatta attctgtgga atgtgtgtca gttagggtgt ggaaagtccc caggctcccc 2640 agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccaggt gtggaaagtc 2700 cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccat 2760 agtcccgccc ctaactccgc ccatcccgcc cctaactccg cccagttccg cccattctcc 2820 gccccatggc tgactaattt tttttattta tgcagaggcc gaggccgcct ctgcctctga 2880 gctattccag aagtagtgag gaggcttttt tggaggccta ggcttttgca aaaagctccc 2940 gggagcttgt atatccattt tcggatctga tcaagagaca ggatgaggat cgtttcgcat 3000 gattgaacaa gatggattgc acgcaggttc tccggccgct tgggtggaga ggctattcgg 3060 ctatgactgg gcacaacaga caatcggctg ctctgatgcc gccgtgttcc ggctgtcagc 3120 gcaggggcgc ccggttcttt ttgtcaagac cgacctgtcc ggtgccctga atgaactgca 3180 ggacgaggca gcgcggctat cgtggctggc cacgacgggc gttccttgcg cagctgtgct 3240 cgacgttgtc actgaagcgg gaagggactg gctgctattg ggcgaagtgc cggggcagga 3300 tctcctgtca tctcaccttg ctcctgccga gaaagtatcc atcatggctg atgcaatgcg 3360 gcggctgcat acgcttgatc cggctacctg cccattcgac caccaagcga aacatcgcat 3420 cgagcgagca cgtactcgga tggaagccgg tcttgtcgat caggatgatc tggacgaaga 3480 gcatcagggg ctcgcgccag ccgaactgtt cgccaggctc aaggcgcgca tgcccgacgg 3540 cgaggatctc gtcgtgaccc atggcgatgc ctgcttgccg aatatcatgg tggaaaatgg 3600 ccgcttttct ggattcatcg actgtggccg gctgggtgtg gcggaccgct atcaggacat 3660 agcgttggct acccgtgata ttgctgaaga gcttggcggc gaatgggctg accgcttcct 3720 cgtgctttac ggtatcgccg ctcccgattc gcagcgcatc gccttctatc gccttcttga 3780 cgagttcttc tgagcgggac tctggggttc gaaatgaccg accaagcgac gcccaacctg 3840 ccatcacgag atttcgattc caccgccgcc ttctatgaaa ggttgggctt cggaatcgtt 3900 ttccgggacg ccggctggat gatcctccag cgcggggatc tcatgctgga gttcttcgcc 3960 caccccaact tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat 4020 ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat 4080 gtatcttatc atgtctgtat accgtcgacc tctagctaga gcttggcgta atcatggtca 4140 tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat acgagccgga 4200 agcataaagt gtaaagcctg gggtgcctaa tgagtgagct aactcacatt aattgcgttg 4260 cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc 4320 caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc gctcactgac 4380 tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata 4440 cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa 4500 aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct 4560 gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa 4620 agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg 4680 cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca 4740 cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa 4800 ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg 4860 gtaagacacg acttatcgcc actggcagca gccactggta acaggattag cagagcgagg 4920 tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga 4980 acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc 5040 tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag 5100 attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac 5160 gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc 5220 ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag 5280 taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt 5340 ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag 5400 ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca 5460 gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact 5520 ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca 5580 gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg 5640 tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatcccc 5700 atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg 5760 gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca 5820 tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt 5880 5940 agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc 6000 ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca 6060 tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa 6120 aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat 6180 tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa 6240 aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctga cgtc 6294 <210> 6 <211> 7795 <212> DNA <213> Artificial sequence <400> 6 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg 60 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg 120 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc 180 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt 240 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata 300 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc 360 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc 420 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt 480 atcatatgcc aagtacgcccc cctattgacg tcaatgacgg taaatggccc gcctggcatt 540 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca 600 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg 660 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc 720 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg 780 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca 840 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagaccacaa gctggctagc 900 atgtccgtcc tgacgccgct gctgctgcgg ggcttgacag gctcggcccg gcggctccca 960 gtgccgcgcg ccagatcca ttcgttgccg ccggagcg aatcagctc cggaagttct 1020 ggttcaatgg ctgacagccg agatcccgcc agcgaccaga tgcagcactg 1080 cggggccgcgc agaagctga tgtcctgacc actggagctg gtaacccagt aggagacaaa 1140 cttaatgtta ttacagtagg gccccgtggg ccccttctg tcaggatgt ggtttcact 1200 gatgaaatgg ctcattttga ccgagagagg attcctgaga gagttgtgca tgctaagga 1260 gcaggggcct ttggctactt tgaggtcaca catgacatta ccaatactc caggcaag 1320 gtatttgagc atattggaaa gaagactccc atcgcagttc gttctccac tgttgctgga 1380 gatcggggtt cagctgacac agttcgggac cctcgtggtt ttgcagtgaa attttacaca 1440 gaagagatgta actgggatct cgttggaat aacaccccca ttttctcat cagagatccc 1500 atattgtttc catctttt ccacagccaa agagaaatc ctcagacaca tctgaagat 1560 ccggacatgg tctgggactt ctggagccta cgtcctgagt ctctgcatca ggttctttc 1620 ttgttcagtg atcgggggat tccagatgga catcgccaca tgaatgata tggatcacat 1680 actttcaagc tggttaatgc aaatggggag gcagtttatt gcaaattcca ttataagact 1740 gaccagggca tcaaaaacct ttctgttgaa gatgcggcga gactttccca ggaagatcct 1800 gactatggca tccgggatct ttttaacgcc attgccacag gaaagtaccc ctcctggact 1860 ttttacatcc aggtcatgac atttaatcag gcagaaactt ttccatttaa tccattcgat 1920 ctcaccaagg tttggcctca caaggactac cctctcatcc cagttggtaa actggtctta 1980 aaccggaatc cagttaatta ctttgctgag gttgaacaga tagccttcga cccaagcaac 2040 atgccacctg gcattgaggc cagtcctgac aaaatgcttc agggccgcct ttttgcctat 2100 cctgacactc accgccatcg cctgggaccc aattatcttc atatacctgt gaactgtccc 2160 taccgtgcac gagtggccaa ctaccagcgt gacggcccga tgtgcatgca ggacaatcag 2220 ggtggtgctc caaattacta ccccaacagc tttggtgctc cggaacaaca gccttctgcc 2280 ctggagcaca gcatccaata ttctggagaa gtgcggagat tcaacactgc caatgatgat 2340 aacgttactc aggtgcgggc attctatgtg aacgtgctga atgaggaaca gaggaaacgt 2400 ctgtgtgaga acattgccgg ccacctgaag gatgcacaaa ttttcatcca gaagaagcg 2460 gtcaagaact tcactgaggt ccaccctgac tacgggagcc acatccaggc tcttctggac 2520 aagtacaatg ctgagaagcc taagaatgcg attcacacct ttgtgcagtc cggatctcac 2580 ttggcggcaa gggagaaggc aaatctgtga agtggttccg gctcggcaag tggttccggc 2640 ggtagtggcg gccgcatggt gagcaagggc gaggcagtga tcaaggagtt catgcggttc 2700 aaggtgcaca tggagggctc catgaacggc cacgagttcg agatcgaggg cgagggcgag 2760 ggccgcccct acgagggcac ccagaccgcc aagctgaagg tgaccaaggg tggccccctg 2820 cccttctcct gggacatcct gtcccctcag ttcatgtacg gctccagggc cttcatcaag 2880 caccccgccg acatccccga ctactataag cagtccttcc ccgagggctt caagtgggag 2940 cgcgtgatga acttcgagga cggcggcgcc gtgaccgtga cccaggacac ctccctggag 3000 gacggcaccc tgatctacaa ggtgaagctc cgaggcacca acttccctcc tgacggcccc 3060 gtaatgcaga agaagacaat gggctgggaa gcgtccaccg agcggttgta ccccgaggac 3120 ggcgtgctga agggcgacat taagatggcc ctgcgcctga aggacggagg ccgctacctg 3180 gcggacttca agaccaccta caaggccaag aagcccgtgc agatgcccgg cgcctacaac 3240 gtcgaccgca agttggacat cacctcccac aacgaggact acaccgtggt ggaacagtac 3300 gaacgctccg agggccgcca ctccaccggc ggcatggacg agctatacaa gtagtctaga 3360 gggcccgttt aaacccgctg atcagcctcg actgtgcctt ctagttgcca gccatctgtt 3420 gtttgcccct cccccgtgcc ttccttgacc ctggaaggtg ccactcccac tgtcctttcc 3480 taataaaatg aggaaattgc atcgcattgt ctgagtaggt gtcattctat tctggggggt 3540 ggggtggggc aggacagcaa gggggaggat tgggaagaca atagcaggca tgctgggggat 3600 gcggtgggct ctatggcttc tgaggcggaa agaaccagct ggggctctag ggggtatccc 3660 cacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc 3720 gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc ctttctcgcc 3780 acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt 3840 agtgctttac ggcacctcga ccccaaaaaa cttgattagg gtgatggttc acgtagtggg 3900. ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt ctttaatagt 3960. ggactcttgt tccaaactgg aacaacactc aaccctatct cggtctattc ttttgattta 4020. tagggattt tgccgatttc ggcctattgg ttaaaaaatg agctgattt acaaaaattt aacgcgaatt aattctgtgg aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc 4140. cagcaggcag aagtatgcaa to catchcatc tcaattagtc to catch tgtggaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca 4320. 4320. 4320. 4320. 4320. tagtcccgcc cgccccatgg ctgactaatt ttttttattt atgcagaggc cgaggccgcc tctgcctctg 4380. agctattcca gaagtagtga ggaggctttt ttggaggcct aggcttttgc aaaaagctcc 4440 cgggagcttg tatatccatt ttcggatctg atcaagagac aggatgagga tcgtttcgca tgattgaaca agatggattg cacgcaggtt ctccggccgc ttgggtggag aggctattcg 4560. gctatgactg ggcacaacag acaatcggct gctctgatgc cgccgtgttc cggctgtcag 4620 cgcaggggcg cccggttctt tttgtcaaga ccgacctgtc cggtgccctg aatgaactgc 4680 aggacgaggc agcgcggcta tcgtggctgg ccacgacggg cgttccttgc gcagctgtgc 4740 tcgacgttgt cactgaagcg ggaagggact ggctgctatt gggcgaagtg ccggggcagg 4800 atctcctgtc atctcacctt gctcctgccg agaaagtatc catcatggct gatgcaatgc 4860 ggcggctgca tacgcttgat ccggctacct gcccattcga ccaccaagcg aaacatcgca 4920 tcgagcgagc acgtactcgg atggaagccg gtcttgtcga tcaggatgat ctggacgaag 4980 agcatcaggg gctcgcgcca gccgaactgt tcgccaggct caaggcgcgc atgcccgacg 5040 gcgaggatct cgtcgtgacc catggcgatg cctgcttgcc gaatatcatg gtggaaaatg 5100 gccgcttttc tggattcatc gactgtggcc ggctgggtgt ggcggaccgc tatcaggaca 5160 tagcgttggc tacccgtgat attgctgaag agcttggcgg cgaatgggct gaccgcttcc 5220 tcgtgcttta cggtatcgcc gctcccgatt cgcagcgcat cgccttctat cgccttcttg 5280 acgagttctt ctgagcggga ctctggggtt cgaaatgacc gaccaagcga cgcccaacct 5340 gccatcacga gatttcgatt ccaccgccgc cttctatgaa aggttgggct tcggaatcgt 5400 tttccgggac gccggctgga tgatcctcca gcgcggggat ctcatgctgg agttcttcgc 5460 ccaccccaac ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa 5520 tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa 5580 tgtatcttat catgtctgta taccgtcgac ctctagctag agcttggcgt aatcatggtc 5640 atagctgttt cctgtgtgaa attgttatcc gctcacaatt ccacacaaca tacgagccgg 5700 aagcataaag tgtaaagcct ggggtgccta atgagtgagc taactcacat taattgcgtt 5760 gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg 5820 ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga 5880 ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat 5940 acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca 6000 aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc 6060 tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata 6120 aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc 6180 gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc 6240 acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga 6300 accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc 6360 ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag 6420 gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag 6480 aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag 6540 ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca 6600 gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga 6660 cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat 6720 cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga 6780 gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg 6840 tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga 6900 gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc 6960 agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac 7020 tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc 7080 agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc 7140 gttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc 7200 catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt 7260 ggccgcagtg tttcactca tggttatggc agcactgcat aattctctta ctgtcatgcc 7320 atccgtaaga tgcttttg tgactggtga gtactcaacc aagtcattct gagaatagtg 7380 tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag 7440 cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat 7500 cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc 7560 atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa 7620 aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta 7680 ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa 7740 aaataaaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg acgtc 7795 <210> 7 <211> 6873 <212> DNA <213> artificial sequence <400> 7 gacggatcgg gagatctccc gatcccctat ggtgcactct footcaatc tgctctgatg 60 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg 120 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc 180 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt 240 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata 300 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc 360 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc 420 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt 480 atcatatgcc aagtacgcccc cctattgacg tcaatgacgg taaatggccc gcctggcatt 540 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca 600 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg 660 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc 720 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg 780 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca 840 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagaccacaa gctggctagc 900 ggatccatgc ctggatcagc actgctatgc tgcctgctct tactgactgg catgaggatc 960 agcaggggcc agtacagccg ggaagacaat aactgcaccc acttcccagt cggccagagc 1020 cacatgctcc tagagctgcg gactgcctt agccaggtga agactttctt tcaaacaaag 1080 gaccagctgg acaacatact gctaaccgac tccttaatgc aggactttaa gggttacttg 1140 ggttgccaag ccttatcgga aatgatccag ttttacctgg tagaagtgat gccccaggca 1200 gagaagcatg gcccagaaat caaggagcat ttgaactccc tgggtgagaa gctgaagacc 1260 ctcaggatgc ggctgaggcg ctgtcatcga tttctcccct gtgaaaataa gagcaaggca 1320 gtggagcagg tgaagagtga ttttaataag ctccaagacc aaggtgtcta caaggccatg 1380 aatgaatttg acatcttcat caactgcata gaagcataca tgatgatcaa aatgaaaagc 1440 tcggaattcg gaagttctgg ttcaagtggt tccggctcgg caagtggttc cggcatggtg 1500 agcaagggcg aggagagataa catggcctct ctcccagcga cacatgagtt acacatcttt 1560 ggctccatca acggtgtgga ctttgacatg gtgggtcagg gcaccggcaa tccaaatgat 1620 ggttatgagg agttaaaacct gaagtccacc aagggtgacc tccagttctc cccctggatt 1680 ctggtccctc atatcgggta tggcttccat cagtacctgc cctaccctga cgggatgtcg 1740 cctttccagg ccgccatggt agatggctcc ggataccaag tccatcgcac aatgcagttt 1800 gaagatggtg cctcccttac tgttaactac cgctacacct acgagggaag ccacatcaaa 1860 ggagaggccc aggtgaaggg gactggtttc cctgctgacg gtcctgtgat gaccaactcg 1920 ctgaccgctg cggactggtg caggtcgaag aagacttacc ccaacgacaa aaccatcatc 1980 agtaccttta agtggagtta caccactgga aatggcaagc gctaccggag cactgcgcgg 2040 accacctaca cctttgccaa gccaatggcg gctaactatc tgaagaacca gccgatgtac 2100 gtgttccgta agacggagct caagcactcc aagaccgagc tcaacttcaa ggagtggcaa 2160 aaggccttta ccgatgtgat gggcatggac gagctgtaca agtccagctc gggctccagt 2220 ggagggtcag gctcgtctag tagctccgga gggtcaagcg gtaccggaat ggaaccgcag 2280 aacaaccagg cgaaagaact ggaacgcctg attgataaag cgatgaaaaa agatggcctg 2340 gtgggcatgg cgattgtggg cggcatggcg ctgggcgtgg cgggcctggc gggcctgatt 2400 ggcctggcgg tgagcaaaag caaaagctag gcggccgcgg gcccgtttaa acccgctgat 2460 cagcctcgac tgtgccttct agttgccagc catctgttgt ttgcccctcc cccgtgcctt 2520 ccttgaccct ggaaggtgcc actcccactg tcctttccta ataaaatgag gaaattgcat 2580 cgcattgtct gagtaggtgt cattctattc tggggggtgg ggtggggcag gacagcaagg 2640 gggaggattg ggaagacaat agcaggcatg ctggggatgc ggtgggctct atggcttctg 2700 aggcggaaag aaccagctgg ggctctaggg ggtatcccca cgcgccctgt agcggcgcat 2760 taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc agcgccctag 2820 cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc tttccccgtc 2880 aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg cacctcgacc 2940 ccaaaaaact tgattagggt gatggttcac gtagtgggcc atcgccctga tagacggttt 3000 ttcgcccttt gacgttggag tccacgttct ttaatagtgg actcttgttc caaactggaa 3060 caacactcaa ccctatctcg gtctattctt ttgatttata agggattttg ccgatttcgg 3120 cctattggtt aaaaaatgag ctgatttaac aaaaatttaa cgcgattaa ttctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag 3240 catgcatctc aattagtcag caaccaggtg tggaagtcc ccaggctccc cagcaggcag aagtatgcaa to catch tcaattagtc to cry gtcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc tgcctctgag ctattccaga agtagtgagg 3480 aggctttttt ggaggcctag gcttttgcaa aaagctcccg ggagcttgta tatccatttt 3540 cggatctgat caagagacag gatgaggatc gtttcgcatg attgaacaag atggattgca cgcaggttct ccggccgctt gggtggagag gctattcggc tatgactggg cacaacagac 3660 aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg caggggcgcc cggttctttt 3720 tgtcaagacc gacctgtccg gtgccctgaa tgaactgcag gacgaggcag cgcggctatc 3780 gtggctggcc acgacgggcg ttccttgcgc agctgtgctc gacgttgtca ctgaagcggg 3840. aagggactgg ctgctattgg gcgaagtgcc ggggcaggat ctcctgtcat ctcaccttgc 3900. tcctgccgag aaagtatcca tcatggctga tgcaatgcgg cggctgcata cgcttgatcc 3960 ggctacctgc ccattcgacc accaagcgaa acatcgcatc gagcgagcac gtactcggat ggaagccggt cttgtcgatc aggatgatct ggacgaagag catcaggggc tcgcgccagc 4080. cgaactgttc gccaggctca aggcgcgcat gcccgacggc gaggatctcg tcgtgaccca 4140 tggcgatgcc tgcttgccga atatcatggt ggaaaatggc cgcttttctg gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata gcgttggcta cccgtgatat 4260 tgctgaagag cttggcggcg aatgggctga ccgcttcctc gtgctttacg gtatcgccgc 4320 tcccgattcg cagcgcatcg ccttctatcg ccttcttcg gagttcttct gagcgggact 4380. ctggggttcg aaatgaccga ccaagcgacg cccaacctgc catcacgaga tttcgattcc accgccgcct tctatgaaag gttggggcttc ggaatcgttt tccgggacgc cggctggatg atcctccagc gcggggatct catgctggag ttcttcgccc accccaactt gtttattgca 4560. gcttataatg gttacaaata aagcaatagc atcacaaatt tcacaaataa agcatttttt 4620 tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca tgtctgtata 4680 ccgtcgacct ctagctagag cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat 4740 tgttatccgc tcacaattcc acacaacata cgagccggaa gcataaagtg taaagcctgg 4800 ggtgcctaat gagtgagcta actcacatta attgcgttgc gctcactgcc cgctttccag 4860 tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt 4920 ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg 4980 ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg 5040 gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag 5100 gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga 5160 cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct 5220 ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc 5280 tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg 5340 gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc 5400 tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca 5460 ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag 5520 ttcttgaagt ggtggcctaa ctacggctac actagaagaa cagtatttgg tatctgcgct 5580 ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc 5640 accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga 5700 tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca 5760 cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat 5820 taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac 5880 caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt 5940 gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt 6000 gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc aaaaaccag 6060 ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgccctc catccagtct 6120 attaattgtt gccgggaagc taagtaagt agttcgccag ttaatagttt gcgcaacgtt 6180 gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc 6240 tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaagcggtt 6300 agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg 6360 gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg 6420 actggtgagt actcaaccaa gtcattctga gatagtgta tgcggcgacc gagttgctt 6480 tgcccggcgt caatacggga taataccggcg ccacatagca gaactttaaa agtgctcatc 6540 attggaaaac gttcttcggg gcgaaaactc tcaggatct taccgctgtt gagatccagt 6600 tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttactt caccagcgtt 6660 tctgggtgag siaaaacagg aaggcaaat gccgcaaaa agggaataag ggcgacacgg 6720 aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta tcagggttat 6780 tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg 6840 cgcacatttc cccgaaaagt gccacctgac gtc 6873 <210> 8 <211> 7620 <212> DNA <213> Artificial synthesis (artificial sequence) <400> 8 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg 60 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg 120 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc 180 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt 240 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata 300 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc 360 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc 420 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt 480 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt 540 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg 660 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc 720 780. aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca 840 ctgcttactg gcttatcga attack ctcactatag ggagacccaa gctggctagc ctagcatgag cagcgtgctg gcgagctgcc cgaaaaaacc ggtgagcagc tatctgcgct ttagcaaaga acagctgccg atttttaaag cgcagaaccc ggatgcgaaa accaccgaac tgattcgccg cattgcgcag cgctggcgcg aactgccgga tagcaaaaaa aaaatttatc aggatgcgta tcgcgcgga tggcaggtgt ataaagaaga aattagccgc tttaaagaac agctgacccc gagccagatt atgagcctgg aaaaagaat tatggataa catctgaaac gcaaagcgat gaccaaaaaa aaagaactga ccctgctggg caaaccgaaa cgcccgcgca gcgcgtata cgtgtatgtg gcggaacgct ttcaggaagc gaaaggcgat agcccgcagg aaaaactgaa aaccgtgaaa gaaactgga aaaacctgag cgatagcgaa aaagaactgt atattcagca tgcgaaaga gatgaaaccc gctatcataa cgaaatgaa agctgggaag 1500. 1500. 1500. 1500. 1500. 1500. 1500. 1500. 1500. 1500. 1500 aatatggcgc ggaagaatgc tcgggatcct ccagctcggg cgggatggcg aaaaaagtga aaaaacgcaa cccgtggagc gatgatgaa gcaaaagcga aagcgatctg gagaaaccg aaccggtggt gattccgcgc gatagcctgc tgcgccgcgc ggcggcggaa cgcccgaaat 1680. atacctttga ttttagcgaa gaagaagatg atgatgcgga tgatgatgat gatgataaca acgatctgga agaactgaaa gtgaaagcga gcccgattac caacgatggc gaagatgaat ttgtgccgag cgatggcctg gataaagatg aatatacctt tagcccgggc aaaagcaaag cgaccccgga aaaaagcctg catgataaaa aaagccagga ttttggcaac ctgtttagct ttccgagcta tagccagaaa agcgaagatg atagcgcgaa atttgatagc aacgaagaag 1980 atagcgcgag cgtgtttagc ccgagctttg gcctgaaaca gaccgataaa gtgccgagca 2040 aaaccgtggc ggcgaaaaaa ggcaaaccga gcagcgatac cgtgccgaaa ccgaaacgcg 2100 cgccgaaaca gaaaaaagtg gtggaagcgg tgaacagcga tagcgatagc gaatttggca 2160 ttccgaaaaa aaccaccacc ggatccgaaa atctctactt ccagagtgga agttctggtt 2220 caagtggttc cggctcggca agtggttccg gcatggtgag caagggcgag gcagtgatca 2280 aggagttcat gcggttcaag gtgcacatgg agggctccat gaacggccac gagttcgaga 2340 tcgagggcga gggcgagggc cgcccctacg agggcaccca gaccgccaag ctgaaggtga 2400 ccaagggtgg ccccctgccc ttctcctggg acatcctgtc ccctcagttc atgtacggct 2460 ccagggcctt catcaagcac cccgccgaca tccccgacta ctataagcag tccttccccg 2520 agggcttcaa gtgggagcgc gtgatgaact tcgaggacgg cggcgccgtg accgtgaccc 2580 aggacacctc cctggaggac ggcaccctga tctacaaggt gaagctccga ggcaccaact 2640 tccctcctga cggccccgta atgcagaaga agacaatggg ctgggaagcg tccaccgagc 2700 ggttgtaccc cgaggacggc gtgctgaagg gcgacattaa gatggccctg cgcctgaagg 2760 acggaggccg ctacctggcg gacttcaaga ccacctacaa ggccaagaag cccgtgcaga 2820 tgcccggcgc ctacaacgtc gaccgcaagt tggacatcac ctcccacaac gaggactaca 2880 ccgtggtgga acagtacgaa cgctccgagg gccgccactc caccggcggc atggacgagc 2940 tatacaagtc cagctcgggc tccagtggag ggtcaggctc gtctagtagc tccggagggt 3000 caagcggtac cggaatggaa ccgcagaaca accaggcgaa agaactggaa cgcctgattg 3060 ataaagcgat gaaaaaagat ggcctggtgg gcatggcgat tgtgggcggc atggcgctgg 3120 gcgtggcggg cctggcgggc ctgattggcc tggcggtgag caaaagcaaa agctagtcta 3180 gataggggcc cgtttaaacc cgctgatcag cctcgactgt gccttctagt tgccagccat 3240 ctgttgtttg cccctcccc gtgccttcct tgaccctgga aggtgccact cccactgtcc 3300 tttcctaata aaatgaggaa attgcatcgc attgtctgag taggtgtcat tctattctgg 3360 ggggtggggt ggggcaggac agcaaggggg aggattggga agacaatagc aggcatgctg 3420 gggatgcggt gggctctatg gcttctgagg cggaaagaac cagctggggc tctagggggt 3480 atccccacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg 3540 tgaccgctac acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc 3600 tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc 3660 gatttagtgc tttacggcac ctcgacccca aaaaacttga ttagggtgat ggttcacgta 3720 gtgggccatc gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta 3780 atagtggact cttgttccaa actggaacaa cactcaaccc tatctcggtc tattcttttg 3840 atttataagg gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa 3900 aatttaacgc gaattaattc tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg 3960 ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg 4020 aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc 4080 aaccatagtc ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca 4140 ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg ccgcctctgc 4200 ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctaggct tttgcaaaaa 4260 gctcccggga gcttgtatat ccattttcgg atctgatcaa gagacaggat gaggatcgtt 4320 tcgcatgatt gaacaagatg gattgcacgc aggttctccg gccgcttggg tggagaggct 4380 attcggctat gactgggcac aacagacaat cggctgctct gatgccgccg tgttccggct 4440 gtcagcgcag gggcgcccgg ttctttttgt caagaccgac ctgtccggtg ccctgaatga 4500 actgcaggac gaggcagcgc ggctatcgtg gctggccacg acgggcgttc cttgcgcagc 4560 tgtgctcgac gttgtcactg aagcgggaag ggactggctg ctattgggcg aagtgccggg 4620 gcaggatctc ctgtcatctc accttgctcc tgccgagaaa gtatccatca tggctgatgc 4680 aatgcggcgg ctgcatacgc ttgatccggc tacctgccca ttcgaccacc aagcgaaaca 4740 tcgcatcgag cgagcacgta ctcggatgga agccggtctt gtcgatcagg atgatctgga 4800 cgaagagcat caggggctcg cgccagccga actgttcgcc aggctcaagg cgcgcatgcc 4860 cgacggcgag gatctcgtcg tgacccatgg cgatgcctgc ttgccgaata tcatggtgga 4920 aaatggccgc ttttctggat tcatcgactg tggccggctg ggtgtggcgg accgctatca 4980 ggacatagcg ttggctaccc gtgatattgc tgaagagctt ggcggcgaat gggctgaccg 5040 cttcctcgtg ctttacggta tcgccgctcc cgattcgcag cgcatcgcct tctatcgcct 5100 tcttgacgag ttcttctgag cgggactctg gggttcgaaa tgaccgacca agcgacgccc 5160 aacctgccat cacgagattt cgattccacc gccgccttct atgaaaggtt gggcttcgga 5220 atcgttttcc gggacgccgg ctggatgatc ctccagcgcg gggatctcat gctggagttc 5280 ttcgcccacc ccaacttgtt tattgcagct tataatggtt acaaataaag caatagcatc 5340 acaaatttca caaataaagc atttttttca ctgcattcta gttgtggttt gtccaaactc 5400 atcaatgtat cttatcatgt ctgtataccg tcgacctcta gctagagctt ggcgtaatca 5460 tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca caacatacga 5520 gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact cacattaatt 5580 gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct gcattaatga 5640 atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc 5700 actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg 5760 gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc 5820 cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc 5880 ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga 5940 ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc 6000 ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat 6060 agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg 6120 cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc 6180 aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga 6240 gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact 6300 agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt 6360 ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag 6420 cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg 6480 tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa 6540 aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata 6600 tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg 6660 atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata 6720 cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg 6780 gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct 6840 gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt 6900 tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc 6960 tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga 7020 tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt 7080 aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc 7140 atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa 7200 tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca 7260 catagcagaa cttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaactctca 7320 aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct 7380 tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc 7440 gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa 7500 tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt 7560 tagaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc 7620 <210> 9 <211> 6970 <212> DNA <213> Artificial sequence <400> 9 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg 60 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg 120 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc 180 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt 240 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata 300 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc 360 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc 420 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt 480 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt 540 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca 600 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg 660 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc 720 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg 780 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca 840 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagaccacaa gctggctagc 900 atgtccgtcc tgacgccgct gctgctgcgg ggcttgacag gctcggcccg gcggctccca 960 gtgccgcgcg ccaagatcca ttcgttgccg ccggagagcg aattcagcgg tatgagcaag 1020 ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg agctggacgg cgacgtaaac 1080 ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg ccacctacgg caagctgacc 1140 ctgaagttca tctgcaccac cggcaagctg cccgtgccct ggcccaccct cgtgaccacc 1200 ctgacctacg gcgtgcagtg cttcagccgc taccccgacc acatgaagca gcacgacttc 1260 ttcaagtccg ccatgcccga aggctacgtc caggagcgca ccatcttctt caaggacgacgac 1320 ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg acaccctggt gaaccgcatc 1380 gagctgaagg gcatcgactt caaggaggac ggcaacatcc tggggcacaa gctggagtac 1440 aactacaaca gccacaacgt ctatatcatg gccgacaagc agaagaacgg catcaaggtg 1500 aacttcaaga tccgccacaa catcgaggac ggcagcgtgc agctcgccga ccactaccag 1560 cagaacaccc ccatcggcga cggccccgtg ctgctgcccg acaaccacta cctgagcacc 1620 cagtccgccc tgagcaaaga ccccaacgag aagcgcgatc acatggtcct gctggagttc 1680 gtgaccgccg ccgggatcac tcacggcatg gacgagctgt acaaggcgac caactttagc 1740 ctgctgaaac aggcgggcga tgtggaagaa aacccgggcc cgaagaagaa gaggaaggtg 1800 agtggcggcc gcatgagcaa gggcgaggag ctgttcaccg gggtggtgcc catcctggtc 1860 gagctggacg gcgacgtaaa cggccacaag ttcagcgtgt ccggcgaggg cgagggcgat 1920 gccacctacg gcaagctgac cctgaagttc atctgcacca ccggcaagct gcccgtgccc 1980 tggcccaccc tcgtgaccac cctgacctac ggcgtgcagt gcttcagccg ctaccccgac 2040 cacatgaagc agcacgactt cttcaagtcc gccatgcccg aaggctacgt ccaggagcgc 2100 accatcttct tcaaggacga cggcaactac aagacccgcg ccgaggtgaa gttcgagggc 2160 gacaccctgg tgaaccgcat cgagctgaag ggcatcgact tcaaggagga cggcaacatc 2220 ctggggcaca agctggagta caactacaac agccacaacg tctatatcat ggccgacaag 2280 cagaagaacg gcatcaaggt gaacttcaag atccgccaca acatcgagga cggcagcgtg 2340 cagctcgccg accactacca gcagaacacc cccatcggcg acggccccgt gctgctgccc 2400 gacaaccact acctgagcac ccagtccgcc ctgagcaaag accccaacga gaagcgcgat 2460 cacatggtcc tgctggagtt cgtgaccgcc gccgggatca ctcacggcat ggacgagctg 2520 tacaagtaat ctagagggcc cgtttaaacc cgctgatcag cctcgactgt gccttctagt 2580 tgccagccat ctgttgtttg cccctccccc gtgccttcct tgaccctgga aggtgccact 2640 cccactgtcc tttcctaata aaatgaggaa attgcatcgc attgtctgag taggtgtcat 2700 tctattctgg ggggtggggt ggggcaggac agcaaggggg aggattggga agacaatagc 2760 aggcatgctg gggatgcggt gggctctatg gcttctgagg cggaaagaac cagctggggc 2820 tctagggggt atccccacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt 2880 acgcgcagcg tgaccgctac acttgccagc gccctagcgc ccgctccttt cgctttcttc 2940 ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct 3000 ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga ttagggtgat 3060 ggttcacgta gtgggccatc gccctgatag acggtttttc gccctttgac gttggagtcc 3120 acgttcttta atagtggact cttgttccaa actggaacaa cactcaaccc tatctcggtc 3180 tattcttttg atttataagg gattttgccg atttcggcct attggttaaa aaatgagctg 3240 atttaacaaa aatttaacgc gaattaattc tgtggaatgt gtgtcagtta gggtgtggaa 3300 agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa 3360 ccaggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca 3420 attagtcagc aaccatagtc ccgcccctaa ctccgcccat cccgccccta actccgccca 3480 gttccgccca ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg 3540 ccgcctctgc ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctaggct 3600 tttgcaaaaa gctcccggga gcttgtatat ccattttcgg atctgatcaa gagacaggat 3660 gaggatcgtt tcgcatgatt gaacaagatg gattgcacgc aggttctccg gccgcttggg 3720 tggagaggct attcggctat gactgggcac aacagacaat cggctgctct gatgccgccg 3780 tgttccggct gtcagcgcag gggcgcccgg ttctttttgt caagaccgac ctgtccggtg 3840 ccctgaatga actgcaggac gaggcagcgc ggctatcgtg gctggccacg acgggcgttc 3900 cttgcgcagc tgtgctcgac gttgtcactg aagcgggaag ggactggctg ctattgggcg 3960 aagtgccggg gcaggatctc ctgtcatctc accttgctcc tgccgagaaa gtatccatca 4020 tggctgatgc aatgcggcgg ctgcatacgc ttgatccggc tacctgccca ttcgaccacc 4080 aagcgaaaca tcgcatcgag cgagcacgta ctcggatgga agccggtctt gtcgatcagg 4140 atgatctgga cgaagagcat caggggctcg cgccagccga actgttcgcc aggctcaagg 4200 cgcgcatgcc cgacggcgag gatctcgtcg tgacccatgg cgatgcctgc ttgccgaata 4260 tcatggtgga aaatggccgc ttttctggat tcatcgactg tggccggctg ggtgtggcgg 4320 accgctatca ggacatagcg ttggctaccc gtgatattgc tgaagagctt ggcggcgaat 4380 gggctgaccg cttcctcgtg ctttacggta tcgccgctcc cgattcgcag cgcatcgcct 4440 tctatcgcct tcttgacgag ttcttctgag cgggactctg gggttcgaaa tgaccgacca 4500 agcgacgccc aacctgccat cacgagattt cgattccacc gccgccttct atgaaaggtt 4560 gggcttcgga atcgttttcc gggacgccgg ctggatgatc ctccagcgcg gggatctcat 4620 gctggagttc ttcgcccacc ccaacttgtt tattgcagct tataatggtt acaaataaag 4680 caatagcatc acaaatttca caaataaagc atttttttca ctgcattcta gttgtggttt 4740 gtccaaactc atcaatgtat cttatcatgt ctgtataccg tcgacctcta gctagagctt 4800 ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca 4860 caacatacga gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact 4920 cacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct 4980 gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc 5040 ttcctcgctc actgactcgc tgcgctcggt cgtcggctg cggcgagcgg tatcagctca 5100 ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg 5160 agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca 5220 taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa 5280 cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc 5340 tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc 5400 gctttctcat agctcacgct gtaggtatct cagttcggtg tagtcgttc gctccaagct 5460 gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg 5520 tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag 5580 gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta 5640 cggctacact agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg 5700 aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt 5760 5820 ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag 5880 attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat 5940 ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc 6000 tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat 6060 aactacgata cgggaggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc 6120 acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag 6180 aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag 6240 agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt 6300 ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg 6360 agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt 6420 tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc 6480 tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc 6540 attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa 6600 taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg 6660 aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc 6720 caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag 6780 gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt 6840 cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt 6900 tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc 6960 acctgacgtc 6970 <210> 10 <211> 6505 <212> DNA <213> Artificial sequence <400> 10 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg 60 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg 120 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc 180 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt 240 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata 300 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc 360 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc 420 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt 480 atcatatgcc aagtacgcccc cctattgacg tcaatgacgg taaatggccc gcctggcatt 540 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca 600 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg 660 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc 720 aaaatcaacg ggactttcca aaatgcgta acaactccgc cccattgacg caaatgggcg 780 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca 840 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagaccacaa gctggctagc 900 atgtctaagt cagagtctcc taaagagccc gaacagctga ggaagctctt cattggaggg 960 1020 acggactgtg tggtaatgag agatccaaac accaagcgct ccaggggctt tgggtttgtc 1080 1140 ggaagagttg tggaaccaaa gagagctgtc tccagagaag attctcaaag accaggtgcc 1200 cacttaactg tgaaaaagat atttgttggt ggcattaaag aagacactga agaacatcac 1260 ctaagagatt attttgaaca gtatggaaaa attgaagtga ttgaaatcat gactgaccga 1320 ggcagtggca agaaaagggg ctttgccttt gtaacctttg acgaccatga ctccgtggat 1380 aagattgtca ttcagaaata ccatactgtg aatggccaca actgtgaagt tagaaaagcc 1440 ctgtcaaagc aagagatggc tagtgcttca tccagccaaa gaggtcgatc gggatccgga 1500 agttctggtt caagtggttc cggctcggca agtggttccg gctccagctc gggctccagt 1560 ggagggtcag gctcgtctag tagctccgga gggtcaagcg gtaccggaca tactctttat 1620 gcccccggtg gatatgacat tatgggatac ctgaggcaga tcaggaaccg gccaaacccg 1680 caggtggaac tgggacccgt ggatacatcc tgcgccttga ttctttgtga cctgaaacag 1740 aaagacaccc cgatagttta cgcgagtgaa gccttcctct acatgacagg ttacagcaac 1800 gcagaggtgc tgggccggaa ttgccggttt ctgcaaagcc ctgacggcat ggtgaagccc 1860 aagagcaccc ggaagtacgt ggatagtaac acaatcaata ctatgcgcaa ggcaatcgac 1920 aggaatgccg aggtgcaggt tgaagtagtc aattttaaaa agaatggaca gcgatttgtt 1980 aatttcctga ctatgatacc tgttagggac gaaacaggcg agtatcgata ctctatggga 2040 ttccagtgcg aaacagaata gtctagatag gggcccgttt aaacccgctg atcagcctcg 2100 actgtgcctt ctagttgcca gccatctgtt gtttgcccct cccccgtgcc ttccttgacc 2160 ctggaaggtg ccactcccac tgtcctttcc taataaaatg aggaaattgc atcgcattgt 2220 ctgagtaggt gtcattctat tctggggggt ggggtggggc aggacagcaa gggggaggat 2280 tgggaagaca atagcaggca tgctggggat gcggtgggct ctatggcttc tgaggcggaa 2340 agaaccagct ggggctctag ggggtatccc cacgcgccct gtagcggcgc attaagcgcg 2400 gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgccct agcgcccgct 2460 cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg tcaagctcta 2520 aatcgggggc tccctttagg gttccgattt agtgctttac ggcacctcga ccccaaaaaa 2580 cttgattagg gtgatggttc acgtagtggg ccatcgccct gatagacggt ttttcgccct 2640 ttgacgttgg agtccacgtt ctttaatagt ggactcttgt tccaaactgg aacaacactc 2700 aaccctatct cggtctattc ttttgattta taagggattt tgccgatttc ggcctattgg 2760 ttaaaaaatg agctgattta acaaaaattt aacgcgaatt aattctgtgg aatgtgtgtc 2820 agttagggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc 2880 tcaattagtc agcaaccagg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc 2940 aaagcatgca tctcaattag tcagcaacca tagtcccgcc cctaactccg cccatcccgc ccctaactcc gcccagttcc gcccattctc cgccccatgg ctgactaatt ttttttattt 3060 atgcagaggc cgaggccgcc tctgcctctg agctattcca gaagtagtga ggaggctttt ttggaggcct aggcttttgc aaaaagctcc cggggagcttg tatatccatt ttcggatctg 3180 atcaagagac rich tcgtttcgca tgattgaaca rich cacgcaggtt ctccggccgc ttgggtggag aggctatttcg gctatgactg ggcacaacag acaatcggct 3300. gctctgatgc cgccgtgttc cggctgtcag cgcaggggcg cccggttctt tttgtcaaga 3360 ccgacctgtc cggtgccctg aatgaactgc aggacgaggc agcgcggcta tcgtggctgg 3420 ccacgacggg cgttccttgc gcagctgtgc tcgacgttgt cactgaagcg ggaagggact 3480. ggctgctatt gggcgaagtg ccggggcagg atctcctgtc atctcacctt gctcctgccg 3540 agaaagtatc catcatggct gatgcaatgc ggcggctgca tacgcttgat ccggctacct gcccattcga ccaccaagcg aaacatcgca tcgagcgagc acgtactcgg atggaagccg gtcttgtcga tcaggatgat ctggacgaag agcatcaggg gctcgcgcca gccgaactgt 3720 tcgccaggct caaggcgcgc atgcccgacg gcgaggatct cgtcgtgacc catggcgatg 3780 cctgcttgcc gaatatcatg gtggaaaatg gccgcttttc tggattcatc gactgtggcc 3840 ggctgggtgt ggcggaccgc tatcaggaca tagcgttggc tacccgtgat attgctgaag 3900 agcttggcgg cgaatgggct gaccgcttcc tcgtgcttta cggtatcgcc gctcccgatt 3960 cgcagcgcat cgccttctat cgccttcttg acgagttctt ctgagcggga ctctggggtt 4020 cgaaatgacc gaccaagcga cgcccaacct gccatcacga gatttcgatt ccaccgccgc 4080 cttctatgaa aggttgggct tcggaatcgt tttccgggac gccggctgga tgatcctcca 4140 gcgcggggat ctcatgctgg agttcttcgc ccaccccaac ttgtttattg cagcttataa 4200 tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt tttcactgca 4260 ttctagttgt ggtttgtcca aactcatcaa tgtatcttat catgtctgta taccgtcgac 4320 ctctagctag agcttggcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc 4380 gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta 4440 atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa 4500 cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat 4560 tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg 4620 agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc 4680 aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt 4740 gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaaatc gacgctcaag 4800 tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc 4860 cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc 4920 ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt 4980 cgttcgctcc aagctggggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt 5040 atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc 5100 agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa 5160 gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa 5220 gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg 5280 tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga 5340 agatccttg atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg 5400 gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg 5460 aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt 5520 aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact 5580 ccccgtcgtg tagataacta cgatacgggga gggcttacca tctggcccca gtgctgcaat 5640 gataccgga gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg 5700 aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg 5760 ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat 5820 tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc 5880 ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta 6240. acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg 6300. agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat 6480. cgcgcacatt cgcgcacatt cgcgcacatt tccccgaaaa gtgccacctg acgtc6505 <210> 11 <211> 6163 <212> DNA <213> Synthetic (artificial sequence) <400> 11 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg 60 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg 120 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc 180 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt 240 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata 300 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc 360 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc 420 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt 480 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt 540 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca 600 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg 660 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc 720 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg 780 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca 840 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagaccacaa gctggctagc 900 atgtccgtcc tgacgccgct gctgctgcgg ggcttgacag gctcggcccg gcggctccca 960 gtgccgcgcg ccaagatcca ttcgttgccg ccggagagcg aattcagcgg tagtggcggc 1020 cgcatggtga gcaagggcga ggcagtgatc aaggagttca tgcggttcaa ggtgcacatg 1080 gagggctcca tgaacggcca cgagttcgag atcgagggcg agggcgaggg ccgcccctac 1140 gagggcaccc agaccgccaa gctgaaggtg accaagggtg gccccctgcc cttctcctgg 1200 gacatcctgt cccctcagtt catgtacggc tccagggcct tcatcaagca ccccgccgac 1260 atccccgact actataagca gtccttcccc gagggcttca agtgggagcg cgtgatgaac 1320 ttcgaggacg gcggcgccgt gaccgtgacc caggacacct ccctggagga cggcaccctg 1380 atctacaagg tgaagctccg aggcaccaac ttccctcctg acggccccgt aatgcagaag 1440 aagacaatgg gctgggaagc gtccaccgag cggttgtacc ccgaggacgg cgtgctgaag 1500 ggcgacatta agatggccct gcgcctgaag gacggaggcc gctacctggc ggacttcaag 1560 accacctaca aggccaagaa gcccgtgcag atgcccggcg cctacaacgt cgaccgcaag 1620 ttggacatca cctcccacaa cgaggactac accgtggtgg aacagtacga acgctccgag 1680 ggccgccact ccaccggcgg catggacgag ctatacaagt agtctagagg gcccgtttaa 1740 acccgctgat cagcctcgac tgtgccttct agttgccagc catctgttgt ttgcccctcc 1800 cccgtgcctt ccttgaccct ggaaggtgcc actcccactg tcctttccta ataaaatgag 1860 gaaattgcat cgcattgtct gagtaggtgt cattctattc tggggggtgg ggtggggcag 1920 gacagcaagg gggaggattg ggaagacaat agcaggcatg ctggggatgc ggtgggctct 1980 atggcttctg aggcggaaag aaccagctgg ggctctaggg ggtatcccca cgcgccctgt 2040 agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc 2100 agcgccctag cgcccgctcc ttcgcttc ttccctcct tctcgccac gttcgccggc 2160 tttccccgtc aagctctaaa tcggggctc cctttaggt tccgatttag tgctttacgg 2220 cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc atcgccctga 2280 tagacggttt ttcgccttt gacgttggag tccacgttct ttaatagtgg actcttgttc 2340 caaactggaa siacactca ccctatctcg gtctattctt ttgatttata agggattg 2400 ccgatttcgg cctattggtt aaaaatgag ctgatttaac aaaaatttaa cgcgaattaa 2460 ttctgtggaa tgtgtgtcag ttaggtgtg gaaagtcccc aggctcccca gcaggcagaa 2520 gtatgcaaag catgcatctc aattagtcag caccaggtg tggaagtcc ccaggctccc 2580 cagcaggcag aagtatgca agcatgcatc tcattagtc agcaccaca gtcccgcccc 2640 taactccgcc catcccgccc ctactccgc ccagttccgc ccattctccg cccatggct 2700 gactaattt ttttatt gcagaggccg aggccgcctc tgcctctgag ctattccaga 2760 agtagtgagg aggcttttt ggaggcctag gctttgca aaagctcccg ggagcttgta 2820 tatccatttt cggatctgat caagagacag gatgaggatc gtttcgcatg attgaacaag 2880 atggattgca cgcaggttct ccggccgctt gggtggagag gctattcggc tatgactggg 2940 cacaacagac aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg caggggcgcc 3000 cggttctttt tgtcaagacc gacctgtccg gtgccctgaa tgaactgcag gacgaggcag 3060 cgcggctatc gtggctggcc acgacgggcg ttccttgcgc agctgtgctc gacgttgtca 3120 ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc ggggcaggat ctcctgtcat 3180 ctcaccttgc tcctgccgag aaagtatcca tcatggctga tgcaatgcgg cggctgcata 3240 cgcttgatcc ggctacctgc ccattcgacc accaagcgaa acatcgcatc gagcgagcac 3300 gtactcggat ggaagccggt cttgtcgatc aggatgatct ggacgaagag catcaggggc 3360 tcgcgccagc cgaactgttc gccaggctca aggcgcgcat gcccgacggc gaggatctcg 3420 tcgtgaccca tggcgatgcc tgcttgccga atatcatggt ggaaaatggc cgcttttctg 3480 gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata gcgttggcta 3540 cccgtgatat tgctgaagag cttggcggcg aatgggctga ccgcttcctc gtgctttacg 3600 gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg ccttcttgac gagttcttct 3660 gagcgggact ctggggttcg aaatgaccga ccaagcgacg cccaacctgc catcacgaga 3720 tttcgattcc accgccgcct tctatgaaag gttgggcttc ggaatcgttt tccgggacgc 3780 cggctggatg atcctccagc gcggggatct catgctggag ttcttcgccc accccaactt 3840 gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt tcacaaataa 3900 agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca 3960 tgtctgtata ccgtcgacct ctagctagag cttggcgtaa tcatggtcat agctgtttcc 4020 tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa gcataaagtg 4080 taaagcctgg ggtgcctaat gagtgagcta actcacatta attgcgttgc gctcactgcc 4140 cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg 4200 gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc 4260 ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac 4320 agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa 4380 ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca 4440 caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc 4500 gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata 4560 cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta 4620 tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca 4680 gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga 4740 cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg 4800 tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa cagtatttgg 4860 tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg 4920 caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag 4980 aaaaaaagga tctcaagaag atccttgat ctttctacg gggtctgacg ctcagtggaa 5040 cgaaaactca cgttaaggga ttttgtcat gagattatca aaaaggatct tcacctagat 5100 ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc 5160 tgacagttac caatgcttaa tcagtgaggc acctactca gcgatctgtc tatttcgttc 5220 atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccac 5280 tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc 5340 aaaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgccctc 5400 catccagtct atttattgtt gccgggaagc tagtagt agttcgccag ttaatagttt 5460 gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc 5520 ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa 5580 aaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt 5640 atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg 5700 cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc 5760 gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa 5820 agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt 5880 gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt 5940 caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag 6000 ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta 6060 tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat 6120 aggggttccg cgcacatttc cccgaaaagt gccacctgac gtc 6163

Claims

1. A mitochondrial-based drug delivery system, comprising: (1) Mitochondria, and (2) a drug loaded into the mitochondria, wherein the drug is loaded into the mitochondria by non-free diffusion; The drug is a nucleic acid drug, and the nucleic acid drug is bound to the mitochondria through a nucleic acid binding protein or a nucleic acid binding domain thereof; After the nucleic acid drug is delivered into the cell, it exists in a state of being combined with or separated from the mitochondria; The nucleic acid binding domain is at least one of hTFAM1, CRT and UP1, wherein hTFAM1 is the DNA binding domain of human TFAM1, CRT is the DNA binding domain of TopII alpha, and UP1 is the non-specific RNA binding domain of human hnRNPA1 protein.

2. The drug delivery system according to claim 1, wherein the mitochondria are derived from autologous, allogeneic or xenogeneic genes and combinations thereof.

3. The drug delivery system according to claim 1, wherein the nucleic acid drug comprises DNA, RNA or its analogs, and combinations thereof.

4. drug delivery system according to claim 1, wherein the drug delivery system realizes the delivery of drug into cells, wherein the cells include inner surface of cell membrane, cytoplasm, nucleus and combination thereof.

5. The drug delivery system according to claim 1, wherein the nucleic acid binding protein or its nucleic acid binding domain is directly or indirectly bound to the mitochondrial outer membrane through the signal peptide of the mitochondrial outer membrane protein FIS1.

6. The drug delivery system according to any one of claims 1 to 5, wherein the drug delivery system is prepared by the following method: (1) Design X-Y1-pMag protein and nMagHigh1-Y2-FIS1.MTS protein respectively; wherein, X is any target drug, Y1 and Y2 are fluorescent proteins for labeling, pMag and nMagHigh1 are a pair of optogenetic proteins, and FIS1.MTS is a targeting peptide of the mitochondrial outer membrane protein FIS1; (2) constructing plasmids of the two proteins using pcDNA3.1 vector and transfecting HEK 293T cells to simultaneously express the two proteins; (3) 24 hours later, cells were irradiated with blue light for 10 min to allow the pMag and nMagHigh1 protein pairs to bind; (4) Y1- and Y2-labeled mitochondria were extracted from the cells and then co-cultured with unlabeled cells.

7. The drug delivery system according to claim 6, wherein the fluorescent protein is at least one of GFP, EGFP, mScarlet-i, YFP and CFP.

8. The drug delivery system according to claim 6, wherein the plasmid vector can also be pEGFP, pCMVp-NEO-BAN, pSV2pUC19DNA, pUC18DNA or PQE30.

9. The drug delivery system according to claim 6, wherein the transfected cells can also be CHO-K1, COS-7, HeLa, NIH-3T3, PC-3, A549, MCF-7, HuH-7, U-2OS or HepG2.

10. The drug delivery system according to claim 6, wherein the transfection reagent is Lipofectamine, X-tremeGENE 9, Escort TM III. Escort TM IV. NeuroPorter TM DOTAP methosulfate or Calcium Phosphate.

11. The drug delivery system according to claim 6, wherein the transfection method is liposome transfection, calcium phosphate transfection or electroporation transfection.

12. The drug delivery system according to claim 1, wherein the drug delivery system is prepared by the following method: The plasmid pcDNA3.1-hTFAM1-CRT-mScarlet-i-FIS1.MTS was constructed, wherein hTFAM1 is the DNA binding domain of human TFAM1, CRT is the DNA binding domain of TopII alpha, both of which can bind to DNA, mScarlet-i is a red fluorescent protein used to mark mitochondria carrying fusion proteins; FIS1.MTS is a signal peptide targeting the outer membrane of mitochondria; mouse cells were transfected with pcDNA3.1-hTFAM1-CRT-mScarlet-i-FIS1.MTS, the fusion protein was expressed, mitochondria were extracted from the cells, the mitochondria were resuspended in DMEM complete medium, plasmid pcDNA3.1-Mito-EGFP-2A-n.EGFP was added, and incubated at room temperature to obtain a drug delivery system capable of delivering Mito-EGFP and n.EGFP; wherein, 2A can translate one mRNA into two proteins, Mito-EGFP marks mitochondria and n.EGFP marks the nucleus.

13. The drug delivery system according to claim 12, wherein: The pcDNA3.1-hTFAM1-CRT-mScarlet-i-FIS1.MTS is the sequence shown in SEQ ID NO:8, and the pcDNA3.1-Mito-EGFP-2A-n.EGFP is the sequence shown in SEQ ID NO:

9.

14. The drug delivery system according to claim 1, wherein the drug delivery system is prepared by the following method: The plasmid pcDNA3.1-UP1-FIS1.MTS was constructed, wherein UP1 is the non-specific RNA binding domain of human hnRNPA1 protein, and FIS1.MTS is a signal peptide targeting the outer membrane of mitochondria. The cells were transfected with pcDNA3.1-UP1-FIS1.MTS, and mitochondria were extracted from the cells. The mitochondria were resuspended in DMEM complete medium, and Mito-mScarlet-i mRNA was added and incubated at room temperature to obtain a drug delivery system capable of delivering Mito-mScarlet-i mRNA.

15. The drug delivery system according to claim 14, wherein: The pcDNA3.1-UP1-FIS1.MTS is the sequence shown in SEQ ID NO: 10; The Mito-mScarlet-i mRNA is obtained by in vitro transcription of the plasmid pcDNA3.1-Mito-mScarlet-I, and the pcDNA3.1-Mito-mScarlet-I is the sequence shown in SEQ ID NO:11.

Images