KR101803088B1 - Vaccine composition using streptococcus parauberis cytoplasmic membrance protein in fishes - Google Patents

Abstract

The present invention relates to a vaccine composition using a Streptococcus parauberis cell membrane protein which is a causative organism of Streptococcus strains of fish, and more particularly to a vaccine composition using Streptococcus parauberis strain I, which is a causative organism of Streptococcus, Streptococcus parauberis type II and Streptococcus iniae were selected for the protein having antigenicity against one or more selected from the group consisting of a vaccine for the prevention and treatment of streptococci And can be usefully used as a composition.

Description

[0001] VACCINE COMPOSITION USING STREPTOCOCCUS PARAUBERIS CYTOPLASMIC MEMBRANCE PROTEIN IN FISHES [0002]

The present invention relates to a vaccine composition using a Streptococcus parauberis cell membrane protein which is a causative organ of Streptococcus strains of fish.

Streptococcus parauberis ( Streptococcus parauberis ) has a high segregation rate in domestic flounder farms and is emerging as a major cause of streptococcal infection. In flukes infected with S. parauberis , dark coloration, nocturnal hemorrhage, abnormal swimming and poor feeding, and death due to cachexia.

Flounder is a species that occupies a large part of domestic marine aquaculture. Domestic flounder farms are mostly located in the form of aquaculture tanks in Jeju, South and West coasts, and the damage caused by streptococcal infections is frequent. However, epidemiological studies have been lacking in response to major causative agents Than in those who were treated with antibiotics.

Streptococcus iniae is a representative marine gram-positive pathogen, and is a major cause of streptococcosis, which causes hemorrhagic septicemia in fish, especially in olive flounder (Paralichthys olivaceus). S. iniae infection of flounder is accompanied by symptoms such as meningoencephalitis, ocular hypertrophy and corneal opacity. It causes high mortality at the time of onset, which not only causes a significant economic impact on fish culture, but also poses a risk factor for workers in the flounder industry.

In addition, recent epidemiological studies have shown that S. parauberis is a major pathogen of streptococcal infection in flounder (Non-Patent Documents 1 and 2).

Currently, S. parauberis has only been identified, but the research on the prevention and treatment of S. parauberis has been limited. In addition, antibiotic resistant genes for erythromycin and tetracycline were found in S. parauberis , and the treatment with antibiotics was limited (Non-Patent Document 3). Therefore, development of a vaccine is urgent as a fundamental preventive measure.

Accordingly, the inventors of the present invention have studied a method of effectively preventing Streptococcus, and as a result, selected proteins having antigenicity against Streptococcus parauberis , and found that excellent vaccine efficacy was confirmed by inoculation with zebrafish Thus completing the present invention.

1. Nho SW, Shin GW, Park SB, Jang HB, Cha IS, Ha MA, Kim YR, Park YK, Dalvi RS, Kang BJ, Joh SJ, Jung TS. FEMS Microbiol Lett. 2009 Apr; 293 (1): 20-7; 2. Shin GW. Palaksha KJ. Yang HH. Shin YS. Kim YR. Lee EY. Kim HY. Kim YJ. Oh MJ. Yoshida T. Jung TS. Bull. Eur. Assoc. Fish Pathol. (2006) 26; 2 3. Park YK, Nho SW, Shin GW, Park SB, Jang HB, Cha IS, Ha MA, Kim YR, Dalvi RS, Kang BJ, Jung TS. Vet Microbiol. 2009 Apr 14; 136 (1-2): 76-81. Epub 2008 Oct 14.

It is an object of the present invention to provide a vaccine composition for treating or preventing a streptococcal infection of fish comprising a polypeptide consisting of any one or more amino acid sequences selected from the group consisting of SEQ ID NOS: 1 to 13 as an active ingredient.

Another object of the present invention is to provide an expression vector comprising a gene encoding the polypeptide.

It is still another object of the present invention to provide a host cell transformed with the expression vector.

It is another object of the present invention to provide a feed composition for the treatment or prevention of streptococcal infectious disease of fish comprising the vaccine composition.

It is yet another object of the present invention to provide a method for treating or preventing streptococcal infection of fish by administering the vaccine composition to fish.

In order to achieve the above object, the present invention provides a vaccine composition for treating or preventing a streptococcal infection of fish comprising, as an active ingredient, a polypeptide consisting of any one or more amino acid sequences selected from the group consisting of SEQ ID NOS: 1 to 13 to provide.

The present invention also provides an expression vector comprising the gene encoding the polypeptide.

The present invention also provides a host cell transformed with the expression vector.

The present invention also provides a feed composition for the treatment or prevention of streptococcal infectious disease of fish comprising the vaccine composition.

In addition, the present invention provides a method for treating or preventing a streptococcal infection of a fish by administering the vaccine composition to a fish.

Hereinafter, the present invention will be described in detail.

The present invention relates to a pharmaceutical composition comprising a compound of SEQ ID NO: SEQ ID NO: 2; SEQ ID NO: 3; SEQ ID NO: 4; SEQ ID NO: 5; SEQ ID NO: 6; SEQ ID NO: 7; SEQ ID NO: 8; SEQ ID NO: 9; SEQ ID NO: 10; SEQ ID NO: 11; SEQ ID NO: 12; Which comprises a polypeptide consisting of one or more amino acid sequences selected from the group consisting of SEQ ID NO: 13 to SEQ ID NO: 13 as an active ingredient. The present invention also provides a vaccine composition for treating or preventing a streptococcal infection of a fish.

The streptococcus is preferably Streptococcus parauberis .

SEQ ID NO: 1; SEQ ID NO: 2; SEQ ID NO: 3; SEQ ID NO: 4; SEQ ID NO: 5; SEQ ID NO: 6; SEQ ID NO: 7; SEQ ID NO: 8; SEQ ID NO: 9; SEQ ID NO: 10; SEQ ID NO: 11; SEQ ID NO: 12; SEQ ID NO: 13 is the antigen protein SP1 (STP-1656) present in the cell membrane of Streptococcus parauberis in the specific embodiment of the present invention; SP 2 (STP-0786); SP 3 (STP-1141); SP 4 (STP-1063); SP 5 (STP-0832); SP 6 (STP-1434); SP 7 (STP-0468); SP 8 (STP-0412); SP 9 (STP-0876); SP 10 (STP-1157); SP 11 (STP-1781); SP 12 (SPB-0027); SP 13 (SPB-1862); to be.

The fish is preferably at least one species selected from the group consisting of flounder, perch, trout, oyster, and dome, but is not limited thereto.

In the embodiment, the streptococcus is selected from the group consisting of Streptococcus para Ube Russ (Streptococcus parauberis) I-type, Streptococcus para Ube Russ (Streptococcus parauberis) II-type and (Streptococcus iniae) Streptococcus am.

In an embodiment of the present invention, SEQ ID NO: 1; SEQ ID NO: 2; SEQ ID NO: 5; SEQ ID NO: 7; SEQ ID NO: 8; SEQ ID NO: 9; Or the polypeptide consisting of the amino acid sequence of SEQ ID NO: 12 has been found to be antigenic in both Streptococcus parauberus type I and Streptococcus parauberus type II. Thus, SEQ ID NO: 1; SEQ ID NO: 2; SEQ ID NO: 5; SEQ ID NO: 7; SEQ ID NO: 8; SEQ ID NO: 9; Or a polypeptide consisting of the amino acid sequence of SEQ ID NO: 12 may serve as a multivalent vaccine for the treatment or prevention of fish streptococcal infection.

In an embodiment of the present invention, the polypeptide consisting of the amino acid sequence of SEQ ID NO: 7 or SEQ ID NO: 12 has a cross-antigen response to both Streptococcus parabeus type I, Streptococcus parabeus type II and Streptococcus species Respectively. Accordingly, the polypeptide consisting of the amino acid sequence of SEQ ID NO: 7 or SEQ ID NO: 12 may serve as a multivalent vaccine for the treatment or prevention of fish streptococcal infection.

The present invention also provides an expression vector comprising the gene encoding the polypeptide.

The present invention also relates to the above-mentioned composition comprising the amino acid sequence of SEQ ID NO: SEQ ID NO: 2; SEQ ID NO: 3; SEQ ID NO: 4; SEQ ID NO: 5; SEQ ID NO: 6; SEQ ID NO: 7; SEQ ID NO: 8; SEQ ID NO: 9; SEQ ID NO: 10; SEQ ID NO: 11; SEQ ID NO: 12; Or an amino acid sequence of SEQ ID NO: 13; and a gene comprising a nucleotide sequence encoding each polypeptide consisting of the amino acid sequence of SEQ ID NO: 13.

In an embodiment of the present invention, the nucleotide sequence is SEQ ID NO: 14; SEQ ID NO: 15; SEQ ID NO: 16; SEQ ID NO: 17; SEQ ID NO: 18; SEQ ID NO: 19; SEQ ID NO: 20; SEQ ID NO: 21; SEQ ID NO: 22; SEQ ID NO: 23; SEQ ID NO: 24; SEQ ID NO: 25; Or SEQ ID NO: 26; SEQ ID NO: 2; SEQ ID NO: 3; SEQ ID NO: 4; SEQ ID NO: 5; SEQ ID NO: 6; SEQ ID NO: 7; SEQ ID NO: 8; SEQ ID NO: 9; SEQ ID NO: 10; SEQ ID NO: 11; SEQ ID NO: 12; Or SEQ ID NO: 13, respectively.

The expression vector is preferably pET-30a, but is not limited thereto.

Also, variants of the above sequences are included within the scope of the present invention. SEQ ID NO: 14; SEQ ID NO: 15; SEQ ID NO: 16; SEQ ID NO: 17; SEQ ID NO: 18; SEQ ID NO: 19; SEQ ID NO: 20; SEQ ID NO: 21; SEQ ID NO: 22; SEQ ID NO: 23; SEQ ID NO: 24; SEQ ID NO: 25; Or SEQ ID NO: 26, respectively. Specifically, the sequence includes a nucleotide sequence having at least 70% sequence identity, more preferably at least 80% sequence identity, even more preferably at least 90% sequence identity, and most preferably at least 95% sequence identity with each of the above base sequences .

 The "% of sequence homology" for the polynucleotide is identified by comparing the comparison region with two optimally aligned sequences, and a portion of the polynucleotide sequence in the comparison region is the reference sequence for the optimal alignment of the two sequences (I. E., Gap), as compared to < / RTI >

The term "vector" is used to refer to a DNA fragment (s), nucleic acid molecule, which is transferred into a cell. The vector replicates the DNA and can be independently regenerated in the host cell. The term "expression vector" is often used interchangeably with a "recombinant vector ". The term "recombinant vector" means a recombinant DNA molecule comprising a desired coding sequence and a suitable nucleic acid sequence necessary for expressing a coding sequence operably linked in a particular host organism. Promoters, enhancers, termination signals and polyadenylation signals available in eukaryotic cells are known.

The present invention also provides a host cell transformed with the expression vector.

SEQ ID NO: 14; SEQ ID NO: 15; SEQ ID NO: 16; SEQ ID NO: 17; SEQ ID NO: 18; SEQ ID NO: 19; SEQ ID NO: 20; SEQ ID NO: 21; SEQ ID NO: 22; SEQ ID NO: 23; SEQ ID NO: 24; SEQ ID NO: 25; Or SEQ ID NO: 26; Each gene sequence can be inserted into a recombinant expression vector. The term "recombinant expression vector" means a bacterial plasmid, a phage, a yeast plasmid, a plant cell virus, a mammalian cell virus, or other vector. In principle, any plasmid and vector can be used if it can replicate and stabilize within the host. An important characteristic of the expression vector is that it has a replication origin, a promoter, a marker gene and a translation control element.

The present invention also provides a feed composition for the treatment or prevention of streptococcal infectious disease of fish comprising the vaccine composition.

The streptococcus is preferably Streptococcus parauberis .

In addition, the present invention provides a method for treating or preventing a streptococcal infection of a fish by administering the vaccine composition to a fish.

The streptococcus is preferably Streptococcus parauberis .

In the examples of the present invention, it has been confirmed that the effective antigen protein can be separated using the membrane protein separation method. Streptococcus parauberis type I, Streptococcus species , para to Ube Russ (Streptococcus parauberis) II type and Streptococcus am for the prevention and therapeutic vaccine composition for Streptococcus When selection was used this protein having the antigenicity to the one or more selected from the group consisting of (Streptococcus iniae) And it can be used effectively.

Use of a protein having excellent antigenicity against Streptococcus parauberus, which is a causative organism of streptococci according to the present invention, can be effectively utilized as a vaccine composition for the prevention and treatment of streptococci.

FIG. 1 shows the result of separation of cell membrane proteins of isolated streptococcal strains according to an embodiment of the present invention by SDS-PAGE.
Figure 2 is a graph showing the effect of S. parauberis on mouse antisera produced by antigen challenge of streptococcus according to an embodiment of the present invention serotype Type I and S. parauberis This is the result of Western blotting to identify proteins showing antigenicity in serotype II cell membrane proteins.
FIG. 3 shows the results of amplifying the isolated streptococcal antigen gene according to the embodiment of the present invention.
FIG. 4 shows the results of SDS-PAGE analysis of the expression of the identified protein according to the embodiment of the present invention.
FIG. 5 is a graph showing the relationship between the protein identified in FIG. 4 and the S. parauberis serotype Type I and S. parauberis This is the result of western blotting of antiserum produced by serotype II attack.
FIG. 6 is a Western blotting result showing that the antigen protein according to the embodiment of the present invention shows antigenicity to rabbit antiserum against S. iniae .

Hereinafter, the present invention will be described in more detail with reference to Examples. It is to be understood by those skilled in the art that these embodiments are merely illustrative of the present invention and that the scope of the present invention is not construed as being limited by these embodiments.

< Example  1> Isolation of Membrane Protein from Streptococcus

S. parauberis serotype I (S. parauberis strain S2 and S. parauberis strain YSFST02-111) were incubated at 37 ℃ and BHI (brain heart infusion, 0.5% glucose) for 18 hours, centrifuged at 5000 rpm for 10 minutes, and the cells were recovered and washed with solution A (80 mM Tris-HCl pH 7.4 and 1.2 M NaCl). And then sonicated for 90 seconds at 40 W for 5 seconds to prepare a cell extract. Solution B (40 mM Tris-HCl pH 7.4, 600 mM NaCl and 4% Triton X-114) corresponding to 1/3 of the solution was added. Then, the reaction was carried out at 4 DEG C for 1 hour using a rotor shaker. Cells were removed by centrifugation at 4 ° C at 12000 rpm for 20 minutes. The supernatant was reacted at 37 ° C for 10 minutes and centrifuged at room temperature for 5 minutes at 5000 rpm to remove the supernatant. The same amount of Solution C (20 mM Tris-HCl pH 7.4, 130 mM NaCl and 10 mM EDTA) was added to the supernatant. The reaction was carried out at 37 ° C for 10 minutes and centrifuged at 5000 rpm for 10 minutes at room temperature to remove the supernatant. The reaction was repeated twice, and the supernatant was removed. 2.5 times of ethanol as a detergent phase was added thereto, and the reaction was performed at -20 ° C to concentrate the protein. The cell membrane proteins of streptococci of the flounder were isolated by centrifugation at 4 ° C at 12,000 rpm for 10 minutes.

< Example  2> Streptococcus ( S. parauberis Serotype  I and S. parauberis Serotype  II) on cell membrane protein analysis

<2-1> Separation of Membrane Protein from Streptococcus

Streptococcal cell membrane proteins isolated in Example 1 were loaded on SDS-PAGE and isolated.

As a result, streptococcus S. parauberis Serotype I, S. parauberis Several tens of membrane protein bands isolated from serotype II were observed (Fig. 1).

<2-2> Streptococcal cell membrane protein

The cell membrane proteins of streptococcal streptococci isolated from Example 1 were subjected to SDS-PAGE and transferred to a nitrocellulose membrane (Nitrocellulose (NC) membrane). The nitrocellulose membrane was blocked with 5% skim milk in PBS for 1 hour at room temperature to remove the nonspecific reaction and washed with PBST (PBS, 0.05% Tween 20) three times for 15 minutes. Then, the streptococcal antiserum type I and II, which were produced in mouse, were added to PBST in which 0.5% skim milk was dissolved, respectively, and subjected to a first reaction at room temperature for 1 hour. The nitrocellulose membrane was washed with PBST three times for 15 minutes, and then subjected to secondary reaction for 1 hour at room temperature with anti-mouse IgG and HRP-linked antibody (cell signaling). The nitrocellulose membrane was washed with PBST three times for 15 minutes each, and the antigen protein was identified using a Western bright ™ Quantum Western blotting detection kit (Advansta).

As a result, S. parauberis Serotype I and S. parauberis against antiserum produced by the attack of streptococcal antigen A variety of proteins showing antigenicity in serotype II cell membrane proteins were identified (Fig. 2).

< Example  3> Antigenic Protein Analysis of Streptococcus

A band predicted to be an antigenic protein of streptococci showing reactivity in the result obtained with Western blot assay in Example 2 was commissioned to Daejeon Korea Basic Science Institute. Proteins were extracted by gel-in-digestion method and the antigenic protein of streptococcus was identified by Matrix-assisted Laser Desorption / Ionization Time of Flight Mass Spectrometry (MALDI-TOF MS).

Specifically, in the result obtained with the Western blot assay in Example 2, the band predicted to be the antigenic protein of the streptococcal strain was the best because the reactivity was the highest, and the band completely destined using 50% ACN / 10 mM NH4HCO3 The gel slice was washed with distilled water (dw) 2 to 3 times, dehydrated with acetonitrile (ACN), and dried using Speedvac. Then, 10 mM DTT / 100 mM NH ₄ HCO ₃ was added to the reaction solution for 1 hour, and 100 mM N NH ₄ HCO ₃ containing 55 mM IAA was added thereto. After washing twice more with distilled water, the gel was dehydrated using acetonatril. Then, the dried g-gel was incubated at 4 ° C for 30 minutes with 10 μl of a solution of 20 ng / μl of trypsin dissolved in 50 mM NH ₄ HCO ₃ buffer, and the insufficient amount was filled with 50 mM NH ₄ HCO ₃ buffer . Digestion at 37 ° C for 16 hours. The reaction supernatant was transferred to a tube and vortexed with 5% formic acid / 50% acetonatril and then allowed to react down at room temperature for 30 minutes. The supernatant thus obtained was collected and 100% acetonatril was added thereto, and the solutions obtained repeatedly 2 to 3 times were added together. This was dried using a Speedvac. The dried samples are dissolved in 0.5% TFA before MALDI analysis. Matrix is selected by HCCA (α-cyano-4-hydroxycinnamic acid) and dissolved in 0.01% TFA / 33% ACN. Ziptip c18 was mixed with a sample at a ratio of 1: 2 and loaded onto a plate for analysis of MALDI TOF. Peptide analysis was performed using Ultraflextreme MALDI TOF / TOF MS (Bruker, USA). The mass value of the obtained peptide MS / MS peak was identified using NCBI database (or DB of user) using Mascot.

As a result, the antigenic proteins of Streptococcus were listed in Table 1 below.

name MW Score ^a Similarity Putative function Localization Protein ^e % Identity SP 1
(STP-1656) 60.4 522 ABC transporter substrate-binding protein ( Streptococcus agalactiae ) 69 dppA (peptide binding protein) Extracellular
(lipid anchored) SP 2
(STP-0786) 37.0 104 Membrane protein
( Streptococcus didelphis ) 85 lipoprotein Extracellular
(lipid anchored) SP 3
(STP-1141) 33.8 85 D-ribose-binding protein
( Lactococcus lactis ) 62 D-ribose-binding protein Extracellular
(lipid anchored) SP 4
(STP-1063) 35.3 48 Foldase protein PrsA 1
( Streptococcus uberis 0140J ) 74 foldase protein PrsA Cytoplasmic membrane SP 5
(STP-0832) 54.6 115 Pyruvate kinase
( Streptococcus uberis 0140J ) 93 pyruvate kinase cytoplasmic SP 6
(STP-1434) 47.5 78 Trigger factor ( Streptococcus canis ) 89 문기기 cytoplasmic SP 7
(STP-0468) 47.2 288 Phosphopyruvate hydratase ( Streptoccus iniae SF1) 98 enolase cytoplasmic SP 8
(STP-0412) 43.8 244 Elongation factor Tu
( Streptococcus pyogenes MGAS315) 97 EF-Tu (elongation factor Tu) cytoplasmic SP 9
(STP-0876) 44.8 32 30S riobosomal protein S1 ( Streptococcus dysgalactiae ) 95 rpsA (30S ribosomal protein S1) cytoplasmic SP 10
(STP-1157) 46.6 22 Arginine deiminase
( Streptococcus pyogenes NZ131) 91 arA (arginine deiminase) cytoplasmic SP 11
(STP-1781) 28.3 17 Hypothetical protein
( Streptococcus porcinus ) 65 심포치 cytoplasmic SP 12
(SPB-0027) 34.8 277 Lipoprotein MtsA
(Streptococcus pyogenes) 79 Metal ABC transporter substrate-binding lipoprotein Cytoplasmic membrane SP 13
(SPB-1862) 34.3 148 ABC transporter substrate-binding lipoprotein ( Streptococcus urinalis ) 67 ABC transporter substrate binding protein Extracellular
(lipid anchored)

< Example  4> Identified  Of Streptococcal Antigenic Protein Cloning

In order to obtain an antigen gene of the protein identified by MALDI-TOF / MS in Example 3, gDNA (genomic DNA) of streptococcal strain was cloned using DNeasy Blood & Tissue kit (Qiagen, Cat No. 69506) .

Specifically, the isolated gDNA was obtained as a primer capable of amplifying the antigen gene, and the introduction into the expression vector was carried out by the In-fusion ^™ Advantage PCR cloning kit (Clontech, USA). The primers and the amplified genes used for obtaining the antigen gene are shown in Table 2 and FIG. The obtained antigen gene was cloned into pET-30a expression vector linealized with NdeI and XhoI using In-Fusion HD Enzyme Premix (Clontech, Cat No. ST0345). The prepared plasmids were transformed into E. coli BL21 (DE3). The transformants were cultured at 37 占 폚 in LB medium containing 50 占 퐂 / ml of antibiotic. When 0.5 at OD 600 was reached, 0.5 mM IPTG (isopropyl-β-thio-D-galactopyranoside) was added and the culture was further incubated at 37 ° C. for 3 hours. For further analysis, cells were collected by centrifugation and the pellet was disrupted by sonication. The supernatant was then recovered and the antigen-candidate recombinant fusion protein was analyzed by SDS-PAGE.

name The sequence (5'-3 ') SP1 Forward primer taagaaggagatatacatatgACAAGTTCAAAAAGCACTAATTG Reverse primer gtggtggtggtggtgctcgagTTTTTTGTAAGCGTAAGTGAAG SP2 Forward primer taagaaggagatatacatatgAACAAGAAAATTATTGGTC Reverse primer gtggtggtggtggtgctcgagGTTATTATTTTTCAGGAAC SP3 Forward primer taagaaggagatatacatatgAAATTTGGAAAAAAACTTGG Reverse primer gtggtggtggtggtgctcgagCCAGTTGTATTTATCAACGT SP4 Forward primer taagaaggagatatacatatgAAAAAATCTAATAAGTTAATTAC Reverse primer gtggtggtggtggtgctcgagTTTATCAACTGATGTTTTCAATG SP5 Forward primer taagaaggagatatacatatgAATAAACGCGTAAAAATTG Reverse primer gtggtggtggtggtgctcgagTTTAACAGTACGAACACGCAT SP6 Forward primer taagaaggagatatacatatgTCTACATCATTTGAAAACAA Reverse primer gtggtggtggtggtgctcgagTTTTACTTTTGCTGATTCTG SP7 Forward primer taagaaggagatatacatatgTCAATTATTACTGATGTTTAC Reverse primer gtggtggtggtggtgctcgagTTTTTTTAGGTTATAGAATGA SP8 Forward primer taagaaggagatatacatatgGCAAAAGAAAAATACGATC Reverse primer gtggtggtggtggtgctcgagAGCTTCGATTTCTGAAACGATAC SP9 Forward primer taagaaggagatatacatatgAATGAATTTGAAGATTTGC Reverse primer gtggtggtggtggtgctcgagTAATTCAATATCACCGAAC SP10 Forward primer taagaaggagatatacatatgACTGCTAAATCACCGATTC Reverse primer gtggtggtggtggtgctcgagGAGATCTTCACGTTCAAATG SP11 Forward primer taagaaggagatatacatatgTTAAAATTCCTGATTCCAAC Reverse primer gtggtggtggtggtgctcgagGGCCTCTTTTTTAATATAAAC SP12 Forward primer taagaaggagatatacatatgAAAACAATTGGAAAATCAG Reverse primer gtggtggtggtggtgctcgagTTTTTTATCTATATTGAC SP13 Forward primer taagaaggagatatacatatgAAATTTAAACAATTATTTTTTA Reverse primer gtggtggtggtggtgctcgagTTTGCTTTCTACTAATTGTTTTTCAAG

< Example  5> Identified  Analysis of antigenic candidate protein expression

Antigen candidate recombinant fusion proteins transformed in Example 4 were subjected to SDS-PAGE followed by Western blot analysis to confirm antigenicity.

Specifically, the candidate antigen recombinant fusion protein transformed in Example 4 was subjected to SDS-PAGE, and the resultant gel was transferred to a nitrocellulose membrane (Nitrocellulose (NC) membrane). The nitrocellulose membrane was blocked with 5% skim milk in PBS for 1 hour at room temperature to remove the nonspecific reaction and washed with PBST (PBS, 0.05% Tween 20) three times for 15 minutes. Then, the streptococcal antiserum type I and II, which were produced in mouse, were added to PBST in which 0.5% skim milk was dissolved, respectively, and subjected to a first reaction at room temperature for 1 hour. The nitrocellulose membrane was washed with PBST three times for 15 minutes, and then subjected to secondary reaction for 1 hour at room temperature with anti-mouse IgG and HRP-linked antibody (cell signaling). The nitrocellulose membrane was washed with PBST three times for 15 minutes each, and the antigen protein was identified using a Western bright ^™ Quantum Western blotting detection kit (Advansta).

As a result, the antigen protein reacted with the mouse serum generated from the antigen candidates identified in Example 4 was examined. S. parauberis In serotype I, SP1, SP2, SP5, SP7, SP8, SP9 and SP12 were found to be antigenic, and S. parauberis In serotype II, SP, SP2, SP3, SP5, SP6, SP7, SP8, SP9, SP10, SP12 and SP13 were found to be antigenic. The results are shown in FIG. 4 and FIG.

FIG. 4 shows the results of SDS-PAGE analysis of the expression of the identified proteins,

FIG. 5 is a graph showing the relationship between the protein identified in FIG. 4 and S. parauberis serotype Type I and S. parauberis Western blot results for antiserum produced by serotype II attack.

SP9 (rpsA; 30S ribosomal protein S1), SP2 (SPP), SP7 (enolase), SP8 (EF-Tu) , And SP12 (metal ABC transporter substrate-binding lipoprotein) antigen proteins were found to be antigenic in both S. parauberis serotype I and S. parauberis serotype II.

< Example  6> Crossover antigen response to causative bacteria causing streptococcal streptococcal infection

In the above <Example 5>, the recombinant fusion protein showing antigenicity, S. iniae (Chonnam National University Fisheries and Ocean University, YSFST01-82).

After performing SDS-PAGE in the above <Example 4>, the obtained gel was transferred to a nitrocellulose membrane. The nitrocellulose membrane was blocked with 5% skim milk dissolved in PBS for 1 hour at room temperature to remove the nonspecific reaction and washed 3 times with PBST for 15 minutes. Then, S. aurea antiserum produced in rabbit was added to PBST in which 0.5% skim milk was dissolved, and then primary reaction was performed at room temperature for 1 hour. The nitrocellulose membrane phosphorus was washed with PBST three times for 15 minutes each, and then subjected to secondary reaction for 1 hour at room temperature with anti-rabbit IgG and HRP-linked antibody (KOMABiotech). The nitrocellulose membrane was washed with PBST three times for 15 minutes each, and the antigen protein was identified using a Western bright ™ Quantum Western blotting detection kit (Advansta).

As a result, it was found that the antigenic proteins of SP6 (trigger factor), SP7 (enolase), SP10 (arcA), arginine deiminase (SPA), SP12 (metal ABC transporter substrate binding lipoprotein) It was confirmed that the produced rabbit antiserum showed antigenicity (Fig. 6).

S. parauberis causing streptococcal disease Serotype Type I, S. parauberis Serotype Type II strain and S. iniae SP7 (enolase) and SP12 (metal ABC transporter substrate-binding lipoprotein) proteins having both antigenicity in the strain can be used as vaccines for the prevention and treatment of streptococci.

Based on the above results, it can be applied to other pathogens and it can be applied as a rapid screening method of a vaccine for prevention and treatment based on the above results. do.

Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. In general, the nomenclature used herein is well known and commonly used in the art.

<110> Korea Research Institute of Bioscience & Biotechnology <120> Vaccine composition using Streptococcus parauberis cytoplasmic          membrance protein in fishes <130> P14R16D0835 <160> 52 <170> Kopatentin 2.0 <210> 1 <211> 553 <212> PRT <213> Streptococcus parauberis <400> 1 Met Thr Ser Ser Ser Ser Thr Asn Trp Lys Arg Val Gly Leu Gly Thr   1 5 10 15 Leu Thr Leu Ala Ser Ala Ala Val Leu Met Ala Cys Gly Asn Lys Ser              20 25 30 Thr Ala Ser Asn Asp Asn Lys Asn Glu Ile Asn Trp Tyr Thr Pro Thr          35 40 45 Glu Ile Ile Thr Leu Asp Ile Ser Lys Asn Thr Asp Thr Tyr Ser Ser      50 55 60 Leu Ala Ile Gly Asn Ser Gly Ser Asn Leu Leu Arg Val Asp Asp Lys  65 70 75 80 Gly Lys Leu Gln Pro Asp Leu Ala Glu Lys Val Asp Val Ser Glu Asp                  85 90 95 Gly Leu Thr Tyr Thr Ala Thr Leu Arg Asp Gly Leu Lys Trp Ser Asp             100 105 110 Gly Ser Glu Leu Thr Ala Lys Asp Phe Val Tyr Thr Trp Gln Arg Met         115 120 125 Val Asp Pro Thr Thr Ala Ser Glu Tyr Ala Tyr Leu Ala Thr Glu Ser     130 135 140 His Leu Lys Asn Ala Glu Asp Ile Asn Thr Gly Lys Asn Lys Asp Leu 145 150 155 160 Thr Ala Leu Gly Val Lys Ala Glu Gly Asn Lys Val Ile Phe Thr Leu                 165 170 175 Ser Glu Pro Ala Pro Gln Phe Lys Ser Leu Leu Ser Phe Ala Asn Phe             180 185 190 Val Pro Gln Lys Glu Asp Phe Val Lys Lys Ala Gly Lys Asp Tyr Gly         195 200 205 Thr Ser Glu Lys Gln Val Tyr Ser Gly Pro Tyr Thr Val Lys Asp     210 215 220 Trp Asn Gly Thr Ser Gly Thr Phe Lys Leu Val Lys Asn Thr Lys Tyr 225 230 235 240 Trp Asp Ala Lys Asn Val Lys Thr Lys Thr Ile Asn Val Gln Thr Val                 245 250 255 Lys Lys Pro Asp Thr Ala Val Gln Met Tyr Lys Gln Gly Lys Leu Asp             260 265 270 Tyr Ala Asn Ile Ser Gly Thr Ser Ala Ile Tyr Asn Ala Asn Lys Lys         275 280 285 Asn Lys Asp Val Thr Pro Val Leu Glu Ala Thr Thr Ala Tyr Met Val     290 295 300 Tyr Asn Gln Thr Gly Lys Val Glu Gly Leu Ser Asn Thr Lys Ile Arg 305 310 315 320 Gln Ala Leu Asn Leu Ala Thr Asp Arg Lys Gly Ile Val Ser Ala Ala                 325 330 335 Val Asp Thr Gly Ser Lys Pro Ala Thr Ala Ile Ala Pro Thr Gly Leu             340 345 350 Ala Lys Leu Lys Asn Gly Thr Asp Leu Thr Lys His Val Ala Pro Gly         355 360 365 Tyr Ser Tyr Asp Glu Lys Lys Ala Gly Glu Leu Phe Lys Glu Gly Leu     370 375 380 Lys Glu Leu Gly Lys Asp Ser Leu Lys Val Thr Val Thr Ala Asp Ala 385 390 395 400 Asp Ser Pro Val Ala Lys Ala Ser Ile Asp Tyr Ile Lys Glu Thr Trp                 405 410 415 Glu Lys Ala Leu Pro Gly Leu Thr Val Glu Glu Lys Phe Val Thr Phe             420 425 430 Lys Gln Arg Leu Glu Asp Thr Lys Asn Gln Asn Phe Glu Val Ala Leu         435 440 445 Val Leu Trp Gly Gly Asp Tyr Pro Glu Gly Ser Thr Phe Tyr Gly Leu     450 455 460 Met Lys Ser Gly Ser Ala Tyr Asn Tyr Gly Lys Phe Thr Asn Glu Thr 465 470 475 480 Phe Asp Lys Ala Tyr Glu Lys Ala Leu Thr Thr Asp Ala Leu Asp Thr                 485 490 495 Asn Ala Ala Asp Asp Tyr Lys Asp Gly Glu Lys Ala Leu Tyr Glu             500 505 510 Ala Ala Asn Tyr Asn Pro Ile Tyr Phe Arg Ser Gly Glu Gly Leu Gln         515 520 525 Asn Pro Ser Ile Lys Gly Leu Ile Arg Asn Ser Thr Gly Leu Asn Val     530 535 540 Asp Phe Thr Tyr Ala Tyr Lys Lys *** 545 550 <210> 2 <211> 354 <212> PRT <213> Streptococcus parauberis <400> 2 Met Asn Lys Lys Ile Ile Gly Leu Gly Leu Thr Ser Val Ala Leu Leu   1 5 10 15 Ser Leu Ala Ala Cys Gly Asn Arg Gly Ala Ser Lys Ser Asp Ser Lys              20 25 30 Asp Ala Lys Thr Asp Leu Lys Ala Ala Ile Val Thr Asp Thr Gly Gly          35 40 45 Val Asp Asp Lys Ser Phe Asn Gln Ser Ala Trp Glu Gly Leu Glu Ala      50 55 60 Trp Gly Lys Glu Asn Gly Leu Lys Lys Gly Ala Gly Phe Asp Tyr Phe  65 70 75 80 Gln Ser Asn Ser Glu Ser Glu Tyr Ala Thr Asn Leu Asp Thr Ala Val                  85 90 95 Ser Asn Gly Tyr Lys Val Val Tyr Gly Ile Gly Phe Ala Leu Lys Asp             100 105 110 Ala Ile Ala Lys Ala Ala Gly Asp Asn Thr Asp Val Asn Phe Val Ile         115 120 125 Ile Asp Asp Val Ile Glu Gly Lys Asp Asn Val Ala Ser Val Thr Phe     130 135 140 Ala Asp Asn Ala Ala Lys Thr 145 150 155 160 Thr Lys Thr Lys Val Val Gly Phe Val Gly Gly Met Lys Gly Thr Val                 165 170 175 Ile Thr Arg Phe Glu Lys Gly Phe Glu Ala Gly Val Lys Ser Val Asp             180 185 190 Asp Ser Ile Lys Val Lys Val Asp Tyr Ala Gly Ser Phe Gly Asp Ala         195 200 205 Ser Lys Gly Lys Ala Ile Ala Ala Ala Gln Tyr Ala Gly Gly Ala Asp     210 215 220 Val Ile Tyr Gln Ala Gly Aly Gly Aly Gly Aly Gly Aly Gly Val Phe Asn Glu 225 230 235 240 Ala Lys Ser Ile Asn Glu Lys Arg Ser Glu Asp Glu Lys Val Trp Val                 245 250 255 Val Gly Val Asp Arg Asp Gln Glu Ala Glu Gly Lys Tyr Thr Thr Asn             260 265 270 Asp Lys Lys Glu Ser Asn Phe Val Leu Ala Ser Ser Ile Lys Gln Val         275 280 285 Gly Lys Ser Val Glu Leu Ile Asn Lys Leu Val Ala Asp Lys Lys Phe     290 295 300 Pro Gly Gly Lys Ser Thr Val Tyr Gly Leu Lys Asp Gly Gly Val Asp 305 310 315 320 Leu Ala Thr Thr Asn Leu Asn Asp Asp Ala Ile Lys Ala Val Lys Glu                 325 330 335 Ala Lys Glu Lys Leu Phe Leu Val Thr Ser Lys Phe Leu Lys Asn Asn             340 345 350 Asn ***         <210> 3 <211> 323 <212> PRT <213> Streptococcus parauberis <400> 3 Met Lys Phe Gly Lys Lys Leu Gly Phe Phe Ala Leu Leu Met Ser Leu   1 5 10 15 Val Leu Val Leu Gly Ala Cys Gly Lys Thr Gly Leu Gly Asn Ser Ser              20 25 30 Gly Ser Ser Ser Lys Glu Val Thr Lys Lys Ala Ala Lys Asp Leu Lys          35 40 45 Leu Gly Val Ser Ser Ser Thr Asn Asn Pro Tyr Phe Val Ser Met      50 55 60 Lys Asp Gly Leu Asp Ser Tyr Ala Lys Asp Lys Lys Val Ser Leu Lys  65 70 75 80 Val Ala Asp Ala Gln Asp Asp Ala Ala Arg Gln Ala Asp Asp Ile Gln                  85 90 95 Asn Phe Ile Ser Gln Asn Val Asp Ala Ile Leu Ile Asn Pro Val Asp             100 105 110 Ser Lys Ala Val Val Thr Ser Ile Lys Ala Ala Asn Ser Ala Asn Ile         115 120 125 Pro Val Ile Leu Ile Asp Arg Gly Ser Glu Gly Gly Asp Val Leu Thr     130 135 140 Thr Val Ala Ser Asp Asn Val Ala Ala Gly Lys Met Ala Ala Glu Phe 145 150 155 160 Ala Val Lys Glu Leu Gly Glu Lys Ala Lys Thr Phe Glu Leu Ser Gly                 165 170 175 Val Pro Gly Ala Ser Ala Thr Val Asp Arg Gly Lys Gly Phe Asn Asp             180 185 190 Ile Ala Lys Thr Lys Leu Asp Ile Leu Ser Ser Gln Ser Ala Asn Phe         195 200 205 Asp Arg Ala Lys Ala Leu Asn Thr Ala Gln Asn Met Ile Gln Gly Asn     210 215 220 Lys Asp Val Gln Val Ile Phe Ala Gln Asn Asp Glu Met Ala Leu Gly 225 230 235 240 Ala Ala Gln Ala Ala Lys Ser Ala Gly Leu Lys Asp Val Met Ile Ile                 245 250 255 Gly Ile Asp Gly Gln Pro Asp Ala His Asp Ala Ile Lys Lys Gly Asp             260 265 270 Ile Thr Ala Thr Ile Ala Gln Gln Pro Ala Lys Met Gly Glu Ile Ala         275 280 285 Ile Gln Ala Ile Asp His Tyr Gln Gly Lys Lys Val Glu Lys Ser     290 295 300 Thr Val Ser Pro Ile Tyr Leu Val Lys Lys Asp Asn Val Asp Lys Tyr 305 310 315 320 Asn Trp ***             <210> 4 <211> 315 <212> PRT <213> Streptococcus parauberis <400> 4 Met Lys Lys Ser Asn Lys Leu Ile Thr Gly Phe Ile Thr Leu Ala Ser   1 5 10 15 Val Met Thr Leu Ala Ala Cys Asn Ser Thr Asn Asp Asn Thr Lys Leu              20 25 30 Val Ser Met Lys Gly Asp Thr Ile Thr Val Thr Asp Phe Tyr Asn Glu          35 40 45 Ala Lys Thr Thr Gln Ala Gln Gln Ser Met Leu Ser Leu Ile Leu      50 55 60 Ser Arg Val Phe Glu Gly Gln Tyr Gly Lys Lys Val Ser Asp Lys Glu  65 70 75 80 Val Glu Lys Ser Tyr Asn Lys Thr Ala Glu Gln Tyr Gly Ala Ser Phe                  85 90 95 Ser Glu Ala Leu Ser Gln Ala Gly Met Thr Thr Glu Thr Tyr Lys Lys             100 105 110 Gln Ile Arg Thr Thr Met Leu Val Thr Tyr Ala Val Lys Gln Glu Ala         115 120 125 Glu Lys Glu Leu Thr Asp Ala Asn Tyr Lys Lys Ala Tyr Asp Asn Tyr     130 135 140 Thr Pro Lys Met Thr Thr Gln Val Ile Thr Leu Asp Asp Glu Glu Lys 145 150 155 160 Ala Lys Lys Val Leu Glu Glu Thr Lys Ala Glu Gly Ala Asn Phe Glu                 165 170 175 Asp Ile Ala Lys Lys Asn Thr Val Asn Glu Lys Lys Lys Val Asp Tyr             180 185 190 Thr Phe Asp Ser Ser Asp Lys Ser Leu Pro Ser Asp Val Ile Lys Glu         195 200 205 Ala Gly Lys Leu Lys Ala Gly Glu Lys Ser Glu Leu Ile Ser Val Met     210 215 220 Asp Pro Ser Thr Tyr Gln Lys Lys Tyr Tyr Ile Val Asn Met Ala Lys 225 230 235 240 Lys Glu Asp Lys Lys Ala Asp Trp Lys Glu Tyr Lys Thr Arg Leu Lys                 245 250 255 Glu Ile Ile Leu Asp Glu Lys Asn Asn Asp Thr Ala Phe Gln Asn Lys             260 265 270 Val Ile Ala Lys Glu Leu Asp Arg Ala Asn Val Lys Ile Lys Asp Lys         275 280 285 Ala Phe Ser Asn Ile Leu Ser Gln Phe Ala Ser Asp Lys Gln Ser Ala     290 295 300 Asn Ser Ala Leu Lys Thr Ser Val Asp Lys *** 305 310 315 <210> 5 <211> 501 <212> PRT <213> Streptococcus parauberis <400> 5 Met Asn Lys Arg Val Lys Ile Val Ala Thr Leu Gly Pro Ala Val Glu   1 5 10 15 Ile Arg Gly Gly Lys Lys Tyr Gly Glu Asp Gly Tyr Trp Ala Gly Lys              20 25 30 Leu Asp Val Glu Ala Ser Ala Lys Lys Ile Ala Glu Leu Ile Glu Ala          35 40 45 Gly Ala Asn Val Phe Arg Phe Asn Phe Ser His Gly Asn His Gln Glu      50 55 60 Gln Gly Asp Arg Met Ala Thr Val Arg Arg Ala Glu Glu Met Ala Arg  65 70 75 80 Gln Lys Val Ala Phe Leu Leu Asp Thr Lys Gly Pro Glu Met Arg Thr                  85 90 95 Glu Leu Phe Glu Asp Gly Ser Lys Glu Phe Ser Tyr Thr Thr Gly Glu             100 105 110 Thr Ile Arg Val Ala Thr Glu Gln Gly Ile Lys Ser Thr Arg Glu Val         115 120 125 Ile Ala Leu Ser Val Ala Gly Asn Leu Asp Ile Tyr Asp Glu Val Glu     130 135 140 Val Gly Asn Thr Ile Leu Ile Asp Asp Gly Lys Leu Gly Leu Lys Ile 145 150 155 160 Val Glu Lys Asp Ile Ser Thr Arg Gln Phe Val Thr Val Val Glu Asn                 165 170 175 Asp Gly Leu Ile Ala Asn Gln Lys Gly Val Asn Ile Pro Asn Thr Lys             180 185 190 Ile Pro Phe Pro Ala Leu Ala Glu Arg Asp Asn Ala Asp Ile Arg Phe         195 200 205 Gly Leu Glu Gln Gly Leu Asn Phe Ile Ala Ile Ser Phe Val Arg Thr     210 215 220 Ala Lys Asp Val Glu Glu Val Arg Glu Ile Cys Arg Glu Thr Gly Asn 225 230 235 240 Glu His Val Gln Leu Phe Ala Lys Ile Glu Asn Gln Gln Gly Ile Asp                 245 250 255 Asn Leu Asp Glu Ile Glu Ala Asp Gly Ile Met Ile Ala Arg             260 265 270 Gly Asp Met Gly Ile Glu Val Pro Phe Glu Met Val Pro Val Phe Gln         275 280 285 Lys Met Ile Ile Thr Lys Val Asn Ala Ala Gly Lys Ala Val Ile Thr     290 295 300 Ala Thr Asn Met Leu Glu Thr Met Thr Asp Lys Pro Arg Ala Thr Arg 305 310 315 320 Ser Glu Val Ser Asp Val Phe Asn Ala Val Ile Asp Gly Thr Asp Ala                 325 330 335 Thr Met Leu Ser Gly Glu Ser Ala Asn Gly Lys Tyr Pro Val Glu Ser             340 345 350 Val Arg Thr Met Ala Thr Ile Asp Lys Asn Ala Gln Thr Leu Leu Asn         355 360 365 Glu Tyr Gly Arg Leu Asp Ser Ser Lys Phe Pro Arg Thr Asn Lys Thr     370 375 380 Asp Val Ile Ala Ser Ala Val Lys Asp Ala Thr His Ser Met Asp Ile 385 390 395 400 Lys Leu Val Val Thr Ile Thr Glu Ser Gly Tyr Thr Ala Arg Ser Ile                 405 410 415 Ser Lys Phe Arg Pro Asp Ala Asp Ile Leu Ala Ile Thr Phe Asp Glu             420 425 430 Lys Val Gln Lys Ala Leu Met Ile Tyr Trp Gly Val Ile Pro Val Leu         435 440 445 Ala Thr Lys Pro Ala Ser Thr Asp Asp Met Phe Glu Val Ala Glu Glu     450 455 460 Val Ala Leu Lys Ala Gly Leu Val Lys Ser Gly Asp Asn Ile Val Ile 465 470 475 480 Val Ala Gly Val Val Gly Ser Gly Gly Thr Asn Thr Met Arg Val                 485 490 495 Arg Thr Val Lys ***             500 <210> 6 <211> 428 <212> PRT <213> Streptococcus parauberis <400> 6 Met Ser Thr Ser Phe Glu Asn Lys Ala Thr Asn Arg Gly Val Ile Thr   1 5 10 15 Phe Thr Ile Gly Gln Glu Gln Ile Lys Pro Ala Leu Asp Gln Ala Phe              20 25 30 Asn Lys Ile Lys Lys Asp Leu Asn Val Pro Gly Phe Arg Lys Gly His          35 40 45 Met Pro Arg Pro Val Phe Asn Gln Lys Phe Gly Glu Glu Val Leu Tyr      50 55 60 Glu Asp Ala Leu Asn Ile Val Leu Pro Ala Thr Tyr Glu Ala Ala Val  65 70 75 80 Thr Glu Leu Gly Leu Glu Val Val Ala Gln Pro Lys Ile Asp Val Val                  85 90 95 Ser Met Glu Lys Gly Gln Asp Trp Lys Ile Asn Ala Glu Val Val Thr             100 105 110 Lys Pro Glu Val Lys Leu Gly Asp Tyr Lys Asn Leu Ala Val Glu Val         115 120 125 Glu Ala Ser Lys Glu Val Ser Asp Glu Glu Ile Asp Glu Lys Ile Glu     130 135 140 Arg Glu Arg Lys Asn Leu Ala Glu Leu Val Val Lys Glu Asp Ala Ala 145 150 155 160 Ala Glu Gly Asp Thr Val Val Ile Asp Phe Leu Gly Ser Val Asp Gly                 165 170 175 Val Glu Phe Asp Gly Gly Lys Gly Asp Asn Phe Ser Leu Glu Leu Gly             180 185 190 Ser Gly Gln Phe Ile Pro Gly Phe Glu Glu Gln Leu Val Gly Val Lys         195 200 205 Ala Gly Glu Thr Val Asp Val Asn Val Thr Phe Pro Glu Glu Tyr Gln     210 215 220 Ala Glu Asp Leu Ala Gly Lys Asp Ala Lys Phe Val Thr Thr Val His 225 230 235 240 Glu Val Lys Thr Lys Glu Val Pro Glu Leu Asp Asp Glu Leu Ala Lys                 245 250 255 Asp Ile Asp Glu Glu Val Glu Thr Leu Asp Glu Leu Lys Ala Lys Tyr             260 265 270 Arg Lys Glu Leu Glu Ala Ala Lys Glu Ile Ala Tyr Asp Asp Ala Val         275 280 285 Glu Gly Ala Ala Ile Glu Leu Ala Val Glu Asn Ala Glu Ile Val Glu     290 295 300 Leu Pro Glu Glu Met Val His Asp Glu Val His Arg Ala Met Asn Glu 305 310 315 320 Phe Met Gly Asn Met Gln Arg Gln Gly Ile Ser Pro Glu Met Tyr Tyr                 325 330 335 Gln Leu Thr Gly Thr Ser Glu Asp Asp Leu His Lys Gln Tyr Glu Ser             340 345 350 Glu Ala Asp Lys Arg Val Lys Thr Asn Leu Val Ile Glu Ala Ile Ala         355 360 365 Lys Ala Glu Gly Phe Glu Ala Ser Asp Glu Glu Ile Glu Lys Glu Ile     370 375 380 Asn Asp Leu Ala Thr Glu Tyr Asn Met Pro Val Glu Gln Val Arg Thr 385 390 395 400 Leu Leu Ser Ala Asp Met Leu Lys His Asp Ile Ala Met Lys Lys Ala                 405 410 415 Val Glu Val Ile Thr Glu Ser Ala Lys Val Lys ***             420 425 <210> 7 <211> 436 <212> PRT <213> Streptococcus parauberis <400> 7 Met Ser Ile Ile Thr Asp Val Tyr Ala Arg Glu Val Leu Asp Ser Arg   1 5 10 15 Gly Asn Pro Thr Leu Glu Val Glu Val Tyr Thr Glu Ser Gly Ala Phe              20 25 30 Gly Arg Gly Met Val Ser Ser Gly Ala Ser Thr Gly Glu His Glu Ala          35 40 45 Val Glu Leu Arg Asp Gly Asp Lys Ser Arg Tyr Asn Gly Leu Gly Thr      50 55 60 Glu Lys Ala Val Asp Asn Val Asn Asn Ile Ile Ala Glu Ala Ile Ile  65 70 75 80 Gly Phe Asp Val Arg Asp Gln Gln Ala Ile Asp Arg Ala Met Ile Ala                  85 90 95 Leu Asp Gly Thr Pro Asn Lys Gly Lys Leu Gly Ala Asn Ala Ile Leu             100 105 110 Gly Val Ser Ile Val Ala Arg Ala Ala Asp Tyr Leu Glu Val         115 120 125 Pro Leu Tyr Asn Tyr Leu Gly Gly Phe Asn Thr Lys Val Leu Pro Thr     130 135 140 Pro Met Met Asn Ile Ile Asn Gly Gly Ser His Ser Asp Ala Pro Ile 145 150 155 160 Ala Phe Gln Glu Phe Met Ile Met Pro Val Gly Ala Thr Thr Phe Lys                 165 170 175 Glu Ala Leu Arg Trp Gly Ala Glu Val Phe His Ala Leu Lys Lys Ile             180 185 190 Leu Lys Glu Arg Gly Leu Glu Thr Ala Val Gly Asp Glu Gly Gly Phe         195 200 205 Ala Pro Lys Phe Asp Gly Thr Glu Asp Gly Val Glu Thr Ile Leu Lys     210 215 220 Ala Ile Gly Ala Gly Aly Gly Aly Gly Glu Asn Gly Ile Met Ile 225 230 235 240 Gly Phe Asp Cys Ala Ser Ser Glu Phe Tyr Asp Lys Glu Arg Lys Val                 245 250 255 Tyr Asp Tyr Ala Lys Phe Glu Gly Glu Gly Gly Ala Val Arg Thr Ser             260 265 270 Ala Glu Gln Ile Asp Tyr Leu Glu Glu Leu Val Asn Lys Tyr Pro Ile         275 280 285 Ile Thr Ile Glu Asp Gly Met Asp Glu Asn Asp Trp Asp Gly Trp Lys     290 295 300 Ala Leu Thr Glu Arg Leu Gly Gly Arg Val Gln Leu Val Gly Asp Asp 305 310 315 320 Phe Phe Val Thr Asn Thr Glu Tyr Leu Ala Arg Gly Ile Lys Glu Glu                 325 330 335 Ala Ala Asn Ser Ile Leu Ile Lys Val Asn Gln Ile Gly Thr Leu Thr             340 345 350 Glu Thr Phe Glu Ala Ile Glu Met Ala Lys Glu Ala Gly Phe Thr Ala         355 360 365 Val Val Ser His Arg Ser Gly Glu Thr Glu Asp Ser Thr Ile Ala Asp     370 375 380 Ile Ala Val Ala Thr Asn Ala Gly Gln Ile Lys Thr Gly Ser Leu Ser 385 390 395 400 Arg Thr Asp Arg Ile Ala Lys Tyr Asn Gln Leu Leu Arg Ile Glu Asp                 405 410 415 Gln Leu Gly Glu Val Ala Gln Tyr Lys Gly Ile Lys Ser Phe Tyr Asn             420 425 430 Leu Lys Lys ***         435 <210> 8 <211> 399 <212> PRT <213> Streptococcus parauberis <400> 8 Met Ala Lys Glu Lys Tyr Asp Arg Ser Lys Pro His Val Asn Ile Gly   1 5 10 15 Thr Ile Gly His Val Asp His Gly Lys Thr Thr Leu Thr Ala Ala Ile              20 25 30 Thr Thr Val Leu Ala Arg Arg Leu Pro Ser Ser Val Asn Gln Pro Lys          35 40 45 Asp Tyr Ala Ser Ile Asp Ala Ala Pro Glu Glu Arg Glu Arg Gly Ile      50 55 60 Thr Ile Asn Thr Ala His Val Glu Tyr Glu Thr Glu Ser Arg His Tyr  65 70 75 80 Ala His Ile Asp Ala Pro Gly His Ala Asp Tyr Val Lys Asn Met Ile                  85 90 95 Thr Gly Ala Glad Met Asp Gly Ala Ile Le Val Val Ala Ser Thr             100 105 110 Asp Gly Pro Met Pro Gln Thr Arg Glu His Ile Leu Leu Ser Arg Gln         115 120 125 Val Gly Val Lys His Leu Ile Val Phe Met Asn Lys Val Asp Leu Val     130 135 140 Asp Asp Glu Glu Leu Leu Glu Leu Val Glu Met Glu Ile Arg Asp Leu 145 150 155 160 Leu Thr Glu Tyr Asp Phe Pro Gly Asp Asp Leu Pro Val Ile Gln Gly                 165 170 175 Ser Ala Leu Lys Ala Leu Glu Gly Asp Thr Ala His Glu Asp Ile Ile             180 185 190 Met Glu Leu Met Lys Thr Val Asp Glu Tyr Ile Pro Glu Pro Glu Arg         195 200 205 Asp Thr Asp Lys Pro Leu Leu Leu Pro Val Glu Asp Val Phe Ser Ile     210 215 220 Thr Gly Arg Gly Thr Val Ala Ser Gly Arg Ile Asp Arg Gly Thr Val 225 230 235 240 Arg Val Asn Asp Glu Ile Glu Ile Val Gly Leu Lys Glu Asp Thr Lys                 245 250 255 Lys Ala Val Val Thr Gly Val Glu Met Phe Arg Lys Gln Leu Asp Glu             260 265 270 Gly Leu Ala Gly Asp Asn Val Gly Ile Leu Leu Arg Gly Val Gln Arg         275 280 285 Asp Glu Ile Glu Arg Gly Gln Val Ile Ala Lys Pro Gly Ser Ile Asn     290 295 300 Pro His Thr Lys Phe Lys Gly Glu Val Tyr Ile Leu Ser Lys Glu Glu 305 310 315 320 Gly Gly Arg His Thr Pro Phe Phe Asn Asn Tyr Arg Pro Gln Phe Tyr                 325 330 335 Phe Arg Thr Thr Asp Val Thr Gly Ser Ile Glu Leu Pro Ala Gly Thr             340 345 350 Glu Met Met Met Pro Gly Asp Asn Val Ala Ile Gln Val Glu Leu Ile         355 360 365 His Pro Ile Ala Val Glu Gln Gly Thr Thr Phe Ser Ile Arg Glu Gly     370 375 380 Gly Arg Thr Val Gly Ser Gly Ile Val Ser Glu Ile Glu Ala *** 385 390 395 <210> 9 <211> 411 <212> PRT <213> Streptococcus parauberis <400> 9 Met Asn Glu Phe Glu Asp Leu Leu Asn Ser Val Ser Glu Val Asn Pro   1 5 10 15 Gly Asp Val Val Thr Ala Glu Val Leu Thr Val Asp Asn Gly Gln Ala              20 25 30 Asn Val Val Ile Asp Gly Thr Gly Val Glu Gly Val Leu Thr Leu Arg          35 40 45 Glu Leu Thr Asn Asp Arg Asp Ala Asp Ile Asn Asp Phe Val Lys Ala      50 55 60 Gly Asp Thr Val Glu Val Leu Val Leu Arg Gln Val Val Gly Lys Asp  65 70 75 80 Thr Asp Thr Val Thr Phe Leu Val Ser Lys Lys Arg Leu Glu Ala Arg                  85 90 95 Lys Ala Trp Asp Lys Leu Val Gly Arg Glu Gly Glu Val Val Thr Val             100 105 110 Lys Gly Thr Arg Ala Val Lys Gly Gly Leu Ser Val Glu Phe Glu Gly         115 120 125 Leu Arg Gly Phe Ile Pro Ala Ser Met Ile Asp Thr Arg Phe Val Arg     130 135 140 Asn Thr Glu Lys Phe Val Gly Gln Glu Phe Glu Ala Lys Ile Lys Glu 145 150 155 160 Val Asp Ala Glu Asn Arg Phe Ile Leu Ser Arg Arg Glu Val Ile                 165 170 175 Glu Glu Ala Ala Lys Glu Ala Arg Gln Glu Val Phe Ser Lys Leu Ser             180 185 190 Glu Gly Ser Val Val Thr Gly Thr Val Ala Arg Leu Thr Ser Phe Gly         195 200 205 Ala Phe Ile Asp Leu Gly Gly Val Asp Gly Leu Val His Val Thr Glu     210 215 220 Leu Ser His Glu Arg Asn Val Ser Pro Lys Ser Val Val Ser Val Gly 225 230 235 240 Glu Glu Val Glu Val Lys Val Leu Ser Ile Asp Glu Glu Ala Gly Arg                 245 250 255 Val Ser Leu Ser Leu Lys Ala Thr Thr Pro Gly Pro Trp Asp Gly Val             260 265 270 Glu Gln Lys Leu Ala Gln Gly Asp Val Val Glu Gly Lys Val Lys Arg         275 280 285 Leu Thr Asp Phe Gly Ala Phe Val Glu Val Leu Ala Gly Ile Asp Gly     290 295 300 Leu Val His Ile Ser Gln Ile Ser His Asn Arg Val Glu Asn Pro Lys 305 310 315 320 Asp Val Leu Ser Val Gly Gln Glu Val Thr Val Lys Val Leu Glu Val                 325 330 335 Asn Ala Ala Asp Glu Aslan Val Ser Leu Ser Ile Lys Ala Leu Glu Glu             340 345 350 Arg Pro Ala Gln Ala Glu Gly Glu Asn Asn Tyr Asn Asn Asn Asn Asn         355 360 365 Asn Asn Asn Ser Glu Arg Gln Ser Arg Pro Arg Arg Pro Lys Arg     370 375 380 Glu Gln Lys Arg Asp Tyr Glu Leu Pro Glu Thr Gln Thr Gly Phe Ser 385 390 395 400 Met Ala Asp Leu Phe Gly Asp Ile Glu Leu ***                 405 410 <210> 10 <211> 412 <212> PRT <213> Streptococcus parauberis <400> 10 Met Thr Ala Lys Ser Pro Ile His Val Tyr Ser Glu Ile Gly Lys Leu   1 5 10 15 Lys Lys Val Leu Leu His Arg Pro Gly Lys Glu Ile Glu Asn Leu Met              20 25 30 Pro Asp Tyr Leu Glu Arg Leu Leu Phe Asp Asp Ile Pro Tyr Leu Glu          35 40 45 Asp Ala Gln Lys Glu His Asp Ala Phe Ala Glu Ala Leu Arg Asn Glu      50 55 60 Gly Val Glu Val Leu Tyr Leu Glu Lys Leu Ala Ala Glu Ser Leu Ile  65 70 75 80 Asn Asp Glu Val Arg Glu Gln Phe Ile Asp Glu Tyr Ile Ser Glu Ala                  85 90 95 Asn Ile Arg Gly Arg Ala Thr Lys Ile Ala Ile Arg Glu Phe Leu Met             100 105 110 Ala Ile Lys Asp Asn Gln Glu Leu Ile Asp Lys Thr Met Ala Gly Val         115 120 125 Gln Lys Ser Glu Leu Pro Glu Ile Ala Ala Glu Glu Lys Gly Leu Thr     130 135 140 Asp Leu Val Glu Ser Asn Tyr Pro Phe Ala Ile Asp Pro Met Pro Asn 145 150 155 160 Leu Tyr Phe Thr Arg Asp Pro Phe Ala Thr Ile Gly Glu Gly Val Ser                 165 170 175 Leu Asn His Met Phe Ser Glu Thr Arg Asn Arg Glu Thr Leu Tyr Gly             180 185 190 Lys Tyr Ile Phe Thr Tyr His Pro Val Tyr Gly Gly Glu Lys Val Pro         195 200 205 Met Val Tyr Asp Arg Ser Glu Asn Thr Arg Ile Glu Gly Gly Asp Glu     210 215 220 Leu Val Leu Ser Lys Asp Val Leu Ala Val Gly Ile Ser Gln Arg Thr 225 230 235 240 Asp Ala Ala Ser Ile Glu Lys Leu Leu Val Asn Ile Phe Lys Gln Asn                 245 250 255 Ile Gly Phe Lys Lys Val Leu Ala Phe Glu Phe Ala Asn Asn Arg Lys             260 265 270 Phe Met His Leu Asp Thr Val Phe Thr Met Val Asp Tyr Asp Lys Phe         275 280 285 Thr Ile His Pro Glu Ile Glu Gly Asp Leu Arg Val Tyr Ser Val Thr     290 295 300 Tyr Glu Asn Asp Asp Leu Lys Ile His Glu Glu Lys Gly Asp Leu Ala 305 310 315 320 Glu Leu Leu Ala Asn Leu Gly Val Glu Lys Val Glu Leu Ile Arg                 325 330 335 Cys Gly Gly Asp Asn Leu Val Ala Ala Gly Arg Glu Gln Trp Asn Asp             340 345 350 Gly Ser Asn Thr Leu Thr Ile Ala Pro Gly Val Val Val Tyr Asn         355 360 365 Arg Asn Thr Ile Thr Asn Ala Ile Leu Glu Ser Lys Gly Leu Arg Leu     370 375 380 Ile Lys Ile Asn Gly Ser Glu Leu Val Arg Gly Arg Gly Gly Pro Arg 385 390 395 400 Cys Met Ser Met Pro Phe Glu Arg Glu Asp Leu ***                 405 410 <210> 11 <211> 244 <212> PRT <213> Streptococcus parauberis <400> 11 Met Leu Lys Phe Leu Ile Pro Thr Ala Lys Glu Met Lys Val Pro Thr   1 5 10 15 Glu Ser Phe Pro Pro Leu Ile Pro Lys Phe Ser Gln Pro Ile Ile Ala              20 25 30 Met Met Ala Glu Leu Thr Leu Glu Glu Leu Ala Ser Val Tyr Lys Ile          35 40 45 Lys Pro Glu Ala Ala Gln Lys Glu Leu Asn Arg Ile Arg Glu Ile His      50 55 60 Gln Asn Gln Ala Leu Thr Tyr Pro Ala Tyr Lys Leu Phe Asn Gly Leu  65 70 75 80 Met Tyr Arg Tyr Ile Lys Arg Glu Asn Leu Ser Lys Lys Glu Glu Asn                  85 90 95 Tyr Leu Thr Lys Arg Thr Tyr Ile Thr Ser Ser Leu Tyr Gly Ile Ile             100 105 110 Pro Met Asp Phe Pro Ile Ala Glu His Arg Leu Asp Phe Gln Thr Lys         115 120 125 Ile Lys Ile Asn Gly Gln Asn Leu Lys Thr Tyr Trp Arg Pro Arg Tyr     130 135 140 Asp Gln Phe Leu Ser Pro Glu Asp Val Tyr Ile Ser Leu Leu Ser Ser 145 150 155 160 Glu Phe Glu Asp Ile Phe Ser Thr Asp Lys Arg Lys Leu Trp Ile Ser                 165 170 175 Ile Thr Phe Leu Glu Glu Lys Asp Gly Ile Pro Lys Ser His Ser Thr             180 185 190 Ile Ser Lys Lys Ala Arg Gly Ala Phe Leu Thr Ala Ala Met Lys Ala         195 200 205 Asn Cys Gln Ser Ser Ser Gln Leu Lys Ser Leu Lys Phe Asn Gly Phe     210 215 220 Ser Tyr Phe Ser Asp Leu Ser Asn Ala Asp His Phe Val Tyr Ile Lys 225 230 235 240 Lys Glu Ala ***                 <210> 12 <211> 311 <212> PRT <213> Streptococcus parauberis <400> 12 Met Lys Lys Lys Phe Thr Phe Ala Leu Thr Ala Ala Leu Ser Leu Phe   1 5 10 15 Met Leu Ser Ala Cys Ser Thr Ser Gly Gln Gln Ala Lys Lys Glu Glu              20 25 30 Lys Leu Asn Val Val Ala Thr Asn Ser Val Ile Ala Asp Met Thr Lys          35 40 45 Asn Ile Ala Gly Asn Lys Val Asn Leu His Ser Ile Val Pro Val Gly      50 55 60 Gln Asp Pro His Glu Tyr Glu Pro Leu Pro Glu Asp Val Glu Lys Thr  65 70 75 80 Ser Ser Ala Asp Leu Ile Phe Tyr Asn Gly Ile Asn Leu Glu Asp Gly                  85 90 95 Gly His Ala Trp Phe Thr Lys Leu Ile Lys Asn Ala Lys Lys Glu Lys             100 105 110 Asn Lys Asp Tyr Phe Ala Val Ser Asp Gly Ile Asp Val Ile Tyr Leu         115 120 125 Glu Gly Gln Asn Glu Lys Gly Lys Glu Asp Pro His Ala Trp Leu Asn     130 135 140 Leu Glu Asn Gly Met Ile Tyr Ser Lys Asn Ile Ala Lys Gln Leu Ile 145 150 155 160 Lys Lys Asp Pro Lys Asn Lys Thr Tyr Tyr Gln Ser His Leu Lys Ser                 165 170 175 Tyr Leu Ala Lys Leu Asp Asn Leu Asp Lys Glu Ala Lys Ser Lys Phe             180 185 190 Asp Lys Ile Pro Ala Asp Lys Lys Met Ile Val Thr Ser Glu Gly Cys         195 200 205 Phe Lys Tyr Phe Ser Lys Ala Tyr Asp Val Ser Ser Ala Tyr Ile Trp     210 215 220 Glu Ile Asn Thr Glu Glu Glu Gly Thr Pro Ser Gln Ile Ser Thr Leu 225 230 235 240 Ile Glu Lys Leu Lys Lys Lys Lys Lys Leu Ser Ala Ile Phe Val Glu Ser                 245 250 255 Ser Val Asp Ser Arg Pro Met Lys Ser Val Ser Lys Asp Ser Gly Ile             260 265 270 Pro Ile Tyr Ser Glu Ile Phe Thr Asp Ser Val Ala Lys Lys Gly Lys         275 280 285 Pro Gly Asp Ser Tyr Tyr Ala Met Met Lys Trp Asn Leu Asp Lys Ile     290 295 300 Ser Glu Gly Leu Ser Lys *** 305 310 <210> 13 <211> 324 <212> PRT <213> Streptococcus parauberis <400> 13 Met Lys Phe Lys Gln Leu Phe Phe Thr Val Gly Val Leu Ser Leu Leu   1 5 10 15 Val Ile Leu Ser Ala Cys Arg Arg Glu Ala Thr Glu Asn Lys Gly Gln              20 25 30 Val Lys Val Gly Ile Ile Gln Tyr Ala Lys His Pro Ala Leu Asp Ala          35 40 45 Ser Arg Lys Gly Phe Ile Glu Ala Leu Arg Asp Gly Gly Tyr Lys Glu      50 55 60 Gly Lys Asn Leu Gln Ile Thr Ser Lys Asn Ala Gln Gly Glu Gln Ala  65 70 75 80 Asn Leu Gln Thr Met Val Asp Gln Leu Thr Gly Lys Asn Asp Leu Asn                  85 90 95 Phe Ala Ile Ala Thr Pro Ala Ala Gln Ala Leu Leu Thr Ser Asp Pro             100 105 110 Asp Thr Pro Ala Val Phe Thr Ala Val Thr Asp Pro Val Gly Ala Gly         115 120 125 Leu Val Asp Ser Met Glu Lys Pro Gly Lys Asn Met Thr Gly Ser Ile     130 135 140 Asp Ala Gly Asn Val Pro Gln Gln Val Ala Leu Leu Leu Lys Ala Val 145 150 155 160 Pro Thr Ser Lys Lys Val Gly Ile Phe Tyr Asn Ser Ser Glu Val Asn                 165 170 175 Ser Gln Met Gln Ala Lys Glu Ala Lys Lys Ala Leu Lys Lys Lys Gly             180 185 190 Ile Ile Ala Val Glu Lys Thr Val Thr Ser Ser Asn Val Gln Gln         195 200 205 Val Met Thr Ser Leu Val Gly Gln Val Asp Ala Val Tyr Leu Pro Thr     210 215 220 Asp Asn Thr Val Ala Ser Thr Ala Met Thr Ile Gly Gln Ile Leu Lys 225 230 235 240 Glu Asn Lys Thr Pro Ala Ile Gly Ser Asp Lys Ala Tyr Leu Asp Ala                 245 250 255 Val Leu Phe Thr Ser Gly Val Asp Tyr Glu Ala Ile Gly Arg Gln Ala             260 265 270 Gly Glu Glu Ala Val Ala Ile Leu Asn Gly Lys Lys Ala Ser Thr Ile         275 280 285 Ala Val Ala Lys Pro Glu Lys Pro Glu Leu Ala Ile Asn Lys Asp Met     290 295 300 Ala Lys Ser Leu Gly Leu Asn Pro Lys Glu Leu Glu Lys Gln Leu Val 305 310 315 320 Glu Ser Lys ***                 <210> 14 <211> 1659 <212> DNA <213> Streptococcus parauberis <400> 14 atgacaagtt caaaaagcac taattggaaa cgtgttggtc ttggaacgtt gactttagct 60 tctgcagcag ttttaatggc atgtggaaat aaatcaactg cttctaatga taataaaaat 120 gaaatcaatt ggtacacacc aactgaaata atcactttag acatttcaaa aaatactgat 180 acgtattcat cacttgctat tggtaactca ggaagtaact tactacgagt tgatgataag 240 gggaaactcc aaccagattt agctgaaaaa gttgacgtat cagaagatgg cttaacttat 300 acagcaactt tacgtgatgg tttgaaatgg tcagacggta gtgaattaac tgcaaaagat 360 ttcgtctata cttggcaacg tatggttgat ccaacaactg cttctgagta tgcctatctt 420 gcaactgaat cacatttgaa aaatgctgaa gatatcaata ctggtaaaaa taaagattta 480 actgctctag gtgttaaagc tgaaggtaat aaagtaatct ttactttgtc agaaccagct 540 ccacaattca agagtcttct atcatttgct aactttgttc cacaaaaaga agattttgtg 600 aaaaaagctg gtaaagatta tggtacttct tcagaaaaac aagtttactc tgggccttat 660 acagttaaag attggaatgg tacaagtggt actttcaaac ttgttaaaaa cactaaatac 720 tgggatgcta aaaacgttaa aactaaaaca atcaacgttc aaacagttaa gaaacctgat 780 actgctgtac aaatgtacaa acaaggtaaa cttgactatg caaacatttc aggaacttca 840 gcgatttaca atgctaacaa gaaaaataaa gatgttacac cagttttaga ggcaactaca 900 gcttatatgg tttataacca aactggtaaa gtagaaggtt tatcaaatac gaagattcgt 960 caagctctta atcttgcaac tgaccgtaaa ggtattgttt ctgcagcagt agatactggt 1020 tctaaaccag cgacagctat tgctccaaca ggacttgcaa aacttaaaaa tggtacagat 1080 ttaacaaaac atgttgctcc tggttactca tatgatgaga agaaggccgg agaactcttt 1140 aaagaaggtc ttaaagaact aggtaaagat agtcttaaag taacagtaac agcggatgct 1200 gatagtcctg ttgccaaagc gtcaattgac tatattaaag aaacttggga aaaagctctt 1260 cctggattaa ccgttgaaga aaaatttgta accttcaaac aacgtttaga agatacaaaa 1320 aatcaaaact ttgaagttgc acttgtactt tggggtggtg actatccaga aggttctact 1380 ttctatggct tgatgaaatc aggttcagct tataactacg gtaaatttac aaatgaaaca 1440 tttgataaag cctacgaaaa agctttaaca actgatgcac ttgatacaaa tgcagcagct 1500 gatgattata aagatggtga aaaagcactt tatgaagcgg ctaactataa tccaatttac 1560 ttccgtagtg gagaaggatt acaaaatcca agtatcaaag gcttaatccg taactcaact 1620 ggtttgaatg ttgacttcac ttacgcttac aaaaaataa 1659 <210> 15 <211> 1062 <212> DNA <213> Streptococcus parauberis <400> 15 atgaacaaga aaattattgg tcttggttta acttcagtag cattattaag tttagctgct 60 tgtggtaatc gtggtgcttc aaaatctgat agtaaagatg ctaaaactga tttgaaagca 120 gcgattgtaa ctgatactgg tggtgttgat gataaatcat tcaaccaatc agcttgggaa 180 ggtttggaag cttggggtaa agaaaatggc ctaaaaaaag gtgctggttt tgattacttc 240 caatctaaca gtgaatctga atatgctact aaccttgata ctgcagtatc aaatggttat 300 aaagttgttt atggtatcgg ttttgctcta aaagacgcaa ttgccaaagc agctggtgat 360 aatactgatg taaactttgt aattattgac gatgtaatcg aaggtaaaga caatgttgca 420 agcgtaactt tcgctgataa tgaagctgca tatcttgctg gtattgctgc tgctaaaact 480 acaaaaacta aagttgttgg ttttgtaggt ggtatgaaag gtacagttat cactcgtttt 540 gaaaaaggtt ttgaagctgg tgttaaatcg gttgacgact caatcaaagt taaagttgac 600 tatgctggat catttggtga tgcctctaaa gggaaagcaa ttgcagctgc ccaatatgct 660 ggtggtgctg atgtaattta tcaagctgct ggtggtactg gtgctggtgt ctttaacgaa 720 gctaagagta tcaatgaaaa acgctctgaa gatgaaaaag tttgggttgt cggtgttgac 780 cgtgaccaag aagctgaagg taaatacaca actaacgata aaaaagaatc taactttgta 840 ttagcttcat ctattaaaca agttggtaaa tcagttgaat taattaacaa acttgttgct 900 gataagaaat tccctggtgg aaaatcaact gtttatggtt tgaaagatgg tggtgttgac 960 cttgctacta caaacttaaa cgacgatgct ataaaagcag ttaaagaagc aaaagaaaaa 1020 ttatttctgg tgacatcaaa gttcctgaaa aataataact aa 1062 <210> 16 <211> 969 <212> DNA <213> Streptococcus parauberis <400> 16 atgaaatttg gaaaaaaact tggtttcttt gctttattaa tgtcattagt attagttcta 60 ggcgcatgtg gaaaaacagg cttaggtaac tcatcaggtt cttcttcaaa agaagtaact 120 aaaaaagctg ccaaagatct taaattaggt gtttcaattt caacaacaaa caacccatat 180 ttcgtatcaa tgaaagatgg tcttgattca tatgcaaaag acaaaaaagt tagcttaaaa 240 gttgctgatg cacaagatga tgcagcgcgc caagctgacg atattcaaaa ctttattagc 300 caaaatgttg atgctatctt aatcaaccca gttgactcaa aagctgttgt tacttcaatc 360 aaagcagcta actcagcaaa cattccagta atcttaattg accgtggtag tgaaggtggc 420 gacgttttaa caactgttgc ttctgataat gttgctgcag gtaaaatggc tgctgaattt 480 gccgttaaag aacttggtga aaaagctaaa acttttgaat tatcaggggt acctggagct 540 tctgcaactg tagaccgtgg taaaggtttt aatgatattg ctaaaacaaa acttgatatt 600 ctttcaagcc aatcagcaaa ctttgaccgt gctaaagcgt taaacacagc tcaaaatatg 660 atccaaggta acaaagatgt acaagttatc tttgctcaaa atgatgaaat ggcactaggt 720 gctgcacaag ctgctaaatc tgctggactt aaagatgtga tgattattgg tattgatgga 780 caaccagatg cacacgatgc cataaaaaaa ggtgatatca cagctacaat tgcacaacaa 840 ccagctaaaa tgggtgaaat tgcaatccaa gcagcaattg accactatca aggtaaaaaa 900 gttgaaaaat caacagtttc tccaatttac ttagttaaaa aagataacgt tgataaatac 960 aactggtaa 969 <210> 17 <211> 945 <212> DNA <213> Streptococcus parauberis <400> 17 atgaaaaaat ctaataagtt aattactggt tttattacac ttgcatcagt aatgacatta 60 gcagcatgta attcaacaaa tgacaatact aaactggttt caatgaaggg tgatacaatc 120 actgtaactg acttttataa tgaagcaaag acaacacaag ctgcacaaca atcaatgctc 180 agtttaattt tatcgcgtgt ttttgaaggc caatatggta aaaaagtatc tgataaagaa 240 gtagaaaagt catacaataa aactgctgaa caatatgggg catcattctc tgaagcactt 300 tcacaagctg gtatgacaac tgaaacatat aaaaaacaaa ttcgtacaac catgctagtc 360 acatatgcag ttaagcaaga agctgaaaaa gaattaacag atgctaatta caaaaaagcc 420 tatgataact acacacctaa gatgactact caagtcatta ctttagatga tgaagaaaaa 480 gctaaaaaag tattagaaga aacaaaagca gaaggtgcta atttcgaaga tattgctaaa 540 aagaatactg ttaatgaaaa gaaaaaagta gattatacct ttgattcgtc tgacaaaagc 600 ttgccaagtg atgttattaa agaagctggc aaattaaaag ctggtgaaaa atcagagtta 660 atttcagtaa tggacccttc tacttatcag aagaaatatt acattgttaa catggctaag 720 aaagaagata agaaagctga ttggaaagaa tataaaactc gtctaaaaga aattattttg 780 gatgaaaaaa acaatgatac tgctttccaa aacaaagtta tcgcaaaaga gttagataga 840 gcaaatgtca aaattaaaga taaagcattc tcaaatatct tatctcaatt tgcttctgat 900 aaacaatctg ctaatagtgc attgaaaaca tcagttgata aataa 945 <210> 18 <211> 1503 <212> DNA <213> Streptococcus parauberis <400> 18 atgaataaac gcgtaaaaat tgttgcaaca cttggtcctg cggtagaaat ccgtggtggt 60 aaaaaatacg gtgaagatgg ctactgggct ggcaaacttg atgttgaagc ttcagctaaa 120 aaaattgctg aattaatcga agctggtgca aatgttttcc gtttcaactt ttctcatggt 180 aatcaccaag agcaaggtga tcgtatggcg acagttcgcc gtgccgaaga aatggctcgt 240 caaaaagtag ctttcttatt ggatacaaaa ggtcctgaga tgagaacaga actatttgaa 300 gatggttcta aagagtttag ttatacaact ggtgaaacta ttcgtgttgc tactgaacaa 360 ggtatcaaat caactcgtga agttattgct ttgagtgtag ctggtaatct tgatatttat 420 gatgaagttg aagttggtaa tacaatcctt atcgatgatg gtaaacttgg cttaaaaatt 480 gttgaaaaag atatttcaac acgtcaattt gttactgtcg ttgaaaacga tggtttgatt 540 gctaatcaaa aaggtgttaa tattccaaac actaagattc ctttcccagc tttagctgaa 600 cgtgataatg ctgacattcg ttttggactt gaacaaggct taaactttat cgctatttca 660 tttgtgcgta ctgcaaaaga cgttgaagaa gttcgcgaaa tttgtcgtga aactggtaat 720 gaacatgtcc aattatttgc taaaatcgaa aaccaacaag gtattgataa tttagatgaa 780 attattgaag ctgctgatgg tatcatgatt gctcgtggag atatgggtat tgaagtacca 840 ttcgaaatgg ttccagtatt ccaaaaaatg attatcacta aagttaatgc tgcaggtaag 900 gcagttatta ctgcaacaaa catgcttgaa acaatgactg ataaacctcg tgcaactcgt 960 tcagaagttt cagacgtatt caatgctgtt attgatggta ctgatgcaac aatgttatca 1020 ggtgaatcag caaatggtaa atacccagta gaatcagtac gtacaatggc tacaatcgat 1080 aagaatgcac aaacattgct taacgaatat ggtcgtttag attcttctaa attcccacgt 1140 acaaataaaa cagatgttat tgcatcagca gttaaagatg ctacacattc aatggacatt 1200 aaattagttg taactattac tgaatcaggt tacactgctc gttcaatttc taaattccgt 1260 cctgatgcgg atatccttgc tattactttc gatgaaaaag ttcaaaaagc attgatgatc 1320 tattggggtg ttattccagt tcttgctact aaaccagcat caactgatga tatgtttgaa 1380 gttgctgaag aagttgcact taaagctggt cttgttaaat caggtgacaa cattgttatt 1440 gttgccggtg taccagttgg ttctggtgga actaacacaa tgcgtgttcg tactgttaaa 1500 taa 1503 <210> 19 <211> 1284 <212> DNA <213> Streptococcus parauberis <400> 19 atgtctacat catttgaaaa caaagctact aaccgtggcg ttattacatt tacaattggt 60 caagaacaaa ttaaaccagc tcttgatcaa gcatttaata aaattaaaaa agacctaaat 120 gtaccaggat tccgtaaagg tcacatgcct cgtccagtat tcaaccaaaa atttggtgaa 180 gaagttcttt atgaagatgc tttgaatatt gttttaccag ctacttatga agctgctgta 240 actgaattag gacttgaagt tgtagctcaa ccaaaaattg acgttgtttc tatggaaaaa 300 ggtcaagatt ggaaaattaa cgctgaagta gttacaaaac cagaagttaa acttggtgat 360 tataaaaatc ttgcagtaga agttgaagct tcaaaagaag ttagtgacga agaaatcgac 420 gaaaaaatcg aacgcgaacg taaaaacctt gctgaacttg ttgttaaaga agacgcagca 480 gcagaaggcg atacagttgt aattgacttc ttaggttcag ttgatggcgt tgaatttgac 540 ggtggtaaag gtgataactt ctcacttgaa cttggttcag gtcaattcat cccaggcttt 600 gaagaacaat tagttggtgt taaagctggt gaaactgttg acgttaatgt tactttccct 660 gaagaatatc aagcagaaga tttagctggt aaagatgcta aattcgtaac aactgttcat 720 gaagtaaaaa ctaaagaagt tccagaatta gatgatgaat tagctaaaga catcgatgaa 780 gaagttgaaa cactagatga attaaaagct aaatatcgta aagaacttga agcagctaaa 840 gaaattgctt atgatgatgc cgttgaaggg gctgcaattg aattagcagt tgaaaatgct 900 gaaatcgttg aattaccaga agaaatggtt catgatgaag tacatcgtgc tatgaatgaa 960 tttatgggta acatgcaacg tcaaggtatt tctccagaaa tgtactacca attaactgga 1020 acttctgaag atgaccttca taaacaatat gagtctgaag ctgacaaacg tgttaaaact 1080 aaccttgtta ttgaagcaat tgctaaagca gaaggatttg aagcaagcga cgaagaaatt 1140 gaaaaagaaa tcaatgatct tgcaactgag tacaatatgc cagttgagca agttcgcact 1200 cttctttcag ctgatatgct taaacatgat attgcaatga aaaaagcagt tgaagttatc 1260 acagaatcag caaaagtaaa ataa 1284 <210> 20 <211> 1308 <212> DNA <213> Streptococcus parauberis <400> 20 atgtcaatta ttactgatgt ttacgctcgc gaagtcttag actcacgcgg taacccaaca 60 cttgaagtag aagtttatac tgaatcaggt gctttcggac gtggaatggt tccttcagga 120 gcatcaactg gtgaacatga agcagttgaa ctacgtgacg gtgacaaatc tcgttataac 180 ggattaggaa ctgaaaaagc agttgacaac gtaaacaaca ttatcgctga agctatcatc 240 ggttttgatg tacgtgatca acaagctatc gaccgtgcta tgatcgctct tgacggaact 300 cctaacaaag gtaaattagg tgctaacgct attcttggtg tttctattgc cgtagctcgt 360 gctgctgcag actaccttga agtacctctt tacaactatc ttggcggatt taacactaaa 420 gtacttccaa ctccaatgat gaacatcatc aatggtggat ctcactcaga tgctccaatc 480 gctttccaag aattcatgat catgcctgta ggtgctacaa catttaaaga agctctacgt 540 tggggtgctg aagtattcca tgctcttaag aaaatcctta aagaacgtgg tcttgaaaca 600 gctgttggtg acgaaggtgg attcgcacct aaatttgatg gcactgaaga tggtgtagaa 660 actatcctta aagctatcga agctgctgga tatgaagctg gcgaaaacgg aatcatgatt 720 ggtttcgact gtgcgtcatc tgaattctat gataaagaac gtaaagtata cgactacgct 780 aaatttgaag gagaaggcgg agcagttcgt acttctgcag aacaaattga ttaccttgaa 840 gaattagtta acaaataccc aatcattact atcgaagatg gtatggatga aaatgactgg 900 gatggttgga aagctcttac tgaacgccta ggcggacgtg ttcaattagt tggtgacgac 960 ttcttcgtta caaacactga atacttagct cgtggtatca aagaagaagc tgctaactca 1020 atccttatca aagttaacca aatcggtact ttgacagaaa cttttgaagc tattgaaatg 1080 gctaaagaag ctggattcac tgcagttgta tctcaccgtt caggtgaaac tgaagattca 1140 acaatcgctg acatcgcagt tgcaactaac gctggacaaa tcaaaactgg ttcattatca 1200 cgtactgacc gtattgctaa atacaaccaa ttacttcgta tcgaagatca acttggtgaa 1260 gttgctcaat acaaaggtat caaatcattc tataacctaa aaaaataa 1308 <210> 21 <211> 1197 <212> DNA <213> Streptococcus parauberis <400> 21 atggcaaaag aaaaatacga tcgtagtaaa ccccacgtta acattggtac aatcggacac 60 gttgaccatg gtaaaactac tttaactgct gcaattacaa cagtacttgc acgtcgctta 120 ccaagctcag ttaaccaacc taaagactat gcttctatcg atgctgctcc agaagaacgc 180 gaacgcggaa tcacaatcaa cactgcacac gttgagtatg aaactgaatc acgtcactat 240 gcgcatatcg atgccccagg acacgcggac tatgttaaaa acatgatcac tggtgctgcc 300 caaatggacg gagctatcct tgtagttgct tcaactgacg gaccaatgcc acaaactcgt 360 gagcacatcc ttctttcacg tcaggttggt gttaaacacc ttatcgtctt catgaacaaa 420 gttgacttag ttgatgatga agaattgctt gaattagttg aaatggaaat tcgtgatctt 480 cttacagaat acgacttccc aggtgatgac ctaccagtta tccaaggttc agctcttaaa 540 gctcttgaag gcgatacagc acatgaagat atcatcatgg aattgatgaa aactgttgat 600 gaatatattc cagaaccaga acgcgacact gacaaaccat tacttcttcc agtcgaagat 660 gttttctcaa tcactggtcg tggtacagta gcttcaggac gtatcgaccg tggtacagtt 720 cgtgtcaacg atgaaatcga aatcgttggt cttaaagaag atactaaaaa agctgttgtt 780 actggtgttg aaatgttccg taaacaactt gacgaaggtc ttgcaggaga caacgtaggt 840 atccttcttc gtggtgttca acgtgacgaa atcgaacgtg gtcaagttat tgctaaacca 900 ggttcaatca acccacatac taaatttaaa ggtgaagtat atatcctttc taaagaagaa 960 ggtggacgtc atactccatt cttcaacaac taccgtccac aattctactt ccgtactact 1020 gacgtaacag gttctatcga acttccagca ggtacagaaa tggttatgcc tggtgataac 1080 gtggcaattc aagttgaatt gattcaccca atcgccgtag aacaaggtac tacattctca 1140 attcgtgagg gtggacgtac tgttggttca ggtatcgttt cagaaatcga agcttaa 1197 <210> 22 <211> 1233 <212> DNA <213> Streptococcus parauberis <400> 22 atgaatgaat ttgaagattt gctaaacagt gttagcgaag taaacccagg tgatgtagtc 60 actgcggaag ttttaacagt tgataacggt caagcaaacg ttgttattga tggtactggc 120 gttgaaggtg tgttaactct tcgcgaattg actaacgatc gtgatgctga tatcaacgat 180 tttgttaaag ctggcgacac tgttgaagtg cttgttcttc gtcaagttgt aggtaaagac 240 actgatacagi ttactttcct agtatctaaa aaacgtttag aagctcgcaa agcatgggac 300 aaacttgttg gtcgcgaagg tgaagttgtt actgttaaag gtacacgcgc tgttaaaggc 360 ggactttcag ttgaatttga aggattacgt ggatttatcc cagcttcaat gattgacact 420 cgtttcgtac gtaacactga aaaatttgtt ggtcaagaat ttgaagctaa aatcaaagaa 480 gttgacgcag cagaaaaccg tttcattttg tctcgtcgtg aagttattga agaagcagct 540 aaagaagctc gccaagaagt cttctctaaa ctttctgagg gttcagttgt aactggtaca 600 gttgctcgtt taacaagctt tggtgctttt atcgaccttg gtggtgttga tggacttgta 660 cacgtgactg aattatcaca cgaacgtaat gtttcaccta aatcagttgt ttctgttggt 720 gaagaagttg aagttaaagt actttcaatt gacgaagaag ctggacgtgt ttcactttct 780 cttaaagcta caacccctgg accatgggat ggtgttgaac aaaaacttgc tcaaggcgat 840 gtagttgaag gtaaagttaa acgtttaact gactttggtg ctttcgtaga agttttagct 900 ggaattgatg gacttgttca catttcacaa atttctcaca accgtgttga aaatccaaaa 960 gatgtacttt ctgtaggtca agaagttact gttaaagttc ttgaagttaa tgctgctgat 1020 gaacgtgtat cactttcaat caaagctctt gaagaacgtc ctgcacaagc tgaaggcgaa 1080 aacaactaca acaacaataa taacaataac aacaacagcg aacgccaatc acgtccacgt 1140 cgtccaaaac gtgaacaaaa acgtgattat gaattaccag aaactcaaac tggtttctct 1200 atggctgact tgttcggtga tattgaatta taa 1233 <210> 23 <211> 1236 <212> DNA <213> Streptococcus parauberis <400> 23 atgactgcta aatcaccgat tcatgtttat tctgaaattg ggaaacttaa aaaagtttta 60 ctacacagac ctggtaaaga aatcgaaaac ttgatgccag actatttaga aagacttttg 120 tttgatgata ttccttattt ggaggatgct cagaaagaac atgatgcgtt tgcagaagcg 180 ttaagaaatg aaggcgtaga agtactttac ttagaaaaat tggctgctga atctttaatt 240 aatgatgaag tccgtgaaca attcattgac gaatacattt cagaagctaa tatcagaggt 300 cgtgctacta aaattgcgat tcgtgaattc ttgatggcta tcaaagataa ccaagaattg 360 attgacaaaa caatggcagg agttcaaaaa tctgaacttc cagaaattgc tgctgaagaa 420 aaaggattga ctgaccttgt tgaatctaac tacccattcg caattgatcc aatgcctaac 480 ctttatttca ctcgcgatcc atttgcaaca attggtgaag gtgtatcact taaccacatg 540 ttctctgaaa cacgtaaccg cgaaacatta tacggaaaat atattttcac atatcaccca 600 gtttacggtg gtgaaaaagt tcctatggtt tatgaccgta gcgaaaacac tcgtatcgaa 660 ggtggagatg aacttgttct ttcaaaagat gttcttgctg taggtatttc acaacgtaca 720 gatgctgcat caattgaaaa attattggta aacatcttca aacaaaacat tggcttcaag 780 aaagtattgg catttgaatt tgctaacaac cgtaaattca tgcacttaga tacagtgttc 840 acaatggttg actatgataa attcactatt cacccagaaa ttgaaggcga cctacgtgtt 900 tactctgtaa catatgaaaa cgatgacctt aagattcacg aagaaaaagg tgatctagca 960 gaattacttg cagctaacct aggtgttgaa aaagttgaat tgattcgttg tggtggtgat 1020 aacttagttg ctgctggacg tgaacaatgg aacgatggtt caaacacatt gactatcgca 1080 ccaggtgttg tagtagtata taaccgtaac acaattacaa acgccatctt agaatctaaa 1140 ggtttacgtt tgattaaaat caacggaagt gaattagttc gcggtcgtgg tggacctcgt 1200 tgtatgtcaa tgccatttga acgtgaagat ctctaa 1236 <210> 24 <211> 732 <212> DNA <213> Streptococcus parauberis <400> 24 atgttaaaat tcctgattcc aacggccaag gaaatgaaag tgccaacaga aagttttcca 60 ccacttatcc ccaagttcag ccagccaatt attgcaatga tggcagagct gactttggag 120 gaacttgcct cagtttacaa aattaagcct gaagctgcgc aaaaagaatt aaatcgaatc 180 agggaaatcc accaaaatca ggccctaact tatcccgctt ataagctctt caatggctta 240 atgtaccgtt atattaaacg cgaaaacctc agtaaaaaag aggaaaacta tttaaccaaa 300 cggacttata ttacttcatc tttatatggt attatcccca tggatttccc aattgcagag 360 catcgacttg acttccaaac gaaaataaaa atcaatggtc aaaatttaaa aacttattgg 420 cgtccacgtt acgaccaatt tttatcacct gaggatgttt atatctcttt actttctagt 480 gaatttgagg atatattctc cacagataaa cgaaaactgt ggataagtat caccttttta 540 gaagaaaaag acggcattcc caaaagccat tccacaattt ccaaaaaagc tcgaggcgca 600 tttttaaccg cagccatgaa agcaaattgc caatcctcat cacagttaaa atccttgaaa 660 ttcaatggtt tcagctattt ttctgattta tctaatgctg atcattttgt ttatattaaa 720 aaagaggcct ag 732 <210> 25 <211> 933 <212> DNA <213> Streptococcus parauberis <400> 25 atgaagaaga aatttacttt tgctttgact gctgctttgt cattattcat gctatctgct 60 tgctcgacta gtggtcaaca ggcaaaaaaa gaagaaaaat taaatgttgt tgcaactaat 120 tctgttattg cagatatgac aaaaaatata gccggtaata aggttaattt gcatagtatt 180 gttccagttg gtcaagaccc ccatgagtat gaaccattac ctgaagatgt tgaaaaaaca 240 agttcggcgg atttgatttt ttacaatgga attaacttag aagatggtgg acatgcttgg 300 tttactaaat tgattaagaa tgccaaaaaa gaaaaaaata aagattattt tgcggtatca 360 gatggtattg atgttatcta tttagagggg caaaatgaaa aaggtaaaga agatcctcat 420 gcttggttaa atcttgaaaa tggtatgatt tattcaaaaa atattgctaa gcaattaatc 480 aagaaagatc ctaaaaataa aacttactat caatctcatt taaaaagtta tttggcaaaa 540 ctagataatt tagataaaga agctaaatct aagtttgata agataccagc agataaaaaa 600 atgattgtaa caagtgaagg ctgcttcaaa tatttctcta aagcctatga tgttccatca 660 gcatatattt gggaaattaa tacagaagaa gaaggaacac cgagtcaaat ttcaactctc 720 attgagaaat taaaaaagaa aaaattatca gctatatttg ttgaatctag tgttgatagt 780 cgacctatga aatctgtttc taaggatagt ggaattccaa tttattctga aatttttaca 840 gattctgttg ccaaaaaagg gaaacctggt gatagttact atgccatgat gaaatggaat 900 ttagataaga tttcagaagg tttatcgaaa taa 933 <210> 26 <211> 972 <212> DNA <213> Streptococcus parauberis <400> 26 atgaaattta aacaattatt ttttacagtc ggtgtattaa gcttattggt tatcttatca 60 gcttgtcgta aagaagcgac agaaaacaag ggacaagtta aagtagggat tattcaatat 120 gccaagcatc ctgctttaga tgcttcacga aaaggcttta tcgaggcact tagagatggt 180 ggttataaag aaggcaagaa cttacaaatt accagtaaaa atgctcaagg tgaacaagct 240 aatctccaaa ccatggttga ccaattaact ggtaagaatg atttgaattt tgccattgca 300 acaccggcag cacaggcact cttgactagt gatccagata cacccgccgt ttttactgcc 360 gttacagatc ccgttggagc aggtttagtt gattcaatgg aaaaacctgg taaaaacatg 420 accggttcaa ttgatgctgg aaatgttcca caacaagtcg ccttgttgct aaaagctgtt 480 ccgacttcta aaaaagtggg catcttttat aattccagtg aagtaaattc acagatgcaa 540 gctaaagaag caaaaaaagc tctgaagaaa aaaggaatta tagctgttga aaagacagtt 600 acaacaagta atgatgtgca acaagtgatg acaagtttag ttggtcaagt tgatgctgtg 660 tacttaccga ctgataatac agttgcatca acagcgatga caattggtca aattttaaaa 720 gaaaataaaa ccccggcaat tggcagtgac aaagcttacc ttgacgcagt actattcaca 780 tcaggcgttg attatgaagc gattggtagg caagccgggg aagaagcggt agcaattctc 840 aatggcaaaa aagcttcaac catcgccgtt gccaaaccag aaaaacctga actagctatt 900 aataaagaca tggctaaatc acttggatta aatcctaaag aacttgaaaa acaattagta 960 gaaagcaaat aa 972 <210> 27 <211> 44 <212> DNA <213> Artificial Sequence <220> <223> Streptococcus parauberis SP1 forward primer <400> 27 taagaaggag atatacatat gacaagttca aaaagcacta attg 44 <210> 28 <211> 43 <212> DNA <213> Artificial Sequence <220> <223> Streptococcus parauberis SP1 reverse primer <400> 28 gtggtggtgg tggtgctcga gttttttgta agcgtaagtg aag 43 <210> 29 <211> 40 <212> DNA <213> Artificial Sequence <220> <223> Streptococcus parauberis SP2 forward primer <400> 29 taagaaggag atatacatat gaacaagaaa attattggtc 40 <210> 30 <211> 40 <212> DNA <213> Artificial Sequence <220> <223> Streptococcus parauberis SP2 reverse primer <400> 30 gtggtggtgg tggtgctcga ggttattatt tttcaggaac 40 <210> 31 <211> 41 <212> DNA <213> Artificial Sequence <220> <223> Streptococcus parauberis SP3 forward primer <400> 31 taagaaggag atatacatat gaaatttgga aaaaaacttg g 41 <210> 32 <211> 41 <212> DNA <213> Artificial Sequence <220> <223> Streptococcus parauberis SP3 reverse primer <400> 32 gtggtggtgg tggtgctcga gccagttgta tttatcaacg t 41 <210> 33 <211> 44 <212> DNA <213> Artificial Sequence <220> <223> Streptococcus parauberis SP4 forward primer <400> 33 taagaaggag atatacatat gaaaaaatct aataagttaa ttac 44 <210> 34 <211> 44 <212> DNA <213> Artificial Sequence <220> <223> Streptococcus parauberis SP4 reverse primer <400> 34 gtggtggtgg tggtgctcga gtttatcaac tgatgttttc aatg 44 <210> 35 <211> 40 <212> DNA <213> Artificial Sequence <220> <223> Streptococcus parauberis SP5 forward primer <400> 35 taagaaggag atatacatat gaataaacgc gtaaaaattg 40 <210> 36 <211> 42 <212> DNA <213> Artificial Sequence <220> <223> Streptococcus parauberis SP5 reverse primer <400> 36 gtggtggtgg tggtgctcga gtttaacagt acgaacacgc at 42 <210> 37 <211> 41 <212> DNA <213> Artificial Sequence <220> <223> Streptococcus parauberis SP6 forward primer <400> 37 taagaaggag atatacatat gtctacatca tttgaaaaca a 41 <210> 38 <211> 41 <212> DNA <213> Artificial Sequence <220> <223> Streptococcus parauberis SP6 reverse primer <400> 38 gtggtggtgg tggtgctcga gttttacttt tgctgattct g 41 <210> 39 <211> 42 <212> DNA <213> Artificial Sequence <220> <223> Streptococcus parauberis SP7 forward primer <400> 39 taagaaggag atatacatat gtcaattatt actgatgttt ac 42 <210> 40 <211> 42 <212> DNA <213> Artificial Sequence <220> <223> Streptococcus parauberis SP7 reverse primer <400> 40 gtggtggtgg tggtgctcga gtttttttag gttatagaat ga 42 <210> 41 <211> 40 <212> DNA <213> Artificial Sequence <220> <223> Streptococcus parauberis SP8 forward primer <400> 41 taagaaggag atatacatat ggcaaaagaa aaatacgatc 40 <210> 42 <211> 44 <212> DNA <213> Artificial Sequence <220> <223> Streptococcus parauberis SP8 reverse primer <400> 42 gtggtggtgg tggtgctcga gagcttcgat ttctgaaacg atac 44 <210> 43 <211> 40 <212> DNA <213> Artificial Sequence <220> <223> Streptococcus parauberis SP9 forward primer <400> 43 taagaaggag atatacatat gaatgaattt gaagatttgc 40 <210> 44 <211> 40 <212> DNA <213> Artificial Sequence <220> <223> Streptococcus parauberis SP9 reverse primer <400> 44 gtggtggtgg tggtgctcga gtaattcaat atcaccgaac 40 <210> 45 <211> 40 <212> DNA <213> Artificial Sequence <220> <223> Streptococcus parauberis SP10 forward primer <400> 45 taagaaggag atatacatat gactgctaaa tcaccgattc 40 <210> 46 <211> 41 <212> DNA <213> Artificial Sequence <220> <223> Streptococcus parauberis SP10 reverse primer <400> 46 gtggtggtgg tggtgctcga ggagatcttc acgttcaaat g 41 <210> 47 <211> 41 <212> DNA <213> Artificial Sequence <220> <223> Streptococcus parauberis SP11 forward primer <400> 47 taagaaggag atatacatat gttaaaattc ctgattccaa c 41 <210> 48 <211> 42 <212> DNA <213> Artificial Sequence <220> <223> Streptococcus parauberis SP11 reverse primer <400> 48 gtggtggtgg tggtgctcga gggcctcttt tttaatataa ac 42 <210> 49 <211> 40 <212> DNA <213> Artificial Sequence <220> <223> Streptococcus parauberis SP12 forward primer <400> 49 taagaaggag atatacatat gaaaacaatt ggaaaatcag 40 <210> 50 <211> 39 <212> DNA <213> Artificial Sequence <220> <223> Streptococcus parauberis SP12 reverse primer <400> 50 gtggtggtgg tggtgctcga gttttttatc tatattgac 39 <210> 51 <211> 43 <212> DNA <213> Artificial Sequence <220> <223> Streptococcus parauberis SP13 forward primer <400> 51 taagaaggag atatacatat gaaatttaaa caattatttt tta 43 <210> 52 <211> 48 <212> DNA <213> Artificial Sequence <220> <223> Streptococcus parauberis SP13 reverse primer <400> 52 gtggtggtgg tggtgctcga gtttgctttc tactaattgt ttttcaag 48

Claims (12)

delete delete delete delete delete An antigen gene for Streptococcus parauberis type I and SEQ ID NO: 14 and Streptococcus parauberis type II, amplified with a primer set having the nucleotide sequences of SEQ ID NOS: 27 and 28, Preparing an expression vector containing the gene;
Culturing the host cell transformed with the expression vector; And
Method from the host cell producing a specific protein antigen of Streptococcus para Ube Russ (Streptococcus parauberis) I-type and p-Ube Russ Streptococcus (Streptococcus parauberis) Ⅱ type. delete 7. The method of claim 6 wherein the host cell is a process for preparing a specific antigen protein to which is characterized in that the Escherichia coli, Streptococcus para Ube Russ (Streptococcus parauberis) I-type and Streptococcus para Ube Russ (Streptococcus parauberis) Ⅱ type. delete delete delete delete

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