MXPA99007722A - Recombinant chimeric viruses and uses thereof - Google Patents

Abstract

This invention provides a recombinant herpesvirus of turkeys-Marek's disease virus chimera comprising a herpesvirus of turkeys unique long viral genome region and a Marek's disease virus unique short viral genome region.

Description

CHIMERIC VIRUSES RECOMMENDERS AND USES THEREOF This application claims the priority of the application of the United States of America Serial No. 08 / 804,372, filed on February 21, 1997. Throughout this application, several publications will be referred to by numbers Arabicas in parentheses. The complete citation of these publications can be found at the end of the specification, before the claims. The disclosure of these publications is considered in its entirety incorporated herein by reference, for a more complete description of the state of the art to which the invention belongs.

BACKGROUND OF THE INVENTION The ability to isolate DNA and clone these DNA isolates to give bacterial plasmids has greatly expanded the approaches available for the development of viral vaccines. The methods used to elaborate the. present invention involve modifying the cloned DNA sequences from various viral pathogens of animals, by insertions, deletions, single or multiple base changes, subsequent insertions of these modified sequences into the virus genome. A utility of the addition of a foreign sequence is achieved when the foreign sequence encodes a foreign protein that is expressed during the P1501 / 99MX viral infection of the animal. The resulting live virus can then be used as a vaccine to produce an immune response in a host animal and provide protection to the animal against the disease. A virus with these characteristics is known as a viral vector, since it becomes a living vector that will carry and express the foreign protein to the host animal. In effect, it becomes an elaborate administration system for the protein or foreign proteins. The application of recombinant DNA techniques to animal viruses has a relatively recent history. The first viruses that were genetically engineered were those with the smallest genomes. In the case of papovavirus, because these viruses are so small and can not accept much additional DNA, their use in genetic engineering has been like defective replicons. The expression of foreign genes from these viruses requires a wild type helper virus and is not limited to cell culture systems. For adenovirus, there is a small amount of non-essential DNA that can be replaced by foreign sequences. The only foreign DNA that appears to have been expressed in adenovirus refers to the T-antigen genes from papovavirus (Mansour, et al., Proc. Nati, Acad. Sci. US, 1985, Thummel, et al., Cell, 1983; Scolnick, et al., Cell, 1981; P1501 / 99MX Thummel, et al., Cell, 1981), and herpes simplex virus (HSV), to the thymidine kinase gene (Ha -Ahmed and Graham, J. of Virology, 1986). These publications do not identify the non-essential regions in HVT where the foreign DNA can be inserted, nor do they show how to achieve the expression of foreign genes in HVT, for example, which promoter sequence and termination sequence it uses. Another group of viruses that have been genetically engineered are poxviruses. One member of this group, vaccinia, has been the subject of extensive research on the expression of foreign genes. Poxviruses are viruses that contain large DNA that replicate in the cytoplasm of the infected cell. They have a structure that is unique in that they do not contain a capsid and is based on icosahedric symmetry or helical symmetry. Poxviruses are very likely to have evolved from bacteria-like organisms through loss of function and degeneration. Due in part to its uniqueness, the advances made in the genetic engineering of poxviruses can not be extrapolated directly to other viral systems, which include herpes viruses and HVT. Vaccinia recombinant virus constructions have been made in several laboratories, so that they express the following inserted foreign genes: P1501 / 99MX HSV thymidine kinase gene (Mackett, et al., Proc. Nati. Acad. Sci. USA, 1982; Panicali and Paolett, Proc. Nati. Acad. Sci. USA, 1982, Hepatitis B surface antigen (Paoletti , et al., Proc. Nati, Acad. Sci. USA, 1984, Smith et al., Nature, 1983), HSV glycoprotein D gene, hemagglutinin influenzae gene (Panicali, et al., Proc. Nati. Acad. Sci. USA, 1983; Smith, et al., Proc. Nati, Acad. Sci. USA,? 983), malaria antigen gene (Smith, et al., Science, 1984, and vesicular stomatitis) G glycoprotein gene ( Mackett, et al., Science, 1986.) The overall global characteristics of recombinant DNA work with vaccinia are similar to the techniques used for all viruses, especially in regard to the techniques in reference (Maniatis, et al., Molecular Cloning, 1982). However, in detail, the techniques for vaccinia are not applicable to herpes viruses and HVT. The usefulness of vaccinxa as a vaccine vector is in question due to its close relationship with human smallpox and its known pathogenicity in humans. Therefore, the use of HVT, host-specific herpes virus, is a better solution for poultry vaccination. Among the primate herpes viruses, only the HSV of humans and, to a limited extent, the herpes saimiri of monkeys, have been engineered to contain sequences of foreign DNA. He P1501 / 99MX first use of recombinant DNA to manipulate HSV involved the cloning of a piece of DNA from the LS junction region to give a single long region of HSV DNA, specifically within the thymidine kinase gene (Moccarski, et al. ., Cell, 1980). This insert was not a strange piece of DNA, but was a piece that occurs naturally in the DNA of the herpes virus, which doubled in another place in the genome. This piece of AD? it was not manipulated genetically to specifically express a protein and, therefore, this work did not involve the expression of the protein in the herpes virus. The following manipulation of HSV involved the creation of deletions in the virus genome by a combination of AD techniques? recombinant and the selection of thymidine kinase. Using this approach, the alpha-22 HSV gene has been canceled (Post, et al., Cell, 1981), and a sequence of 15,000 base pairs has been canceled. from the internal HSV repeat (Poffenberger, et al., Proc. Nati. Acad. Sci. USA, 1981). The following cases involve the insertion of the genes that code for the protein within the herpes virus: the insertion of the glycoprotein C of HSV within a naturally occurring deletion mutant of this gene in HSV (Gibson and Spear, J. of Virology, 1983); the insertion of HSV type 2 glycoprotein D into HSV type 1 P1501 / 99MX (Lee, et al., Proc. Nati. Acad. Sci. USA, 1982), without manipulation of the promoter sequences, since the gene is not "foreign"; the insertion of the hepatitis B surface antigen into the HSV under the control of the HSV ICP4 promoter (Shih, et al., Proc. Nati, Acad. Sci. USA, 1984); and the insertion of bovine growth hormone within the herpes virus saimiri with an SV40 promoter (the promoter did not work in this system and an endogenous upstream promoter was used to transcribe the gene) (Desrosiers, et al., 1984). Two additional extraneous genes (chicken ovalbumin gene and Epstein-Barr virus nuclear antigen) have been inserted into HSV (Arsenakis and Roizman, 1984), and pseudorabies virus X glycoprotein has been inserted into HSV (Post, et al., 1985). These cases of deletion or insertion of genes within the herpes virus demonstrate that it is possible to genetically manipulate herpes virus genomes by recombinant DNA techniques. The methods that have been used to insert genes involve homologous recombination between the viral DNA cloned in the plasmids and the purified viral DNA transfected within the same animal cell. However, the degree to which the location of the deletion and the insertion sites of the foreign genes can be generalized is not known from these previous studies.

P1501 / 99MX An object of the present invention is a vaccine for Marek's disease. The Marek's disease virus (MDV) is the causative agent of Marek's disease that encompasses bird paralysis, a common lymphoproliferative disease of chickens. The disease occurs most commonly in young chickens between 2 and 5 months of age. Prominent clinical signs are progressive paralysis of one or more of the extremities, lack of coordination due to paralysis of the legs, drooping of the limbs due to the involvement of the wing, and a crouched head position due to the involvement of the legs. neck muscles. In acute cases, it can result in severe depression. In the case of highly oncogenic tensions, there is bursal and thymic atrophy characteristics. In addition, there are lymphoid tumors that affect the gonads, lungs, liver, baso, kidneys and thymus (Mohanty and Dutta, 1981). Most chickens are vaccinated against MDV on the first day of birth to protect against MDV for life. Prior to the present invention, the main method of vaccination for MDV involved using strains of turkey herpes virus (HVT) that occurred naturally. It may be advantageous to incorporate other antigens to this vaccination on the first day of birth, but the efforts made to combine the vaccines P1501 / 99MX conventional have not proved satisfactory to date, due to competition and immunosuppression between pathogens. The multivalent HVT-based genetically engineered vaccines of this invention represent a novel way to simultaneously vaccinate against several different pathogens. For the first time, a recombinant HVT with a foreign gene inserted into a non-essential region of the HVT genome is revealed here. The types of genetic manipulation that has been carried out on these herpes viruses consists of cloning parts of the virus DNA to form I plasmids in bacteria, reconstituting the DNA of the virus while it is in the cloned state, so that the DNA contains deletions of some sequences, and in addition, by adding foreign DNA sequences, instead of deletions or at sites removed from the deletions. A foreign gene of interest that is used for insertion into the HVT genome can be obtained from any pathogenic organism of interest. Typically the gene of interest will be derived from pathogens that in poultry cause diseases that have an economic impact on the poultry industry. Genes can be derived from organisms from which vaccines already exist and due to the novel advantages of vector technology, vaccines derived from P1501 / 99MX HVT will be superior. Also, the gene of interest can be derived from pathogens from which there is currently no vaccine, but where there is a requirement for disease control. Typically, the gene of interest codes for immunogenic polypeptides of the pathogen, and may represent surface proteins, secreted proteins and structural proteins. A relevant avian pathogen that is targeted for the formation of HVT vectors is Infectious Laryngotracheitis Virus (ILTV). ILTV is a member of the family herpesviridiae and this pathogen causes an acute disease in chickens, which is characterized by respiratory depression, emission of choppy sounds and expectoration of bloody exudate. Viral replication is limited to airway cells, when the infection is in the trachea, it leads to tissue erosion and hemorrhage. In chickens, no drug has been effective in reducing the degree of lesion formation or in decreasing clinical signs. Vaccination of birds with various modified forms of ILT virus derived from cell subculture and / or tedious administration regimens has conferred acceptable protection in susceptible chickens. Due to the degree of attenuation of current ILT vaccines, care must be taken to ensure that the level P1501 / 99 X correct virus is maintained, enough to ** provide protection, but not enough to cause disease in the flock. An additional target of the approach of the HVT vectors is Ne castle's disease, an infectious, highly contagious and debilitating disease that is caused by the Ne castle disease virus (NDV for its acronym in English). NDV is a single-stranded RNA virus of the paramyxovirus family. The different NDV pathotypes (velogenic, mesogenic, lentogenic) differ in the severity of the disease, specificity and symptoms, but most of the types appear to infect the respiratory system and the nervous system. NDV infects mainly chickens, turkeys and other avian species. Vaccination has historically been used to prevent this disease, however due to the interference of maternal antibodies, the life term of the birds and the route of administration, the producer needs to adapt immunization protocols that adjust to their specific needs. . The therapeutic agent that is administered as a viral vector of the present invention must be a biological molecule that is a byproduct of the replication of swinepox virus. This limits the therapeutic agent in the first analysis either to DNA, RNA or protein. These are examples of agents P1501 / 99MX from which these classes of compounds in the form of antisense DNA, antisense RNA (S. Joshi, et al., J. of Virology, 1991), ribozymes (M. Wachsman, et al., J. of General Virology, 1989), suppressor tRNA (RA Bhat, et al., Nucleic Acids Research, 1989), interferon-inducing double-stranded RNA and several other examples of protein therapeutics, from hormones, for example insulin, from of lymphokines, for example interferons and interleukins, to natural opiates. The discoveries of these therapeutic agents and the elucidation of their structure and function does not make obvious the ability to use them in a viral vector administration system. Several genes that code for cytokines have been cloned in mammalian species. Very few cytokine genes have been cloned into bird species. The genes coding for type I and type II interferons have been cloned and characterized in several mammalian species. The gene for chicken interferon-a was cloned and the amino acid sequence was compared to that of the mammalian type I interphone and showed to have a sequence identity of 20-24% amino acids. ChIFN-a is not related to mammalian IFN-g (59, 65). The chicken interferon-g gene was cloned, and the amino acid sequence shares 15% identity with ChIFN-a, ChIFN-g is 35 and 32% identical to IFN-g, equine and Human P1501 / 99MX, respectively (62, 63, 66). A gene for duck interferon was cloned and is 50% identical at the amino acid level with ChIFN-a (64). Chicken has more than 20 interferon genes that have not been classified yet. It is not known if the cloned duck IFN should be classified as IFN-a, -b or -g. Aviary interferon activities are species specific. An avian interferon is not active in a different mammalian or bird species or in a different bird or mammalian cell line. Due to the lack of interferon activities between species and the low percentage of identity of the amino acid sequences between the interferons of different species, it is not obvious that genes from different species of birds or mammals can be cloned and expressed.

SUMMARY OF THE INVENTION This invention provides a recombinant herpes virus from the turkey Marek's disease virus chimera, which comprises a unique long turkey viral genome region and a unique short viral genome region of a virus of the genome. Marek's disease BRIEF DESCRIPTION OF THE FIGURES Figures 1A-1C: Detail of the HVT Construction and Map Data.

P1501 / 99MX Figure IA shows BamHI restriction fragment map of the HVT genome. The fragments are numbered in order of decreasing size; the letters refer to small fragments whose comparative size has not been determined. Figure IB shows the BamHl # 16 fragment of the HVT genome showing the location of the β-galactosidase gene insertion in S-HVT-001. Figure IC shows the BamHI fragment # 19 of the HVT genome showing the location of the β-galactosidase gene insertion. Legend: B = BamHl; X = Xhol; H = HindIII; P = PstI; S = I left; N = Ndel; R = EcoRI. Figure 2: BamHI, Notl restriction map of the HVT genome. Long unique regions are shown (UL) and single cut (US). The repeats of the long and short regions are indicated in boxes. The BaMHI fragment is numbered in descending order of size. The location of probes P1-P4 is indicated. The origin of each probe is as follows: Pl - BamHl # 6, P2 - BamHl # 2, P3 - BamHl # 13 and P4 - 4.0 kb BglII to sub fragment Stul of genomic Xbal fragment HVT # 5 (8.0 kb). Figures 3A-3B: Show how the unique Xhol site of the BamHI fragment # 10 of the HVT genome was converted to a PacI site and a NotI site by inserting the synthetic DNA sequence into the XhoI site. { Nucleotides # 1333-1338; SEQ ID NO. 12).

P1501 / 99MX Figure 3A shows the Xho site converted to a Pací site to generate Plasmid 654-45.1 (SEQ ID NO: 17) and Figure 3B shows the Xhol site converted to a Notl site to generate Plasmid 686-63. Al (SEQ ID No. 18). Figure 4: Restriction map and open reading frames of the sequence surrounding the insertion site within the single length of HVT (SEQ ID No. 12). This map shows the restriction site Xhol (SEQ ID No. 12; Nuci 1333-1338) used for the insertion of foreign genes. Four open reading frames are also shown within this sequence. The ORF (reading frame for its acronym in English) A is interrupted by the insertion of DNA within the Xhol site. The amino acid sequence of ORF A (SEQ ID No. 14; Nuci, 1402 to 602; 267 amino acids), does not show a significant sequence identity with any known amino acid sequence in the protein databases. UL 54 (SEQ ID No. 13; Nucl.146 to 481; 112 amino acids) and UL55 (SEQ ID No. 15; Nucl 1599 to 2135; 179 amino acids) show significant sequence identity with the UL54 and UL55 proteins of the virus of herpes simplex type I, respectively. The reading frame ORF B (SEQ ID No. 16; Nuci 2634 to 2308; 109 amino acids) does not show significant sequence identity with any of the amino acid sequences known in the database of P1501 / 99MX proteins. The NCBI databases were searched using the Blast software. Figure 5: Restriction map of cosmids 407-32.1C1, 672-01. A40, 672-07. C40 and 654-45.1. The overlap of the genomic DNA fragments of HVT EcoRI # 9 and BamHI # 10 is illustrated. A unique Xhol site within the EcoRI # 9 and BamHl # 10 fragments has become a unique PacI site in Plasmid 654-45.1 or a unique NotI site in Plasmid 686-63. Al. Figure 6: Expression of β-galactosidase from the chicken anemia virus promoter and the HCMV immediate early promoter in transient transfection assays in chicken embryo fibroblasts treated with BES. The expression of β-galactosidase was measured by the ONPG assay at 0, 20, 40 and 60 minutes and expressed as OD4Δ5. The protocol is described in TRANSITORY TRANSFER TEST. Plasmids 388-65.2 contain the immediate early promoter. Plasmid 850-80.2, 850-25.18 and 850-69.1 contains the chicken anemia virus promoters as described in Materials and Methods. Figure 7: DNA sequence of glycoprotein D (gD) of infectious laryngotracheitis virus (ILTV) which is useful for expression of foreign DNA in recombinant turkey herpesvirus or recombinant chimeric viral vaccine comprising a chimaera from the short region of disease virus P1501 / 99MX Marek and the long region of turkey herpesvirus (SEQ ID NO.). Figure 8: DNA sequence of glycoprotein I (gl) of infectious laryngotracheitis virus (ILTV) which is useful for the expression of foreign DNA in the recombinant herpesvirus of turkey or recombinant chimeric viral vaccine comprising a chimera from the region short of the Marek's disease virus and the long region of the turkey herpesvirus (SEQ ID NO.).

DETAILED DESCRIPTION OF THE INVENTION This invention provides a recombinant herpesvirus of Marek-Turkey disease virus chimera comprising a long viral genome region of turkey herpesvirus and a short viral genomic region of the Marek's disease virus. This invention provides a recombinant herpesvirus from Marek-Turkey disease virus quxmera comprising a long viral genomic region, unique, of turkeys herpesvirus and a short, unique viral genomic region of Marek's disease virus. In one embodiment, the foreign DNA sequence is inserted into a non-essential region of the viral genome of the Marek-turkey disease virus chimera, which is capable of being expressed in a host cell. In another embodiment, the foreign DNA sequence is inserted into an EcoRI fragment P1501 / 99 X # 9 of the single long region of the viral genome of the Marek's disease virus chimera - turkeys. In another mode, the above DNA sequence codes for a polypeptide. For example, the above DNA sequence can code for a cytokine, for example chicken millominocyte growth factor (cMGF), chicken interferon (cIFN) or quail interferon (qlFN). Alternatively, the foreign DNA sequence may code for an antigenic polypeptide selected from the group consisting of: Marek's disease virus, Newcastle disease virus, infectious laryngotracheitis virus, infectious bronchitis virus and infectious bursal disease virus. In another embodiment, the foreign DNA sequence is under the control of a herpesvirus promoter, upstream, endogenous. Examples of promoters include, but are not limited to: upstream heterologous promoters such as PRv gX promoters, HSV-l alpha 4 promoters, HCMV immediate early promoters, MDV gA promoters, MDV gB, MDV gD, ILT gB, BHV-1.1 VP8, ILT gD and chicken anemia virus promoter (CAV). Examples of recombinant chimeric viruses include, but are not limited to: S-HVT-145, S-HVY-149, and S-HVY-152. This invention provides a vector comprising the isolated nucleic acid molecule that P1501 / 99MX codes for chicken interferon. This invention provides a host cell that contains the vector. The invention provides a recombinant DNA comprising the complementary sequence of the isolated nucleic acid molecule encoding chicken interferon. The complementary sequence is the antisense strand. In one embodiment, the antisense nucleic acid molecule hybridizes to the chicken interferon mRNA. This invention provides a method for improving the immune response of an animal by vaccinating the animal with the recombinant virus comprising the foreign DNA. Examples of foreign DNAs include, but are not limited to: cytokines, such as, for example, the sense strand of the nucleic acid sequence coding for quail interferon and chicken interferon, and viruses, such as, for example, chicken anemia virus, MDV, NDV, ILT, or IBDV. Alternatively, it is contemplated that an isolated polypeptide that is expressed from the recombinant virus may be used in combination with an exterminated or attenuated virus vaccine, in order to improve the animal's immune response. This invention provides a method for improving the immune response of an animal by vaccination of the animal with a vaccine containing a vector comprising the complementary sequence of P1501 / 99MX the homosentide strand of the isolated nucleic acid molecule encoding chicken interferon. This invention provides a vector comprising the isolated nucleic acid molecule encoding chicken interferon. This invention provides an isolated nucleic acid comprising a promoter sequence normally associated with the transcription of a chicken anemia virus (CAV) gene. The invention provides an isolated nucleic acid encoding a chicken anemia virus promoter. In one embodiment, the isolated nucleic acid encoding a chicken anemia virus promoter has the nucleic acid sequence set forth in SEQ ID NO. 23. This invention provides a vector comprising the isolated nucleic acid encoding a chicken anemia virus promoter. The invention provides a recombinant herpesvirus comprising a foreign DNA sequence under the control of the chicken anemia virus promoter. The invention provides a recombinant herpes virus or recombinant pox virus comprising a foreign DNA sequence inserted into a non-essential site in the HVT genome, wherein the foreign DNA sequence is capable of being expressed in a host cell infected with the recombinant HVT. and his expression is under control P1501 / 99 X? OR of the chicken anemia virus promoter. The invention provides a chimeric virus comprising a foreign DNA sequence inserted into a non-essential site of the HVT genome. The foreign DNA sequence is capable of being expressed in a host cell infected with the recombinant chimeric virus and its expression is under the control of a promoter placed upstream of the foreign DNA sequence. As defined herein, "a non-essential site" in the genome of recombinant chimeric virus or HVT genome refers to a region in the viral genome that is not necessary for viral replication or infection. As defined herein, "viral genome" or "genomic DNA" refers to all the DNA that occurs naturally in the virus. As defined herein, "foreign DNA sequence" or "gene" refers to any DNA or gene that is exogenous to genomic DNA. As defined herein, an "open reading frame" is a DNA segment that contains codons that can be transcribed to RNA that can be translated into an amino acid sequence and that does not contain a stop codon. The invention further comprises several suitable insertion sites in the virus, either a HVT or MDV genome useful for constructing the recombinant chimeric virus of the present invention. The sites of P1501 / 99MX insert include the EcoRI # 9 fragment and the BamHl # 10 fragment of the HVT genome, a preferred insertion site within those fragments is an Xhol restriction endonuclease. This invention provides a recombinant chimeric virus comprising a sequence of Strange DNA inserted within the EcoRI # 9 fragment of the turkey viral genome herpesvirus and the foreign DNA sequence is capable of being expressed in a host cell infected with turkey herpesvirus. In one embodiment, the foreign DNA sequence is inserted into an AORFA open reading frame of the EcoRI # 9 fragment. The insertion of the foreign DNA sequences into the Xhol site of EcoRI # 9 interrupts the ORFA, indicating that the entire ORFA region is not essential for the replication of the recombinant. For the purposes of this invention "a recombinant chimeric virus" and "a turkey recombinant herpes virus" refers to live viruses that have been generated by recombination methods well known to those skilled in the art, for example the methods set forth in the TRANSFECTION OF DNA TO GENERATE RECOMBINANTS, in the section of material and method, and the virus has not had genetic material essential for the replication of the recombinant chimeric virus or the recombinant herpesvirus P1501 / 99MX turkey that has been removed or canceled. The purified recombinant chimera virus and the recombinant turkey herpesvirus result in an insertion of foreign DNA sequences or a gene in the EcoRI # 9 fragment or in the Ba Hl # 10 fragment. The invention further provides a recombinant chimeric virus wherein the foreign DNA sequence encodes a polypeptide that is antigenic in a mammal into which the recombinant chimeric virus is introduced. In one embodiment, the polypeptide is a detectable marker. For the purposes of this invention a "polypeptide that is a detectable marker" includes the dimeric, trimeric and tetrameric forms of the polypeptide. The E. coli B-galactosidase is a tetramer composed of four monomeric polypeptides or subunits. In one embodiment, the polypeptide is B-galactosidase from E. coli. The invention provides a recombinant turkey herpesvirus (HVT) comprising a foreign DNA sequence inserted into a non-essential site in the HVT genome. The foreign DNA sequence is capable of being expressed in a host cell infected with the recombinant HVT and if expression is under the control of a promoter placed upstream of the foreign DNA sequence. In another modality the strange DNA sequence P1501 / 99MX codes for a cytokine. In another embodiment, the cytokine is chicken myelomonocytic growth factor (cMGF), chicken interferon (cIFN) or quail interferon. In a preferred embodiment, the recombinant turkey herpesvirus is designated S-HVT-144 '. The invention also provides a herpesviru? turkey recombinant whose viral genome contains foreign DNA encoding an antigenic polypeptide derived from Marek's disease virus (MDV), Newcastle disease virus (NDV), infectious laryngotracheitis virus (ILTV), infectious bronchitis virus (IBV) ) or infectious bursal disease virus (IBDV). The invention provides a recombinant turkey herpesvirus with a foreign DNA sequence insertion in the EcoRI # 9 fragment, further comprising a foreign DNA sequence encoding the antigenic polypeptide selected from the group consisting of: Marek's disease virus, Newcastle disease virus, infectious laryngotracheitis virus, infectious bronchitis virus and infectious bursal disease virus. In one embodiment, the foreign DNA sequence encoding the antigenic polypeptide is derived from the Marek's disease virus and encodes the glycoprotein gA of Marek's disease virus, the glycoprotein gB of the human disease virus.

P1501 / 99MX Marek or glycoprotein gD of Marek's disease virus. In another embodiment of the foreign DNA sequencing coding for glycoprotein gA, glycoprotein gB or glycoprotein gD of Marek's disease virus are inserted into the unique Stul site of the US2 gene coding for the herpesviru region? of Turkey. The invention further provides turkey recombinant herpesvirus whose genomic DNA contains foreign DNA encoding antigenic polypeptide from the Marek's disease virus. Preferably, the antigenic polypeptide is glycoprotein gB, gA or gD of the Marek's disease virus. In a modality, a recombinant HVT containing a foreign DNA sequence codes for IBDV VP2, MDV gA, and MDV gB. Preferably, these recombinant viruses are designated S-HVT-137 and S-HVT-143. The present invention provides a recombinant chimeric virus that contains a foreign DNA sequence that codes for an antigenic polypeptide from the Newcastle disease virus (NDV). In this case, it is preferred that the antigenic polypeptide be a fusion protein (F) of the Newcastle disease virus or a hemagglutinin-neuraminide (HN) of the Newcastle disease virus or a recombinant protein P1501 / 99MX comprising an E. coli B-galactosidase fused with a hemaglutidine-neuraminidase (HN) of the Newcastle disease virus. The present invention also provides recombinant chimeric viruses engineered to contain one or more foreign DNA sequences encoding an antigenic MDV polypeptide as well as one or more foreign DNA sequences encoding an antigenic NDV polypeptide. Preferably, the MDV antigenic polypeptide is gB, gD or gA of MDV and for NDV it is F or HN. The invention further provides recombinant chimeric viruses whose genomic DNA contains foreign DNA encoding an antigenic polypeptide from the Marek's disease virus and further comprises foreign DNA encoding for antigenic polypeptide of Newcastle disease virus. In addition, in one embodiment, the foreign DNA sequence encodes the antigenic polypeptide from an infectious laryngotracheitis virus and codes for the infectious laryngotracheitis virus glycoprotein gB, infectious laryngotracheitis virus gl glycoprotein gD or infectious laryngotracheitis virus gD glycoprotein . In another embodiment, the foreign DNA sequence encodes an antigenic polypeptide that P1501 / 99MX is derived or derivable from a group consisting of: MDV, gA, MDV gB, MDV gD, NDV HN, NDV F, ILT gB, ILT gl, ILT gD, IBV, IBDV VP2, IBDV VP3, IBDV VP4 , avian encephalomyelitis virus, avian reovirus, avian paramyxovirus, avian influenza virus, avian adenovirus, avian poxvirus, avian coronavirus, avian rotavirus, chicken anemia virus (agent), salmonella spp. E. coli, Pasteurela spp., Bordetala spp., Eimeria spp., His tomonas spp., Trichomonas spp., Poultry nematodes, tapeworm, pterodactyls, pest mites / poultry, protozoa of poultry . In a preferred embodiment the turkey recombinant herpesvirus is designated S-HVT-136. The invention further provides a recombinant turkey herpesvirus containing a foreign DNA sequence encoding an antigenic polypeptide from infectious laryngotracheitis virus. It is preferred that the antigenic polypeptide be glycoprotein gB of ILTV, gD of ILTV or gl of ILTV. In one embodiment, the foreign DNA sequence comes from an infectious laryngotracheitis virus and codes for a gD glycoprotein, or a glycoprotein gl of infectious laryngotracheitis virus. The invention provides a turkey recombinant herpesvirus containing a sequence of P1501 / 99MX Foreign DNA inserted into the EcoRI # 9 fragment of the turkey herpesvirus viral genome, where the foreign DNA sequence comes from the Newcastle disease virus and codes for an HN of Newcastle disease virus or a virus F of Newcastle disease. This antigenic polypeptide can be derived or derivable from the following: feline pathogen, canine pathogen, equine pathogen, bovine pathogen, avian pathogen, porcine pathogen or human pathogen. In another embodiment, the antigenic polypeptide of a human pathogen is derived from human herpes virus, herpes simplex virus-1, herpes simplex virus-2, human cytomegalovirus, Epstein-Barr virus, Varicell-Zoster virus, human herpesvirus-6. , herpesviru? human-7, human influenza, human immunodeficiency virus, rabies virus, measles virus, hepatitis B virus and hepatitis C virus. In addition, the antigenic polypeptide of a human pathogen may be associated with malaria or malignant tumor from the group which consists of pl asmodi um falsiparum, Borde tella pertusi s, and malignant tumor. The invention further provides turkey recombinant herpes virus whose genomic DNA contains foreign DNA encoding the fusion protein (F) of the Newcastle disease virus and which further comprises a foreign DNA encoding a P1501 / 99MX recombinant protein, wherein the E. coli B-galactosidase is fused with the hemagglutinin-neuraminidase (HN) of the Newcastle disease virus. The invention further provides a recombinant chimeric virus whose genomic DNA contains foreign DNA encoding the glycoprotein gB of Marek's disease virus and for the gk glycoprotein of Marek's disease virus, and also comprises foreign DNA encoding the hemagglutinin-neuraminidase (NH) of the Newcastle disease virus. The invention provides a recombinant herpesvirus of Marek-turkey disease virus chimera, comprising a unique long viral genome region of turkey herpesvirus and a unique short region of Marek's disease virus. In one embodiment, the recombinant herpes virus from the Marek-turkey disease virus chimera contains a foreign DNA sequence inserted into the EcoRi # 9 fragment of the turkey herpesvirus viral genome and the foreign DNA sequence capable of being expressed in a Host cell infected with herpesviru? of Turkey. In one embodiment, the recombinant turkey herpesvirus contains a foreign DNA sequence encoding a polypeptide. The polypeptide can be antigenic in an animal into which the recombinant herpes virus is introduced.

P1501 / 99MX In another embodiment, the polypeptide is beta-galacto-idase from E. coli. In another embodiment, the foreign DNA sequence codes for a cytokine. In another embodiment, the cytokine is chicken millomonitoring factor (cMGF), pilon interferon (cIFN) or quail interferon. The invention further provides turkey recombinant herpesvirus wherein the foreign DNA sequence encoding a polypeptide that is antigenic in an animal within which the recombinant herpesvirus is introduced. In addition, the recombinant turkey herpesvirus further comprises a foreign DNA sequence encoding the antigenic polypeptide selected from the group consisting of: Marek's disease virus, Newcastle disease virus, infectious laryngotracheitis virus, infectious blonquitis virus and infectious bursal disease virus. The invention provides a herpesviru? turkey recombinant wherein the foreign DNA sequence is under the control of a hexogene upstream herpesvirus promoter. In one embodiment, the foreign DNA sequence is under the control of a heterologous promoter, upstream. In another embodiment, the promoter is selected from CAV promoter, PRV gX promoter, HSV-l alpha 4 promoter, HCMV immediate early promoter, MDV gA promoter, P1501 / 99MX promoter MDV gB, gD promoter, ILT gB promoter, BHV-1.1 VP8 promoter and ILT gD promoter. This invention provides a homology vector for producing a recombinant chimeric virus by inserting foreign DNA into the viral genome. Examples of homology vectors include: 301-07. YDl, 852-52,114, 864-74.18, 881-23.28, and 739-27.16. This invention provides a homology vector for producing a turkey recombinant herpesvirus by inserting foreign DNA into the viral genome of a turkey herpesvirus comprising a double-stranded DNA molecule consisting essentially of: a) a foreign double-stranded DNA normally not present within the viral genome of turkey herpesvirus b) at one end of the foreign DNA, a double-stranded DNA from turkey herpesvirus, homologous to the viral genome located on one side of the EcoRI # 9 site, the coding region of the viral genome of turkey herpesvirus; and c) at another end of the foreign DNA, a double-stranded DNA of turkey herpes virus, homologous to the viral genome located on the other side of the EcoRI # 9 fragment of the coding region of the turkey herpesvirus viral genome. Examples of homology vectors are designated 751-87.A8. In one embodiment, the polypeptide is antigenic in the animal within which it is introduced P1501 / 99MX the recombinant turkey herpesvirus. In another modality, the antigenic polypeptide comes from a cytokine, Marek's disease virus, Newcastle disease virus, infectious laryngotracheitis virus or infectious bronchitis virus. In a preferred embodiment, the antigenic polypeptide is a chicken milonium growth factor (cMGF) or chicken interferon (cIFN), quail interferon, infectious bursal disease virus polyprotein, infectious bursal disease virus VP2 protein, glycoprotein gB of Marek's disease virus, Marek's disease virus gA glycoprotein, Marek's disease virus gD glycoprotein, Newcastle disease virus fusion protein, Newcastle disease virus hemagglutinin-neuraminidase, gB glycoprotein infectious laryngotracheitis disease virus, gD glycoprotein of infectious laryngotracheitis disease virus, infectious bronchitis virus seed protein or infectious bronchitis virus matrix protein. In another embodiment, the double stranded foreign DNA sequence in the homology vector codes for an antigenic polypeptide derived from an equine pathogen. The antigenic polypeptide of an equine pathogen can be derived from equine influenza virus or equine herpes virus. Examples of this antigenic polypeptide are: virus neuraminidase P1501 / 99MX equine influenza type A / Alasca 91 neuraminidase from equine influenza virus type A / Prague 56, equine influenza virus neuraminidase type A / Miami 63, neuraminidase from equine influenza virus ^ type A / Kentucky 81, glycoprotein B from equine herpesvirus type 1 and equine herpesvirus glycoprotein D type 1. In another embodiment, the double stranded foreign DNA sequence of the homology vector codes for an antigenic polypeptide derived from bovine respiratory syncytial virus or bovine parainfluenza virus. The antigenic polypeptide derived from the equine pathogen of bovine respiratory syncytial virus that can be derived from equine influenza virus is bovxno respiratory syncytial virus binding protein (BRCV G), bovxno respiratory syncytial virus fusion protein (BRSV F), bovine respiratory syncytial virus nucleocapside (BRSV N), bovine parainfluenza virus type 3 fusion protein, and bovine parainfluenza virus type 3 haemagglutinin-neuraminidase. In another embodiment, the double stranded foreign DNA sequence in the homology vector it codes for a cytokine capable of stimulating the human immune response. For example, the cytokine can, inter alia, interleukin-1, interleukin-15, interferons, interferon P1501 / 99MX quail, chicken interferon, granulatory-macrophage colony stimulating factors and interleukin receptors. For the purposes of this invention, a "homology vector" is a plasmid constructed to insert foreign DNA into a specific site of the turkey herpesvirus genome. In one embodiment of the invention, the double-stranded DNA of the turkey herpesvirus is homologous to the DNA sequence present within the EcoRl fragment # 9 of the turkey herpesvirus genome. Preferably, this homology vector is designated 172-63.1. In another embodiment, the foreign DNA sequence codes for a classifiable marker. Examples of classifiable labels include, but are not limited to: E.coli B-galactosidase or E.coli B-glucuronidase. The invention further provides a vaccine comprising an effective immunizing amount of a herpesviru? of recombinant turkey of the present invention and a suitable carrier. The invention provides a vaccine useful for immunizing a bird against the Marek's disease virus comprising an effective immunizing amount of the recombinant turkey herpesvirus and a suitable carrier. The invention provides a useful vaccine P1501 / 99MX for immunizing a bird against the Newcastle disease virus comprising an effective immunising amount of the recombinant turkey herpesvirus and a suitable carrier. The invention provides a vaccine useful for immunizing a bird against infectious laryngotracheitis disease virus, comprising an effective immunizing amount of the recombinant turkey herpesvirus and a suitable carrier. The invention provides a vaccine useful for immunizing a bird against the infectious bronchitis disease virus, which comprises an effective immunising amount of herpesviru? of recombinant turkey and a suitable carrier. The invention provides a vaccine useful for immunizing a bird against the infectious bursal disease virus comprising an effective immunising amount of the recombinant turkey herpesvirus and a suitable carrier. The invention provides a multivalent vaccine for immunizing a bird against the Marek's disease virus and Newcastle disease virus, which comprises an effective immunizing amount of the recombinant turkey herpesvirus. The invention provides a multivalent vaccine useful for immunizing a bird against the Marek's disease virus and virus.

P1501 / 99MX infectious laryngotracheitis, comprising an effective immunising amount of the recombinant turkey herpesvirus and a suitable carrier. The invention provides a multivalent vaccine useful for immunizing a bird against the Marek's disease virus and infectious bronchitis virus, which comprises an effective immunizing amount of the recombinant turkey herpesvirus and a suitable carrier. The invention provides a multivalent vaccine useful for immunizing a bird against the Marek's disease virus and infectious bursal disease virus comprising an effective immunizing amount of the recombinant turkey herpesvirus and a suitable carrier. The present invention also provides a method for immunizing a flock. For the purposes of the invention, this includes immunizing a flock against infectious bursal disease virus, Marek's disease virus, Newcastle disease virus, infectious laryngotracheitis virus or infectious bronchitis virus. The method comprises administering to the flock an effective immunizing dose of the vaccine of the present invention. The vaccine can be administered by any of the methods well known to those skilled in the art, for example, intramuscularly, subcutaneously, intraperitoneally or intravenously. Alternatively, the P1501 / 99MX vaccine can be administered in intranasal or oral form. This invention provides a host cell infected with herpesviru? of recombinant turkey. In one embodiment, host cells is an avian cell. For the purposes of this invention, a "host cell" is a cell used to propagate a vector and its insert. The infection of the cell is achieved by methods well known to the expert? in this field, for example, as set forth in TRANSFECTION OF DNA BY GENERATION OF RECOMBINANT HERPESVIRUS, in the Materials and Methods section. The methods for constructing, selecting and purifying recombinant turkey herpesviruses are detailed below. This invention provides a method for distinguishing chickens or other poultry, which are vaccinated with the above vaccine, from those which are infected with a naturally occurring Marek's disease virus, wherein the method comprises analyzing samples from the body fluids of chickens or other birds, to determine the presence of gG glycoprotein and at least antigen normally expressed in chickens or other birds infected by a Marek's disease virus that occurs naturally, the presence of antigens normally expressed in infected chickens, but the absence of glycoprotein gG are P1501 / 99MX indicative of vaccination with the aforementioned vaccine and non-infection with Marek's disease virus that occurs naturally. This invention provides a recombinant turkey herpesvirus expressing foreign DNA sequence and is useful as a vaccine is mammalian or avian species, including but not limited to: chickens, turkeys, ducks, felines, canines, bovines, equines and primates, including man. This vaccine may contain inactivated or live recombinant viruses. For the purposes of this invention is the term "an effective immunizing amount" of herpesviru? The recombinant feline of the present invention falls within the range of 103 to 109 PFU / dose. In another embodiment, the immunizing amount e? from 105 to 107 PFU / dose. In a preferred embodiment the immunizing amount is 106 PFU / dose. The method comprises administering to the animal an effective immunizing dose of the vaccine of the present invention. The vaccine can be administered by any of the methods well known to those skilled in the art, for example intramuscularly, subcutaneously, intraperitoneally or intravenously. Alternatively, the vaccine may be administered intranasally or intraorally. Suitable carriers for the recombinant virus are well known to those skilled in the art and include, but are not limited to, P1501 / 99MX proteins, sugars, etc. An example of this suitable carrier is a physiologically balanced culture medium containing one or more stabilizing agents for example, hydrolyzed proteins, lactose, etc. Preferably, the live vaccine is created by taking tissue culture fluids and adding stabilizing agents such as stabilizing hydrolyzed proteins. Preferably, inactivated vaccines utilize tissue culture fluids directly after inactivation of the virus. This invention provides an isolated nucleic acid molecule encoding an Inferior Quail Type 1. In one embodiment, the isolated nucleic acid molecule encoding the Interferon Quail Type 1 has the nucleic acid sequence set forth in SEQ ID No. .31. In one embodiment, the isolated nucleic acid molecule is genomic DNA. In another embodiment, the isolated nucleic acid molecule is cDNA. In another embodiment the RNA is derived from the isolated nucleic acid molecule or is capable of hybridizing to the isolated nucleic acid molecule. By "nucleic acid sequence" reference is made to a polymer of one or two strands of deoxyribonucleotide or ribonucleotide bases, read from the 5 'end to the 3' end. This includes both P1501 / 99 X self-replicating plasmids as infectious polymers of DNA or RNA and non-functional DNA or RNA. It will be readily understood by those skilled in the art that when reference is made here to a particular listing, that reference is intended to include sequences that substantially correspond to the listing and its complement, including permission of minor sequencing errors, simple base changes, of lesions, substitutions, and the like, so that any variation in sequence corresponds to the nucleic acid sequence of the pathogenic organism or the disease marker to which the relevant sequence listing relates. The invention provides a nucleic acid molecule of at least 14 nucleotides capable of specifically hybridizing to the isolated nucleic acid molecule encoding the Interferon Quail Type 1. This invention provides a nucleic acid molecule of at least 14 nucleotides capable of of specifically hybridizing to a complementary sequence of the homosense strand of the isolated nucleic acid molecule encoding the Type 1 Quail Inferior. A complementary sequence is the antisense strand of the nucleic acid molecule encoding the Type 1 Quail Interferon. In one embodiment, the molecule is from 8 to 36 P1501 / 99MX nucleotides. In another embodiment, the molecule is 12 to 25 nucleotides. In another embodiment, the molecule is 14 nucleotides. In one modality the molecule is DNA. In another modality the molecule e? RNA The invention provides an antisense molecule capable of hybridizing to the isolated nucleic acid molecule. In one embodiment, the antisense molecule is DNA. In another embodiment, the antisense molecule is RNA. In another embodiment, the antisense molecule is a nucleic acid derivative (eg, DNA or RNA with a protein structure). The present invention extends to the preparation of antisense nucleic acids or fragments thereof or ribozyme which can be used to interfere with the expression of a polypeptide, either by masking the mRNA with an antisense nucleic acid or cleaving it with a ribozyme, respectively. In one embodiment, the antisense nucleic acid molecule is hybridized to the mRNA of a quail Interferon. The approaches that go to the DNA are in several categories. The nucleic acids can be designed to attach to the main groove of the duplex DNA to form a triple helical structure or "triplex". Alternatively, the inhibitory nucleic acids are designed to bind to regions of single-stranded DNA that result from the opening of duplex DNA during replication or P1501 / 99 X transcript. Most commonly, the inhibitory nucleic acids are designed to bind to the mRNA precursors or to the AR? M itself. Inhibitory nucleic acids are used to prevent maturation of pre-AR? M. The acid? Inhibitory nucleics can be designed to interfere with the processing, or splicing or translation of AR ?, The complementary strand of the isolated nucleic acid molecule is also called antisense strand. Finally, inhibitory nucleic acids can be used to induce chemical inactivation or hobby of the target or RA? M genes. Chemical inactivation can occur by the induction of cross-links between the inhibitory nucleic acid and the target nucleic acid within the cell. Other chemical modifications of nucleic acids? induced by inhibitory nucleic acids properly derivatized, may also be used. Very severe hybridization conditions are selected at about 5 ° C less than the thermal fusion point (Tm) of the specific sequence at a defined ionic strength and a defined pH. The Tm is the temperature (under the pH and the defined ionic strength?) At which 50% of the salt concentration is at least about 0.02 molar at pH 7 and the temperature is at least P1501 / 99MX approximately 60 ° C. Like other factors this can significantly affect the stringent conditions of hybridization, including, among others, composition and size of the bases of the complementary strands, presence of organic solvents, ie concentration of salts or formamide, and extension of lack of coupling between the bases, the combination of parameters is more important than the absolute measurement of any of them. For example, the strictest conditions could be achieved by overnight hybridization at approximately 68 ° C in a 6X SSC solution, the ambient temperature band with 6X SSC solution, followed by a wash at approximately 68 ° C in a 0.6X SSC solution. Hybridization with moderate stringent conditions can be achieved, for example, in the following manner: 1) pre-hybridizing filter with a solution of 3X SSC, 50% formamide, bofe 0.1 M Tris at pH 7.5, Denhardt 5X solution; 2) pre-hybridization at 37 ° C for 4 hours; 3) hybridization at 37 ° C with a quantity of probe labeled equal to 3,000,000 cpm in total for 16 hours; 4) washes in x SSC and 0.1% SDS solution; 5) wash 4 times for 1 minute each at room temperature n 4X SSC at 60 ° C for 30 minutes; on each occasion; and 6) dry and expose to film. The nucleic acid probe technology is P1501 / 99MX well known to the experts who will really appreciate that these probes can vary greatly with respect to length and can be labeled with a detectable label, for example a fluorescent dye or a radio isotope, to facilitate detection of the probe. The DNA probe molecules can be produced by inserting a DNA molecule having full length or by a fragment of the nucleic acid molecule isolated from DNA virus into suitable vectors, for example plasmids or bacteriophages, followed by transformation into suitable bacterial host cells, replication in transformed bacterial host cells and collection of DNA probes, using methods well known in the art. Alternatively, probes can be generated chemically from DNA synthesizers. The DNA probes can be generated by inserting the full length isolated nucleic acid molecule or a fragment thereof of the viru DNA, downstream of a bacteriophage promoter such as T3, T7 or SP6. Large quantities of RNA probe can be produced by incubating the labeled nucleotides with an isolated and linearized nucleic acid molecule of the DNA virus by its fragment, which contains an upstream promoter in the presence of an RA? suitable polymer P1501 / 99MX As defined herein, nucleic acid probes can be fragments of DNA or RNA. The DNA fragments can be prepared, for example, by digestion of the plasmid DNA or by the use of PCR, or they can be synthesized either by the phosphoramidite method described by Beaucage and Carruthers 1981, Tetrahedron Lett. 22, 1859-1862 or by the triester method according to Matteucci et al., 1981, Am. Chem Soc. 103: 3185. A double-stranded fragment can be obtained in this case, if desired, by annealing the single strand chemically synthesized together with suitable conditions or by synthesizing the complementary strand using the DNA polymerase with a suitable primer sequence. When a specific sequence for a nucleic acid probe is provided, it is understood that the complementary strand is also defined and included. The complementary strand will work equally well in situations where the target is a double-stranded nucleic acid. It is also understood that when the specific sequence is identified for use as a nucleic probe, a subsequence of the listed sequence that is 25 base pairs (bp) or more in length, is also encompassed to be used as a probe. The nucleic acid molecules of the present invention also include molecules that encode polypeptide analogs, fragments or P1501 / 99MX derived from antigenic polypeptides that differ from naturally occurring forms, in terms of the identity or location of one or more amino acid residues (deletion analogs containing less than all the residues specified for the polypeptide, analogs of substitution in which one or more specified residues are replaced by other residues and analogs of addition where one or more amino acid residues are added to a medial or terminal portion of the polypeptides) and which share some or all of the properties of the forms that They present in a natural way. The term "SSC" refers to a saline and citrate solution with 0.15M sodium chloride and 20mM sodium citrate. The solutions are usually expressed as multiples or fractions of this concentration. For example, 6XSSC refers to a solution having a concentration of sodium chloride and sodium citrate of 6 times that amount or 0. M sodium chloride and 120 mM sodium citrate. 0.2XSSC refers to a solution of 0.2 times the concentration of SSC or 0.03M of sodium chloride and 4mM of sodium citrate. The phrase "specifically hybridizing" describes a nucleic acid probe that hybridizes, duplexes or binds only a particular target DNA or RNA sequence, when the target sequences are present in a cellular DNA or RNA preparation P1501 / 99MX total. By "selectively hybridizing" it is meant that a probe binds to a specific target in a form that is detectable in a manner different from the non-target sequence, under fairly stringent conditions of hybridization. The phrase "nucleic acid molecule encoding" refers to a nucleic acid molecule that is directed to the expression of a specific polypeptide. The nucleic acid sequences include both the strand sequence of the DNA that is transcribed into the RNA and the strand of complementary DNA and the sequence of RNA that is translated into the protein. The nucleic acid molecule includes both the full length nucleic acid sequence and the sequences that are not full length. It is further understood that the sequence includes codons for degeneracy of the native sequence or sequences that can be introduced to provide codon preference in a specific host cell. This invention provides an isolated DNA operably linked to a transcription promoter RNA. The term "operably linked" as used herein refers to the binding of an upstream promoter from a DNA sequence, such that the promoter mediates the transcription of the DNA sequence. This invention provides a vector that P1501 / 99MX comprises the isolated nucleic acid molecule encoding the quail Interferon Type 1: The invention provides a recombinant DNA comprising the isolated nucleic acid molecule encoding the Type 1 quail Interferon. The invention provides a vector comprising the complementary sequence of the homosense strand of the isolated nucleic acid molecule encoding the Type 1 quail Interferon. This invention provides a recombinant DNA comprising the complementary sequence of the homosense strand of the isolated nucleic acid coding for the Type 1 quail Interferon. The vector includes, but is not limited to: a plasmid, a cosmid, a phage, an artificial yeast chromosome (YAC) or a recombinant virus containing the isolated nucleic acid. To obtain the vector, for example, the insert and the vector DNA can both be exposed to a restriction enzyme to close complementary ends in both molecules that form a base pair with each other and then bind together with the DNA ligase. Alternatively, the ligands can be ligated to the insert DNA corresponding to a restriction site in the vector DNA, which is then digested with the restriction enzyme that cuts at that site. Other means are available and are well known P1501 / 99MX for experts in this field. This invention provides a host cell that contains the vector. Suitable host cells include, but are not limited to, bacteria (e.g., E. coli), yeast, fungi, plant cells, insects, and mammal. Suitable animal cells include, but are not limited to: CEF, QT-35, ESK-4 Vero cells, HeLa cells, Cos cells, CV1 cells and several primary mammalian cells. The invention provides an isolated polypeptide having the biological activity of Type 1 quail Interferon. In another embodiment, the isolated polypeptide codes for the Inferior of Type 1 quail having the amino acid sequence set forth in SEQ. ID No. 32. The term "polypeptide", as used herein, refers to either the full-length gene product encoded by the nucleic acid or portions thereof. Thus "polypeptides" include not only the full-length protein but also the partial length fragments, including peptides less than 55 amino acid residues in length. In addition, the isolated polypeptide can be linked to a second polypeptide to form a fusion protein by joining the isolated nucleic acid molecule to a second nucleic acid molecule and performing P1501 / 99MX expression in a suitable host cell. In one embodiment, the second nucleic acid molecule encodes beta-galactosidase. Other nucleic acid molecules that are used to form a fusion protein are known to those skilled in the art. The invention provides an antibody that specifically binds to the polypeptide encoded by the isolated nucleic acid molecule. In one embodiment, the antibody is a monoclonal antibody. In another embodiment, the antibody recognizes an epitope of the polypeptide. In another embodiment, the antibody is a polyclonal antibody. In another embodiment, the antibody recognizes those of an epitope of the polypeptide. In another embodiment, the antibody is an anti-idiotypic antibody. An isolated antibody, polypeptide or nucleic acid molecule can be labeled with a detectable label including, but not limited to: radioactive label or a colorimetric, luminescent or fluorescent label, or gold. Radioactive tags include, but are not limited to: 3H, 1C, 32P, 33P; 35S, 3eCl, 51 Cr, "Co59, Co59, Fe90, Y135, 131I, I186, and Re [sic.] Fluorescent labels include, but are not limited to: fluorescein, rhodamine, and auramine. Colorimetric labels include, but are not limited to: biotin and digoxigenin.The methods to produce P1501 / 99MX polyclonal or monoclonal antibodies are known to those skilled in the art. In addition, the antibody, polypeptide or nucleic acid molecule can be detected by an aliquot that could be linked to an enzyme, for example alkaline phosphatase or horseradish peroxidase. Other enzymes that can be used are well known to those skilled in the art. The invention provides a method for producing a polypeptide encoded with the isolated nucleic acid molecule, which comprises culturing a host-neighbor system under suitable conditions that allow production of the polypeptide and recover polypeptide produced in this form. Suitable suspended cells distribute bacteria, yeast, filamentous fungi, plants, inserts and mammalian cells. Host-vector systems for producing and recovering a polypeptide are well known to those skilled in the art and include, but are not limited to: E. coli and pMAL (New England Biolabs), the Sf9-vaculovirus insect cell expression system, and mammalian cells (such as HeLa, COS, NIH 3T3 and HEK293) transfected with a mammalian expression vector by lipofectin (Gibco-BRL) or calcium phosphate precipitation or with other methods to achieve vector entry into the cell. The invention provides a method for P1501 / 99MX provide specific polypeptide regions encoded by the nucleic acid molecule isolated from the polypeptide to generate antibodies. The amino acid sequences can be analyzed by methods well known to those skilled in the art in order to determine whether they produce hydrophobic or hydrophilic regions of the polypeptides that are constructed. In the case of a cell membrane polypeptide, it is well known that the hydrophobic regions form the part of the polypeptide that is inserted within the bilayer of liquid within the cell membrane, while the hydrophilic regions are placed on the cell surface, a watery environment Normally, the hydrophilic regions will be more immunogenic than the hydrophobic regions. Therefore, the hydrophilic amino acid sequences can be selected and used to generate antibodies specific for the polypeptide encoded by the isolated nucleic acid molecule encoding the DNA of the virus. The selected peptides can be prepared using commercially available machines. As an alternative, the nucleic acid can be cloned and expressed and the resulting polypeptide can be recovered and used as an immunogen. In addition, enzymes can be used as labels. Suitable enzymes include alkaline phosphates, beta-galac osidase, glucose-6-phosphate P1501 / 99 X dehydrogenase, hydrogenase maleate and peroxidase. Two major types of enzyme immunoassays are the enzyme-linked immunosorbent assay (ELISA), and the homogeneous enzyme immunoassay, also known as enzyme-linked immunoassay (EMIT, Syva Corporation, Palo Alto, CA). In the ELISA system, separation can be achieved, for example, by the use of antibodies coupled to a solid phase. The EMIT system depends on the deactivation of the enzyme in the tracer-antibody complex; the activity is therefore measured without the need for a separation step. Additionally, the chemiluminescent compounds? They can be used as labels. The compounds? chemiluminescent? they include luminol, isoluminol, aromatic acridinium esters, imidazoles, acridinium salts and oxalate esters. Similarly, bioluminescent compounds can be used for labeling, bioluminescent compounds include luciferin, luciferase, and aequorin. A description of a radioimmunoassay (RIA) may be found in: Labora tory Techniques in Biology and Biological Processing (1978) North Holland Publishing Company, New York, with particular reference to the article entitled "An Introduction to Radioimmune Assay and Related Techniques "by T. Chard. A general description of the trials P1501 / 99MX general immunometric of the various types may be found in the following United States patents: United Nos. 4,376,110 (David et al.) Or 4,098,876 (Piasio) The invention provides a recombinant virus comprising a foreign DNA inserted into a non-essential region of a viral genome that is capable of being expressed in a host cell, wherein Strange DNA codes for quarantine interferon Type I. The virus is selected from the group consisting of: turkey herpesvirus, swine virus, pseudorabies virus, infectious bovine rhinotracheitis virus, equine herpes virus, herpesviru? feline, poultry pustulation virus, infectious laryngotracheitis virus, Mareck's disease virus, poxvirus, rash (pox) in canary, eruption (pox) in raccoons, vaccinia, adeno-associated virus, adenovirus, canine herpesvirus, infectious bursal disease virus, herpes simplex virus and alpha virus. This invention provides a vaccine comprising an effective immunizing amount of the Type I quaternary interferon polypeptide and a pharmaceutically acceptable carrier. Vaccines can be administered by any conventional method for administration of vaccines, including oral or injection P1501 / 99MX parenterally (for example, subcutaneous or intramuscular). Intramuscular administration is preferred. The treatment may consist of a single dose of vaccine or a plurality of doses over a period of time. It is preferred that the dose administered to a human patient be from the first 8 months of life. The antigen of the injection can be combined with dosi? Suitable for compounds that include antigen? of influenza, for example antigens of influenza type A. Also, the antigen could be a component of the recombinant vaccine that could be adapted for oral administration. The vaccines of the invention can be combined with other vaccines for other diseases in order to produce multivalent vaccines. A pharmaceutically effective amount of the antigen may be employed with a pharmaceutically acceptable carrier, for example a protein or diluent useful for vaccination of mammals, particularly humans. Other vaccines may be prepared according to methods well known to those experts. The experts? they will readily recognize that it is only necessary to suppose a mammal to the appropriate epitopes in order to produce effective immunoprotection. Epitopes are typically segments of amino acids that are a small portion of the total protein. Using recombinant genetics, it is routine to alter a primary structure of P1501 / 99MX natural protein to create derivatives that encompass epitopes that are identical or substantially equal (immunologically equivalent) to epitopes that occur naturally. These derivatives may include peptide fragments, amino acid substitutions, amino acid lesions and amino acid additions in the amino acid sequence for the polypeptide. For example, it is known in the art of proteins that certain amino acid residues can be substituted with amino acids of similar size and polarity, without undue effect on the biological activity of the protein. The invention provides a method for culturing a recombinant virus to a high titer by growing the recombinant virus in a cell line containing the complementary sequence of the homosense strand of the isolated nucleic acid molecule coding for quaternary interferon, which is express. In another embodiment, the virus is an exterminated or attenuated virus. In one embodiment, the nucleic acid e? the strand homosentido. In another embodiment, the nucleic acid is the antisense strand. The invention provides a method for improving the replication of a recombinant virus. In one embodiment, the virus is killed or attenuated. The invention provides a method for inhibiting the IFN production of an avian species. In a P1501 / 99MX modality, the nucleic acid is the strand homosentido. In another embodiment, the nucleic acid is the antisense strand. The invention provides a transgenic avian species that expresses a quail interferon. Avian species include, but are not limited to: quail, turkey, chicken, geese, ducks, ostriches, pigeons or birds in general. This invention provides a method for improving the immune response of an animal by vaccinating the animal with a vaccine containing a vector or a plasmid comprising the isolated nucleic acid molecule encoding quail interferon. The invention provides a method for improving the immune response of an animal by vaccinating the animal with a vaccine containing a vector or plasmid comprising the complementary sequence of the homosense strand of the isolated nucleic acid molecule encoding the interferon quail. It is contemplated by this invention that the vector may be administered concurrently, subsequently, or subsequently to the recombinant virus that is used to vaccinate the animal. The invention provides a method for increasing the weight of an animal, which comprises vaccinating the animal of avian species with a vaccine comprising the isolated nucleic acid molecule.

P1501 / 99MX codes for quail interferon. In one embodiment, the nucleic acid is the homosense strand. In another modality the nucleic acid is the antisense strand. This invention is further illustrated in the following section of Experimental Details. This section is set forth to aid in understanding the invention but is not intended to limit the invention and should not be construed to limit the invention in any way, as set forth in the appended claims.

EXPERIMENTAL DETAILS: Materials and Methods: _ _ t __ _ _ PREPARATION OF TURKEY HERPESVIRUS FROM EXISTING SAMPLES. Existing samples of turkey herpesvirus were prepared by infecting tissue culture cells at a multiplicity of infection of 0.01 PFU / cell in the Dulbecco Modified Eagle Medium (DMEM) containing 2 mM glutamine, 100 units / ml penicillin, 100 units / ml of streptomycin (these components are obtained from Irvine Scientific or an equivalent supplier, and hereinafter referred to as complete DME medium), plus 1% fetal bovine serum. After the effect was realized, the medium and cells were harvested and P1501 / 99MX cells were pelleted at 3000 rpm for 5 minutes in a clinical centrifuge. The infected cells were resuspended in complete medium containing 20% fetal bovine serum, 10% DMSO and stored frozen at -70 ° C.

PREPARATION OF THE DNA OF THE HERPES VIRUS OF TURKEY. All manipulations of turkey herpes virus (HVT) were made using strain FC-126 (ATCC # 584-C). For the preparation of HVT viral DNA from the cytoplasm of infected cells, the primary chicken embryo fibroblasts were infected at a sufficient MOI (muI iplici ty of infection) to cause a profound cytoplasmic effect before cell overgrowth. All incubations were carried out at 39 ° C in a humidified incubation with 5% C02 in air. The best DNA yields were obtained by collecting monolayers that were maximally infected, but showed incomplete cell lysis (typically 5-7 days?). The infected cells were harvested by scraping the cells into the medium, using a cellular scraper (Costar brand). The cell suspension was centrifuged at 3000 rpm for 10 minutes at 5 ° C in a GS-3 rotor (Sorvall Instruments). The resulting pellet was resuspended in PBS (20 ml / revolving bottle) and subjected to another centrifugation for 10 minutes at 3000 rpm cold. After decapting the P1501 / 99MX PBS, the cell pellet was resuspended in 4 ml / rotating bottle of RSB buffer (lOmM tris pH 7.5, 1 mM EDTA and 1.5 mM MgC12). NP40 (Nonidet P-40 ™; Sigma) was added to the sample at a final concentration of 0.5%, minutes with occasional mixing. The mixture was centrifuged for 10 minutes at 3000 rpm cold to form a pellet of the nuclei and remove the cell debris. The supernatant fluid was carefully transferred to a 15 ml Corex centrifuge tube. EDTA (0.5M pH 8.0) and SDS (sodium dodecyl sulfate, 20% existence solution) were added to final concentrations of 5mM and 1%, respectively. 100 μl of proteinase-K (10 mg / ml, Boehringer Mannheim) were added per ml of sample, mixed and incubated at 45 ° C for 1-2 hours. After this period, an equal volume of water-saturated phenol was added to the sample and mixed gently by hand. The sample was centrifuged in a clinical centrifuge for 5 minutes at 3000 rpm to separate the phases. NaAc was added to the aqueous concentration to a final concentration of 0.3M (storage solution 3M pH 5.2) and the nucleic acid was precipitated at -70 ° C for 30 minutes after the addition of .25 volumes of cold absolute ethanol. The DNA in the sample was pelleted by centrifugation for 20 minutes at 8000 rpm in a HB-4 rotor at 5 ° C. The supernatant was carefully removed and the DNA pellet was washed once with P1501 / 99MX 25 ml of 80% ethanol. The DNA pellet was briefly dried under vacuum (2-3 minutes) and resuspended in 50 μl / revolving bottle of infected TE buffer cells (10 mM Tris pH 7.5, 1 M EDTA). Typically, viral DNA yields ranged from 5-10 μg / revolving bottle of infected cells. All viral DNA was stored at approximately 10 ° C.

REACTION OF FILLING WITH POLYMERASE. The DNA was resuspended in buffer containing 50 mM Tris pH 7.4, 50 mM KCl, 5 mM MgCl2 and 400 micromolar, each of the four deoxynucleotides. Ten units of Klenow DNA polymerase (BLR) were added and the reaction was allowed to proceed for 15 minutes at room temperature. The DNA was extracted with phenol and precipitated with ethanol, as above.

DNA SEQUENCING. Sequencing was focused using the USB Secuenase case and '35S-dATP (NEN) The reactions using the two dGTP mixtures and the dITP mixture were carried out to clarify the compression areas. Alternatively, the compressed areas were resolved on formamide gels. The templates were subclones of double-stranded plasmid or single-stranded M13 subclones, and the primers elaborated either for the vector just outside the insert to be sequenced or for the P1501 / 99MX sequence previously obtained. The obtained sequence was assembled and compared using the Dnastar software. The manipulation and comparison of obtained sequences was done with the Superclone and Supersee programs of Coral Software.

BIOLOGICAL TO MOLECULAR TECHNIQUES. Techniques for the manipulation of bacteria and DNA, including procedures such as digestion with restriction endonucleases, gel electrophoresis, extraction of DNA from gels, binding, phosphorylation with kinase, treatment with phosphatase, bacterial culture, transformation of bacteria with DNA and Other methods of molecular biology are described in Maniatis et al (1982) and Sambrook et al (1989). The polymerase chain reaction (PCR) was used to introduce suitable restriction sites for the manipulation of various DNA. The procedures used are described in Innis et al (1990). In general, the amplified fragments were less than 500 base pairs in size and critical regions of amplified fragments were confirmed by DNA sequencing. Except for the observed, these techniques were used with less variation.

SOUTHERN TRANSFERS OF DNA. The general Southern blot procedure was taken from Maniatis et al. (1982). The DNA was transferred to P1501 / 99MX nitrocellulose filters (S &BA85) in 20X SSC (IX ?? C = 0.15M NaCl, 0.015M sodium citrate, pH 7.0), and prehybridized in hybridization solution consisting of 30% formamide, IX of Denhardt's solution (0.02% polyvinylpyrrolidone (PVP), 0.02% bovine serum albumin (BSA), 0.02% Ficoll), 6X SSC, 50 mM NaH2P04, pH 6.8, 200 μg / ml salmon sperm DNA, for 4-24 hours at 55 ° C. The labeled probe DNA was added, once it had been labeled by nick translation using a kit from Bethesda Research Laboratories (BRL) and a 32P-labeled nucleotide. The DNA probe was separated from unincorporated nucleotides by the NACS column (BRL) or a Sephadex G50 column (Pharmacia). After hybridization overnight at 55 ° C, filtered was washed once with 2X SSC at room temperature followed by two washes with 0. IX SSC, 0.1% sodium dodecyl sulfate (SDS) for 30 minutes at 55 ° C. The filter was dried and subjected to autoradiography.

PROCEDURE OF DNA CLONING. The cDNA cloning refers to the methods used to convert the RNA molecules into DNA molecules following the procedures of the state of the art. The methods of the applicant are described in (Glubler and Hoffman, 1983). Bethesda Research Laboratories (Gaithersburg, MD) has designed a cDNA cloning kit that is very similar to P1501 / 99MX procedures used by applicants and contains a set of reagents and protocol that can be used to duplicate our results. To clone virus mRNA species, a host cell line responsive to virus infection was infected at 5-10 plaque-forming units per cell. When the cytopathic effect was evident, but before total destruction, the medium was removed and the cells lysed in 10 ml of lysis buffer (4 M guanidine thiocinate, 0.1% antifoam A, 25 mM sodium citrate pH 7.0 , n-lauroyl sarcosine 0.5%, beta-metcaptoethanol 0.1 M). The cell lysate was poured over a sterilized Dounce homogenizer and homogenized on ice 8-10 times until the solution was homogeneous. For the purification of RNA, 8 ml layers of cell lysate were gently formed on 3.5 ml of CsCl solution (5.7 M CaCl, 25 mM sodium citrate, pH 7.0) in a Beckman S 41 centrifuge tube. The samples were centrifuged for 18 hours at 20 ° C at 35,000 rpm in a Beckman SW41 rotor. The tubes were placed on ice and the supernatants of the tubes were carefully removed by aspiration to leave the RNA pellet undisturbed. The pellets were resuspended in 400 μl of distilled water and 2.6 ml of guanidine solution (7.5 m guanidine-HCl, 25 mM sodium citrate pH 7.0, 5 mM dithiothrethol) and added. 0.37 volumes of 1 M acetic acid were added, followed by 0.75 P1501 / 99MX volumes of cold ethanol to the sample was set at -20 ° C for 1 hour to precipitate the RNA. The precipitate was collected by centrifugation in a Sorvall centrifuge for 10 minutes at 4 ° C, at 10,000 rpm in an SS34 rotor. The pellet was dissolved in 1.0 ml of distilled water, recentrifuged at 13,000 rpm and the supernatant was saved. The RNA was re-extracted from the pellet twice as previously, with 0.5 ml of destined water and the supernatants were pooled. A volume of 0.1 of 2 M potassium acetate solution was added to the sample, followed by two volumes of cold ethanol and the sample was set at -20 ° C for 18 hours. The precipitated RNA was collected by centrifugation in the SS34 rotor at 4 ° C for 10 minutes, at 10,000 rpm. The pellet was dissolved in 1 ml of distilled water and the concentration was taken up by absorption at A260 / 280. The RNA was stored at -70 ° C. The mRNA containing polyadenylate (poly-A) tails was selected using oligo-dT cellulose (Pharmacia # 27 5543-0). Three mg of the total RNA were boiled and cooled and applied to the 100 mg oligo-dT cellulose column in binding buffer (0.1 M Tris pH 7.5, 0.5 M LiCl, 5 mM EDTA pH 8.0, 0.1% dodecyl lithium sulfate). The poly-A RNA retained was eluted from the column with elution buffer (5mM Tris pH 7.5, 1mM EDTA pH 8.0, 0.1% sodium dodecyl sulfate): This mRNA was reapplied to an oligo-dT column in binding buffer and eluted from P1501 / 99MX new in elution buffer. The sample was precipitated with 200 mM sodium acetate and 2 volumes of cold ethanol at -20 ° C for 18 hours. The RNA was resuspended in 50 μl of distilled water. Ten μg of poly-A RNA were denatured in mM of methyl mercury hydroxide for 6 minutes at 22 ° C. β-mercaptoethanol was added at 75 mM and the sample was incubated for 5 minutes at 22 ° C. the reaction mixture for cDNA synthesis of the first strand in 0.25 ml contained 1 μg of oligo-dT primer (PL Bio-chemicals) or a 1 μg of synthetic primer, 28 units of placental ribonucleic inhibitor (Bethesda Research Labs # 5518SA), 100 M Tris pH 8.3, 140 mM KCl, 10 mM MgCl2, 0.8 mM dATP, dCTP, dGTP and dTTP (Pharmacia), 100 microcuries of dCTP labeled with 32p (New England Nuclear # NEG-013H) and 180 units of AMV reverse transcriptase (Molecular Genetics Resources #MG 101). The reaction was incubated at 42 ° C per 90 minutes, and then it was finished with 20mM EDTA pH 8.0. The sample was extracted with an equal volume of phenol / chloroform (1: 1) and precipitated with 2M ammonium acetate and 2 volumes of ethanol frXo -2Q ° C for 3 hours. After precipitation and centrifugation, the pellet was dissolved in 100 μl of distilled water. The sample was placed on a Sephadez G-100 15 ml column (Pharmacia) in buffer (100 mM Tris pH 7.5, 1 mM EDTA pH 8.0, 100 mM NaCl). The leading edge of the diluted DNA fractions was pooled and the DNA was P1501 / 99MX concentrated by lyophilization until the volume was approximately 100 μl, then the DNA was precipitated with ammonium acetate plus ethanol, as before. The whole sample of the first strand was used for the reaction of the second strand that followed the method Gubler and Hoffman (1983) except that 50 μg / ml of dNTP, 5.4 units of DNA polymerase I (Boerhinger Mannheim # 642-711) were used. ), and 100 units / ml of E. coli DNA ligase (New England Biolabs # 205) in a total volume of 509 microliters. After the synthesis of the second strand, the cDNA was extracted with phenol / chloroform and precipitated. The DNA was resuspended in 10 μl of distilled water, treated with one μg of RNase for 10 minutes at 22 ° C, and electrophoresed through 1% agarose gel (Sigma Type II agarose). in 40 mM Tris-acetate pH 6.85. The gel was stained with ethidium bromide and the DNA in the expected size range was extracted from the gel and eluted in 8 mM Tris-acetate pH 6.85. The electroeluted DNA was lyophilized at approximately 100 microliters and precipitated with ammonium acetate and ethanol as before. The DNA was resuspended in 20 μl of water. The oligo-dC tails were added to the DNA to facilitate cloning. The reaction contained DNA, 100 mM potassium cacodylate pH 7.2.0.2 mM dithiothreitol, 2 mM CaC12, 80 μmoles d dCTP and P1501 / 99MX units deosinucleo tidi 1 terminal transferase (Molecular Genetic Resources # S1001) in 50 μl. After 30 minutes at 37 ° C, the reaction was terminated with 10 mM EDTA, and the sample was extracted with phenol / chloroform and precipitated as before. The DNA sample with DC tail was fixed to 200 ng of the plasmid vector pBR322 which contained oligo-dG tails (Bethesda Research Labs # 5355 SA / SB) in 200 μl of 0.01 M Tris pH 7.5, 0.1 M NaCl, 1 mM EDTA pH 8.0 at 65 ° C for 2 minutes and then 57 ° C for 2 hours. Competent fresh DH-1 cells of E. coli were prepared and transformed as described in Hanahan (1983) using half of the cDNA sample fixed in twenty 200 μl aliquots of the cells. The transformed cells were placed on L-broth agar plates plus 10 μg / ml tetracycline. The colonies were screened for the presence of inserts within the ampicillin gene using Ampscreen (Bethesda Researchs Labs # 5537 AU), and the positive colonies were chosen for further analysis.

TRANSFECTION OF DNA TO GENERATE HERPES RECOMBINANT VIRUSES. The method is based on the polybrene-DMSO procedure of Kawai and Nishizawa (1984) with the following modifications. Generation of recombinant HVT viruses depends on homologous recombination between viral HVT DNA and the plasmid homology vector P1501 / 99MX containing the desired foreign DNA flanked by the cloned sequences of suitable herpesviruses. Transfections were carried out in 6 cm plates (Corning plastic) of primary and 50% confluent chicken embryo fibroblast cells (CEF). The cells were plated the previous day in growth medium CEF (IX FIO / 199, 5% fetal calf serum, 2% glutamine, 1% non-essential amino acids and 2% penicillin / streptomycin) containing 4 μg / ml of polybrene (storage solution 4 mg / ml in IX HBSS). For cotransfections to the CEF cells, 5 μg of intact HVT DNA was used and suspended in 1 ml of CEF medium containing 30 μg / ml polybrene (storage solution 4 mg / ml in IX HBSS). The DNA polybrene suspension (1 ml) was subsequently added to a 6 cm dish of CEF cell from which a medium had been aspirated and incubated at 39 ° C for 30 minutes. The plates moved oscillating periodically during this time to redistribute the inoculum. After this period, 4 ml of the CEF growth medium was added directly to the washing plate and incubated for an additional 2.5 hours at 39 ° C. At this time, the medium was removed from each plate and the cells were subjected to shock with 2 ml of 30% DMSO (dimethyl sulfoxide, J.T. Baker Chemical Col), in IX HBSS for 4 minutes at room temperature. The 30% DMSO was carefully removed and the monolayers washed one P1501 / 99MX once with IX HBSS at room temperature. The cells were subsequently incubated at 39 ° C after the addition of 5 ml of the CEF growth medium. The next day, the medium was changed to remove the last possible traces of DMSO and to stimulate cell growth. The cytopathic effect of the virus is evident within six days. The generation of a high-grade warehouse solution (80% -90% CPE) can usually be done within a signal as of this date. HVT stock samples were prepared by resuspending the infected cells in growth medium CEF containing 20% calf serum, 10% DMSO and stored at least -70 ° C.

PROCEDURE TO GENERATE HERPES RECOMBINANT VIRUS FROM SUBGENOMICAL DNA FRAGMENTS. The ability to generate herperviruses by -cotransfection of cloned overlapping subgenomic fragments has been demonstrated for the pseudorabies virus (Zijl et al., 1988). If lesions and / or insertions are manipulated directly into the genomic fragments before co-transfection, this procedure results in a high frequency of viruses containing genomic alterations, greatly reducing the amount of analysis required to purify the recombinant virus. . This procedure was used to construct the recombinant HVT.

P1501 / 99MX A library of subclones containing subgenomic fragments of overlapping HVT was generated in the following manner. The HVT DNA was obtained from the American Type Culture Collection (FC-126 ("Calnek"). It was cut and then selected in size on a glycerol gradient as described by van Zijl et al., (1988) with fragments. of 40-50 kb selected as the insert population.The pooled fractions were doubled with TE, one tenth volume of 3 M NaAc and 2.5 volume of ethanol were added and the DNA was precipitated at 30K rpm in a Beckman SW rotor 41 for one hour.The cut fragments were given blunt ends by an initial treatment with T4 DNA polymerase, using low concentrations of DNTP to try to promote removal of the 3 'overhang, followed by treatment with Klenow polymerase to fill the ends. These fragments of insert were then ligated with a cosmid vector pWE15 (Stratagene) which had been digested with BamHI, treated with calf intestinal phosphatase and blunted with polymerase treatment to Klenow The bound mixture was then packaged using packaging extracts Gigapack XL (Stratagene). The binding and packaging were made as recommended by the manufacturer. Restriction maps published for the enzymes BamHl, HindIII and Xhol allowed the use of P1501 / 99MX fragments subcloned as probes specific for the analysis of the cosmid library for subclone extending from the genome. The probes were generated from subcloned restriction fragments. The fragments were subsequently labeled using a non-radioactive system (Genius, Boehringer Mannheim). Classification was facilitated by choosing the middle colonies and then cultivating them throughout the night. Five-filter sets and a master plate were stamped from the microtitre plate and cultured again throughout the night. Glycerol was added to the cavities at 15% and the plates were frozen at -20 ° C to provide storage culture for each colony. The filters were BioRad Colony Lift membranes and were treated and hybridized with the manufacturer's instructions and washed in 0. IX SSC, 0.1% SDS, 65 ° C. The clones that were hybridized with the non-reactive probe were detected according to the directions of the Genius kit. The colonies were selected for further analysis based on their hybridization in one or more of the specific tests. They were then digested with BamHl, and compared with published HVT maps (Buckmaster et al., 1988). The three cosmids (407-32.2C3, 407-32, IG7, and 407-32.5G6) were obtained in this way. A detailed description of each clone? E provides below. It was found that the P1501 / 99MX amplification of chloramphenicol (Maniatis et al., 1982) was necessary to achieve reasonable DNA yields from these clones. In addition, a cosmid clone (407-32.5G6) was unstable and tube to be cultured from frozen original stock material in order to obtain satisfactory DNA preparations. The vector pWEl5 allows the inserts to be extracted with Notl. However, four Notl sites are present in the HVT genome, so that the inserts that expand these sites can not be extracted with Notl. Two of the Notl sites are present in the BamHl # 2 fragment of HVT, this fragment was directly cloned into pSP64. The other two sites are present in the unique short region within the BamHI # 1 fragment. This fragment was cloned directly into the pWEl5 vector. The three cut cosmids and the two BamHl fragments cover the entire HVT genome except for a small portion of the ends. As these regions are repeated in the inner portions of the genome, all genetic information is available. A Stul site within US2 of HVT was established as a useful site for the insertion of foreign DNA using the HOMOLOGA RECOMBINATION PROCEDURE FOR GENERATING RECOMBINANT HERPESVIRUS (see Example 6). The US2 gene of HVT is located within the BamHl # 1 fragment that contains P1501 / 99MX five? Stul. To facilitate the use of this site for the insertion of foreign DNA by the Stul site within the US2 gene, it was converted to a unique HindIII site. This was achieved by partial digestion of subclone BamHl # 1 with Stul, and then by inserting a 10 kb fragment that confers resistance to kanomycin (NeoR) within the site using HindIII linkers. The kanomycin resistance gene allowed positive selection of the recombinant clones. The NeoR fragment was removed by digestion with HindIII followed by rearing generating the clone 430-84.215. The DNA was prepared by reconstruction experiments by restriction digestion with enzymes that cut out the outer subclones or that falter to the HVT insertions: In some cases, a cosmid in a reconstruction was used undigested. The DNAs were extracted once with phenol and precipitated with ethanol. The DNA was resuspended at a concentration of 0.5 to 1 μg / ml. Viral reconstruction experiments were performed using Lipofectin (BRL) to mediate transfection. Two to three micrograms of each subclone were added to 0.5 ml of MEM medium (Earle salts) supplemented with 1% non-essential amino acids and 2% penicillin / streptomycin (MEMt-). The controls consisted of MEM + without DNA, with several ug of HVT DNA, or with 4 of 5 of the subclones. Separately, 30 μl of Lipofectin were added to 0.5 others P 15 01/9 911X ml of MEM +. These two mixtures were combined and then incubated at room temperature for 15 minutes. Chicken embryo fibroblast cells (CEF) were prepared for transfection in the following manner. The CEFs (Spafas) were grown in plates of 6 cavities at 39 ° C in medium F10 / M199 (1: 1) containing 1% non-essential amino acids, 2% penicillin / streptomycin, and 5% fetal calf serum (CEF +). The cells were transfected to a confluence of 90-95%. For transfection, the cavities were aspirated and rinsed three times with MEM +, and then incubated 4 hours? at 39 ° C with 1 ml of lipofectin / previous DNA mixture. One more ml of CEF + was then added to the wells and the cells were incubated overnight and fed with CEF +. The plates were then examined daily to determine the appearance of plaques. Lipofectin with control HVT DNA resulted in the appearance of plaques within 5 days. When only four of the five subclones were used, no plates were obtained. When the five overlapping genomic fragments of _HVT were used to reconstruct the virus, plaques appeared everywhere between 5 and 19 days after the initial lipofection. In the case of late onset plaques, the plaques were not initially observed in the infected monolayer and it was only P1501 / 99MX after subculture of the monolayer and reseeding in plates on a larger surface, the plates appeared. After the subculture, the plates generally appeared within 3 days. The recombinant viruses that were plaque purified were approximately three and then analyzed for foreign DNA insertion.

BUOGAL SCREENING OR CPRG SCREENING FOR RECOMBINANT HERPESVIRUS When the foreign gene encoded β-galactosidase, the plates containing the gene were visualized more easily. The BluogalTM chemical agent (Bethesda Research Labs) for blue plates was incorporated at the 200-300 μg / ml level into the agarose array during the plate assay, and the plates that expressed active β-galactosidase changed to blue. The chemical agent CPRG (Chlorophenol Red Galactopyranoside, Boehringer Mannheim) for the red plates was incorporated into the agarose array during plaque analysis and the plates that expressed β-galactosidase changed to red. The blue or red plates were collected and purified by additional blue or red plate isolations. Other foreign genes were inserted by homologous recombination so as to replace the β-galactosidase gene; in this case the non-blue plaques were collected for purification of the recombinant virus.P1501 / 99MX SCREENING OF THE EXPRESSION OF THE STRANGE GENE IN HVT RECOMBINANT UTILIZADO NO BLACK PLATE ANALYSIS.

To analyze the expression of foreign antigens expressed by the recombinant HVT viruses, the monolayers of the CEF cells were infected with recombinant HVT, placed on nutrient agarose medium and incubated for 4-5 days at 39 ° C. Once the plates had developed, the agarose arrangement was removed from the plate, the monolayer was rinsed with Ix PBS, fixed with 100% methanol for 10 minutes at room temperature and the cells were air dried. After rehydration of the plate with PBS, the primary antibody was diluted to the appropriate dilution with PBS and incubated with the cell monolayer for 2 hours overnight at room temperature. The unbound antibody was removed from the cells by washing three times with PBS at room temperature. The antibody conjugated with alkaline phosphatase was diluted with PBS and incubated with the cells for 2 hours at room temperature. The unbound secondary antibody was removed by washing the cells three times at room temperature. Then, the monolayer was rinsed in a color developer buffer (100mM Tris pH 9.5 / 100mM NaCl / 5mM MgCl 2), and then incubated for 10 minutes overnight at room temperature with freshly prepared substrate solution (0.3 mg / ml of Blue Nitro P1501 / 99MX tetrazolium + 0.15 mg / ml 5-bromo-4-chloro-3 -indolyl phosphatase in color development buffer). Finally, the reaction was stopped by placing the substrate solution with TE (10 mM Tris, pH 7.5 / 1 mM EDTA). Plates expressing the correct antigen will be stained black.

PRA PLATE HYBRIDIZATION PROCEDURE DETERMINE THE PURITY OF RECOMBINANT HVT STORAGE MATERIAL. When there is no suitable immunological reagent to detect the presence of a particular antigen in a recombinant HVT virus, plaque hybridization can be used to assess the purity of a stock material. Initially, the monolayers of CEF cells are infected with several dilutions of the viral store materials to give approximately 50-100 plates / 10 cm. plate, superimposed on the nutrient agarose medium and incubated for 4-5 days at 39 ° C. Once the development of the plate is presented, the position of each plate is marked on the bottom of the plate. The agarose arrangement is then removed, the plate is washed with PBS and the remaining CEF monolayer is transferred to an NC membrane or to a BioRad nylon membrane pre-moistened with PBS (noting the position of the membrane relative to the plate). The cells contained on the NC membranes are then lysed by placing the membranes in 1.5 ml of NaCl P1501 / 99MX 1.5 M and 0.5 M NaOH for five minutes. The membranes were neutralized by placing them in 1.5 ml of 3M sodium acetate (pH 5.2) for five minutes. The DNA of the lysed cells is then bound to the NE membranes by baking at 80 ° C for one hour. After this period, the membranes are prehybridized in a solution containing 6X SSC, 3% skim milk, 0.5% SDS, (+ _) salmon sperm DNA (50 μl / ml) for one hour at 65 ° C. The radiolabeled probe DNA (alpha 32P-dCTP) is added and the membranes are incubated at 65 ° C overnight (-12 hours). After hybridization, the NE membranes are washed twice (30 minutes each) with 2X SSC at 65 ° C, followed by two additional 65 ° C washes with 0.5X SSC. The NC membranes are then dried and exposed to an X-ray film (Kodak X-OMAT, AR) at -70 ° C for 12 hours. The positive signals are then aligned with the position of the plates on the plate and the purity of the storage material is recorded as the percentage of positive plates on the plate.

CONSTRUCTION OF THE VECTOR OF HOMOLOGY FOR THE INSERTION OF THE ß-GALACTOSIDASE GENE WITHIN THE US2 GENE OF HVT. The ß-galac tosidase (lacZ) gene was inserted into the Ecol # 7 fragment of HVT at the unique Stul site. The marker gene is oriented in the same direction as the US2 gene. It is built P1501 / 99 X using standard recombinant DNA techniques (Maniatis et al, 1982 and Sambrook et al, 1989) binding the restriction fragments from the following sources to the DNA sequences and techniques. Fragment one is a restriction subfragment of approximately 413 base pairs of salt I to BamHI of the restriction fragment of BamHI of PruV, 10 (Lomniczi et al., 1984) Fragment 2 is a restriction fragment of approximately 3010 pairs of BamHl to PvuII bases of plasmid pJF751 (Ferrari et al., 1985). Fragment 3 is a restriction subfragment of approximately 754 base pairs of Ndel to Sali from restriction fragment # 7 of PRV BamHl (Lomniczi et al., 1984).

RNA ISOLATED FROM CONCANAVALIN AND STIMULATED CHICKEN SPLEEN CELLS: Chicken spleens were dissected from three-week-old chickens from SPAFAS, Inc., washed and disintegrated through a syringe / needle to release the cells . After allowing the stroma and the residues to settle, the cells were pelleted and washed twice with PBS. The cell pellet? E deal with a buffer? hypotonic to lyse the erythrocyte cells and the splenocytes were recovered and washed twice with PBS. Splenocytes were resuspended at 5 x 106 cells / ml in RPMl P1501 / 99MX containing 5% FBS and 5 μg / ml of Concanavalin A and incubated at 39 ° for 48 hours ?. Total RNA was isolated from the cells using guanidine isothionate lysis reagents and Promega RNA isolation kit protocols (Promega Corporation, Madison Wl). 4 μg of the total RNA was used in each reaction of the first strand containing the appropriate antisense primers and the AMV reverse transcriptase (Promega Corporation, Madison Wl). The cDNA synthesis was carried out in the same tube after the reverse transcriptase reaction, using the appropriate homosense primers and the Vent® DNA polymerase (Life Technologies, Inc. Bethesda, MD).

RECOMBINATION PROCEDURES HOMOLOGA TO GENERATE SPV OR RECOMBINANT FPV. This method is based on the homologous recombination between FPV DNA and SPV and the plasmid homology vector DNA that occurs in the tissue culture cells that contain the two FPV or SPV DNA and the homology vector. of transfected plasmid. For homologous recombination to occur, monolayers of CEF cells were infected with S-FPV 0D1 (a mild poultrypox vaccine strain, available from Bio-Pox ™ from Agri-Bio Corporation, Gainsville, Georgia) or SPV- 001 (Kasza layer) at a multiplicity of infection of 0.01 PFU / cell for P1501 / 99MX introduce replicating FPV (ie DNA synthesis) or SPV within the cells. The plasmid homology vector DNA is then transfected? in these cells according to "Infection-Transfection Procedure".

PROCEDIMIETO OF INFECTION-TRANSFECTION. CEF cells in 6 cm plates (approximately 80% confluent) were infected with S-FPV-001 at a multiplicity of infection of 0.01 PFU / cell in the negative medium CEF were incubated at 37 ° C in a humidified incubator with 5% of C02 for five hours. ESK-4 cells, CEF cells in 6-centimeter plates (approximately 80% confluent) were infected with S-SPV-001 at a multiplicity of infection of 0.01 PFU / cell in the negative CEF medium and incubated at 37 ° C in a humidified incubator with 5% C02 for 5 hours. The transfection procedure used is essentially that recommended by Lipofectin ™ (BRL). Briefly, for each 6 cm plate, 15 micrograms of plasmid DNA was diluted to 100 microliters with H20. Separately, 50 micrograms of Lipofectin ™ reagent was diluted to 100 microliters with H20. THE? 100 microliter? of diluted Lipofectin ™ reagent were added dropwise to the diluted plastid DNA contained in a 5 ml polystyrene tube, pressurized stopper and mixed gently. Subsequently the mixture was incubated for 15 to 20 P1501 / 99MX minutes at room temperature. During this time, the virus inoculum was removed from the 6 cm plates and the cell monolayers were washed once with CEF negative medium. Three milliliters of negative CEF medium were added to the plasmid DNA / lipof ectin mixture and the contents were pipetted onto the monolayer cell layer. After incubation throughout the car (approximately 16 hours) at 37 ° C in a humidified incubator at 5% COa, the medium was removed and replaced with 5 ml of complete CEF medium. The cells were incubated at 37 ° C in 5% C02 for 3 to 7 days until the cytopathic effect of the virus was 80-100%. The virus was collected as described above for the preparation of virus stock material. This storage material is referred to herein as a storage material for transfection and subsequently was analyzed for recombinant virus by "plaque hybridization method for purification of recombinant PSV.

RECOMBINATION PROCEDURE HOMOLOGA TO GENERATE HVT / MDV HERPESVIRUS RECOMBINANT AVIAN CHEMÉRICOS: Transfectar the first or second monolayer of embryo fibroblast chicken subconfluent (70 to 80% confluent). Day of use, change of medium to maintain medium (medium FIO + medium M199 (1: 1 mixture), fetal bovine serum at 1%, 2% glutamine, 1% NEAA, 1% Penn-Strep). (1) Mix P1501 / 99MX DNA: 20 μl of parental viral DNA (visible amount on gel); 2-3 μg of insertion vector; dH 0 to 300 μl. (2) Calcium phosphate precipitate (300 ul DNA, 37 ul 2.5 M CaCl2: 340 ul 2X HEPES-regulated saline solution. (3) incubate mix 1 minute at room temperature, add to the cells dropwise (within the medium ) Disperse reaction mixture evenly on 2 x 35 mm plates (2 wells in a 6-well plate). (4) Incubate cells at 39 ° C for 3 hours. (5) Undergo shock with glycerol cells for 1 hour. minute, replace medium / reaction mixture with 1 ml of 15% glycerol in PBS After 1 minute, remove glycerone and wash 3X monolayer with PBS. (6) Alxlate cells with maintenance medium and incubate at 39 ° C throughout night. (7) The following day, replace with fresh medium and continue incubation until CPE is observed, change the medium every 2 to 3 days? The infected cells are subcultured, one or more times, in a larger dish until that CEP is achieved.

CLONA SUBGENOMICA 172-07. BA2. Plasmid 172-07. BA2 was constructed in order to generate the recombinant HVT. It contains a region of approximately 25,000 base pairs of genomic HVT DNA. It can be used together with other subgenomic clones according to the PROCEDURE FOR GENERATION OF RECOMBINANT HERPESVIRUS FROM SUBGENOMIC FRAGMENTS P1501 / 99MX TRANSLATORS for the construction of recombinant HVT. This plasmid can be constructed using standard recombinant DNA techniques (Maniatis et al, 1982 and Sambrook et al, 1989), by binding two restriction fragments from the following sources. The first fragment is a restriction fragment of BamH1 to BamHI of 2999 base pairs of pSP64 (Promega). The second fragment is a BamHI # 2 fragment of approximately 25,000 base pairs of HVT (Buckmaster et al., 1988).

VECTOR OF HOMOLOGY 172-29.31. Plasmid 172-29.31 was constructed for the purpose of inserting foreign DNA into the HVT. It contains a unique Xhol restriction enzyme site within which it can be inserted into foreign DNA - when a plasmid containing a foreign DNA insert at the XhoI site is used according to the COTRANSFECTION OF DNA TO GENERATE RECOMBINANT HERPESVIRUS or THE PROCEDURE TO GENERATE RECOMBINANT HERPESVIRUS FROM SUBGENOMIC TRANSLATING FRAGMENTS, a virus containing foreign DNA will be obtained. This plamid can be constructed using standard recombinant DNA techniques (Maniatis et al, 1982 and Sambrook et al, 1989), joining two restriction fragments from the following sources. The first fragment is a restriction fragment of P1501 / 99 X BamH1 to BamH1 of approximately 2999 base pairs of pSP64 (Promega). The second fragment is a BamHl # 16 fragment of approximately 3300 base pairs of HVT (Buckmaster et al., 1988). The complete sequence of the BamHI fragment # 16 is provided in SEQ ID NO. 1. Note that the fragment was cloned so that the ORF UL43 is in the transcriptional orientation opposite to the β-lactamase gene of pSP64.

VECTOR OF HOMOLOGY 172-63.1. Plasmid 172-63.1 e was constructed for the purpose of inserting foreign DNA into HVT. It contains a unique Xhol restriction enzyme site into which foreign DNA can be inserted. When a plasmid containing a foreign DNA insert in the Xh0thite is used according to the COTRANSFECTION OF DNA TO GENERATE RECOMBINANT HERPESVIRUS or the PROCEDURE TO GENERATE RECOMBINANT HERPESVIRUS FROM SUBGENOMIC TRANSLATING FRAGMENTS, a virus containing the foreign DNA is will get. This plasmid can be constructed using standard recombinant DNA techniques (Maniatis et al, 1982 and Sambrook et al 1989), joining the two restriction fragments from the following sources. The first fragment e? an EcoRI to EcoRI restriction fragment of approximately 2999 base pairs of pSP64 (Promega). The second fragment is the EcoRI # 9 fragment of approximately 5500 base pairs of P1501 / 99MX HVT. Note that the EcoRI fragment was cloned such that the unique Xhol site is closest to the unique HindIII site in the vector pSP64.

CLONA SUBGENOMICA 407-32.1C1. Cosmid 407-32.1C1 was constructed for the purpose of generating recombinant HVT and recombinant chimeric viral vaccine comprising a chimera from the short region of the Marek's disease virus and the long region of the turkey herpesvirus. It contains a region of approximately 38,850 base pairs of the genomic HVT DNA (see Figure 2). This region includes BamHI fragments 11, 7, 8, 21, 6, 18, approximately 1250 base pairs of fragment 13, and approximately 6,700 base pairs? of the fragment 1. It can be used together with other subgenomic clones according to the PROCEDURE TO GENERATE RECOMBINANT HERPESVIRUS FROM SUBGENOMIC TRANSLATING FRAGMENTS for the construction of recombinant HVT. This cosmid can be constructed as described above in the PROCEDURE FOR GENERATING HERPES RECOMBINANT VIRUS FROM SUBGENOMIC TRANSLATING FRAGMENTS. It was isolated from the DNA library cut by screening with probes Pl and P4 (which are described in Figure 2). A bacterial strain containing this cosmid has been deposited on March 3, 1993 according to the Budapest Treaty on the International Deposit of Microorganisms for the purposes of P1501 / 99MX Patent Procedures, with the Patent Crop Deposit Agency of the American Type Culture Collection 12301 Parkiawn Drive, Rockville, Maryland 20852 U.S.A. under ATCC Accession No. 75428.

CLONA SUBGENOMICA 407-32.2C3. The cosmid 407-32.2C3 was constructed for the purpose of generating recombinant HVT. It contains a region of approximately 40,170 base pairs of the genomic HVT DNA (see Figure 2). This region includes BamHl 10, 14, 19, 17, 5 and approximately 2100 base pairs of fragment 2. It can be used together with other subgenomic clones according to the PROCEDURE FOR GENERATING RECOMBINANT HERPESVIRUS FROM SUBGENOMIC TRANSLATING FRAGMENTS for the construction of recombinant HVT. . This cosmid can be constructed as described before in the PROCEDURE TO GENERATE RECOMBINANT HERPESVIRUS FROM D? TRANSLATING SUBGENOMICAL FRAGMENTS. It was isolated from the cut DNA library by screening with the probes Pl and P2 (which is described in Figure 2). A bacterial strain containing this cosmid has been deposited according to the Budapest Treaty on the International Deposit of Microorganisms for the Purpose of Patent Procedures, at the Patent Crop Agency of the American Type Crop Collection Parkiawn Drive 12301, Rockville, P150I / 99 X Maryland 20852 U.S.A. under ATCC Accession No. 75430.

CLONA SUBGENOMICA 407-32.5G6. The cosmid 407-32.5G6 was constructed for the purpose of generating recombinant HVT. Does it contain a region of approximately 40,000 base pairs? of genomic HVT DNA (see Figure 2). This region includes BamHI fragments 9, 3, 20, 12, 16, 13, approximately 1650 base pairs of fragment 2 approximately 4000 base pairs of fragment 11. It can be used together with other subgenomic clones according to the PROCEDURE FOR GENERATING RECOMBINANT HERPESVIRUS A DEPARTURE OF SUBGENOMIC TRANSLATING FRAGMENTS for the construction of recombinant HVT. This cosmid can be constructed as described above in the PROCEDURE FOR GENERATING RECOMBINANT HERPESVIRUS FROM SUBGENOMIC TRANSLATING FRAGMENTS. It was isolated from the DNA library cut by screening with probes P2 and P3 (described in Figure 2). The bacterial strain containing this cosmid has been deposited on March 3, 1993 under the Budapest Treaty on the International Deposit of Microorganisms for the Purpose of Patent Procedures before the Patent Crop Agency of the American Type Culture Collection, 12301 Parkiawn Drive, Rockville, Maryland 2085 USA with Accession No. ATCC 75427.

P1501 / 99MX VECTOR OF HOMOLOGY 435-47.1. The plasmid 435-47.1 was constructed for the purpose of inserting foreign DNA into HVT. It contains a unique HindIII restriction enzyme site within which foreign DNA can be inserted. When a plasmid containing a foreign DNA insert in the GinlII site is used according to the COTRANFECTION OF DNA TO GENERATE RECOMBINANT HERPESVIRUS or the PROCEDURE TO GENERATE RECOMBINANT HERPESVIRUS FROM SUBGENOMIC TRANSLATING FRAGMENTS, a virus containing the foreign DNA will be obtained . This plasmid can be constructed using standard recombinant DNA techniques (Maniatis et al, 1982 and Sambrook et al, 1989), joining two restriction fragments a from the following source. The first fragment is an EcoRI to EcoRI restriction fragment of approximately 2999 base pairs of pSP64 (Promega). The second fragment is the EcoRI # 7 fragment of approximately 7300 base pairs of HVT. Note that the HindIII site of vector pSP64 was removed by digestion of the subclone with GinlII followed by a Klenow reagent reaction and religation. A synthetic HindIII ligand (CAAGCTTG) was then inserted into the unique Stul site of the EcoRI # 7 fragment.

CLONA SUBGENOMICA 437-26.26. Plasmid 437-26.26 was constructed for the purpose of generating recombinant HVT.

P1501 / 99MX Contains approximately a region of 15,300 base pairs? of genomic HVT DNA. It can be used together with other subgenomic clones according to the PROCEDURE TO GENERATE RECOMBINANT HERPESVIRUS FROM SUBGENOMIC TRANSLATING FRAGMENTS for the construction of recombinant HVT. This plasmid can be constructed using standard recombinant DNA techniques (Maniatis et al, 1982 and Sambrook et al, 1989), joining two restriction fragments from the following sources. The first fragment is a HindIII to BamH1 restriction fragment of approximately 2970 base pairs of pSP64 (Promega). The second fragment is the BamHl to Stul subfragment of approximately 15,300 base pairs? of the BamHl fragment # 2 of HVT (Buckmaster et al., 1988). Note that the fragment BamHl # 2 contains 5 Stul sites, the site used in this subcloning became a HindIII site as described in the PROCEDURE FOR GENERATING RECOMBINANT HERPESVIRUS FROM SUBGENOMIC TRANSLATING FRAGMENTS.

CLONA SUBGENOMICA 415-09. BAl. The cosmid 415-09. BAl was constructed for the purpose of generating recombinant HVT. It contains a BamHI # 1 fragment of approximately 29,500 base pairs of the genomic HVT DNA. It was used together with other subgenomic clones according to the PROCEDURE TO GENERATEP1501 / 99 X RECOMBINANT HERPESVIRUS FROM D? SUBGENOMIC TRANSLATING FRAGMENTS for the construction of recombinant HVT. This cosmid was constructed by joining two restriction fragments (Sambrook, et al., 1989) from the following sources. The vector is a BamH1 to BamH1 restriction fragment of about 4430 base pairs of pSY1005 derived from pHC79 (Bethesda Research Labs, Inc.) and pWE15 (Stratagene, Inc.). The first fragment is the BamHI # 1 fragment of approximately 29,500 base pairs? of the HVT genome (Buckmaster et al., 1988).

CLONA SUBGENOMICA 67-01. A40. The cosmid 672-01. A40 was constructed for the purpose of generating recombinant HVT. It was isolated as a subclone of cosmid 407-32.1C1 (see Figures 2 and 5). The cosmid 672-01. A40 contains a subfragment Notl to _AscI of approximately 14,000 base pairs and an AscI to BamHl subfragment of approximately 1300 base pairs of cosmid 407-32.1C1. The cosmid was constructed by joining the restriction fragments (Sambrook, et al., 1989) from the following sources. The vector is a Notl to BamHl fragment of approximately 2700 base pairs constructed from pNEB193 (New England Biolabs, Inc.) containing a Notl ligand inserted into the Smal site. Fragment 1 is a region of approximately 15,300 base pairs of genomic HVT DNA. The region includes BamHI 11 fragments and P1501 / 99MX 7 and approximately 1250 base pairs of fragment 13. It was used together with other clones? subgenomics according to the PROCEDURE TO GENERATE RECOMBINANT HERPESVIRUS FROM SUBGENOMIC TRANSLATING FRAGMENTS for the construction of recombinant HVT.

CLONA SUBGENOMICA 654-45.1. Plasmid 654-45.1 was constructed for the purpose of generating recombinant HVT. It was isolated as an Ascl subclone from the acid 407-32.1C1 (see Figures 2 and 5). The cosmid was built by joining the fragment? of restriction (Sambrook, et al., 1989) from the following sources. The vector is an AscI fragment of approximately 2000 base pairs constructed from 2000 bp AatlI to PvuII fragment of pNEB 19 (New England Bilabs, Inc.) whose ends were made blunt with DNA polymerase and AscI ligands were inserted. Fragment 1 is an AscI to AscI fragment of approximately 8600 base pairs of the genomic HVT DNA. This region includes the BamHl fragments 10 and 21 and approximately 1100 base pairs of fragment 6 and approximately 1300 base pairs? of fragment 7. The Xhol (Nucleotide # 1339-1344; SEQ ID NO: 12)? t has become a unique PacI site using the synthetic DNA linkers. The Pací site was used in the insertion and expression of foreign genes in HVT (see Figure 3A). It was used P1501 / 99MX together with other subgenomic clones according to the PROCEDURE TO GENERATE RECOMBINANT HERPESVIRUS FROM SUBGENOMIC TRANSLATING FRAGMENTS for the construction of recombinant HVT.

CLONA SUBGENOMICA 686-63. Al. Plasmid 686-63. Al was constructed for the purpose of generating recombinant HVT. It was isolated as an Ascl subclone from cosmid 407-32.1C1 (see Figures 2 and 5). The cosmid was constructed by joining the restriction fragments (Sambrooks, et al., 1989) from the following sources. The vector is an AscI fragment of approximately 2000 base pairs constructed from an AatlI to PvuII fragment of 2000 base pairs of pNEBl93 (New England Biolabs, Inc.) whose ends were blunt-ended with Klenow DNA polymerase and Ascl linkers inserted. . Fragment 1 is an AscI to AscI fragment of approximately 8600 base pairs of the genomic HVT DNA. This region includes BamHl fragments 10 and 21, and approximately 1100 base pairs of fragment 6 and approximately 1300 base pairs of fragment 7. The Xhol site (Nucleotide # 1339-1344; SEQ ID NO: 12) had become a Unique Notl site using synthetic DNA linkers. The Notl site was used for the insertion and expression of foreign genes in HVT (see Figure 3B). Was it used together with other clones? subgenomics according to the PROCEDURE TO GENERATE H? TRP? SVIRUS P1501 / 99MX RECOMBINANT FROM SUBGENOMIC TRANSLATING FRAGMENTS for the construction of recombinant HVT.

CLONA SUBGENOMICA 672-07. C40 The cosmid 672-07. C40 was constructed for the purpose of generating recombinant HVT. It was isolated as a subclone of cosmid 407-32.1C1 (see Figures 2 and 5) Cosmid 672-07. C40 contains a subfragment of BamHl to AscI of approximately 1100 base pairs and a subfragment of AscI to Notl of approximately 1300 base pairs of cosmid 407-32.1C1. The cosmid was constructed by joining the restriction fragments (Sambrook, et al., 1989) from the following sources. The vector is a Notl to BamHl fragment of approximately 2700 base pairs constructed from pNEBl93 (New England Biolabs, Inc.) containing a Notl lingl inserted into the Smal site. Fragment 1 is a region of approximately 14,100 base pairs of the genomic HVT DNA. This region includes fragments 6 and 18 of BamHI and a BamHI to NotI fragment of approximately 2600 base pairs within the BamH1 # 1 fragment. It was used together with other subgenomic clones according to the PROCEDURE TO GENERATE RECOMBINANT HERPESVIRUS FROM SUBGENOMIC TRANSLATING FRAGMENTS, for the construction of recombinant HVT.

P1501 / 99MX SUBGENOMIC CLONE 720-51.3. The cosmid 720.51.3 was constructed for the purpose of generating recombinant HVT and generating recombinant chimeric viral vaccine comprising a chimera from the short region of the Marek's disease virus and the long region of the turkey herpesvirus. It contains a region of approximately 34,700 base pairs of the genomic HVT DNA (see Figure 2). This region includes the BamHl fragments 11, 7, 8, 21, 6, 18, approximately 1250 base pairs? of the BamHl fragment 13 t approximately 2,600 base pairs of the BamHl 1 fragment. It was used together with other subgenomic clones according to the PROCEDURE TO GENERATE HERPES RECOMBINANT VIRUSES FROM SUBGENOMIC TRANSLATING FRAGMENTS for the reconstruction of the recombinant HVT. Cosmid 720-51.3 was constructed from cosmid 407-32.1C1 in the following procedure. Cosmid 407-32.1C1 was digested with restriction endonuclease NotI to give a HVT genomic fragment of approximately 34,700 base pairs, a genomic fragment of HVT of approximately 4,100 base pairs and a cosmid vector pWEl5 of approximately 8,000 base pairs. The 34,700 base pair HVT genomic fragment and the 8,000 base pair pWEl5 cosmid vector were ligated to form the cosmid 720.51.3. The cosmid 720-53.1 lacks approximately 4,100 base pairs of the BamHl 1 genomic fragment of the P1501 / 99MX HVT, which is present in cosmid 407-32.1C1. The cosmid 720-51.3 contains an Xhol site within the genomic fragment of HVT EcoRI # 9 which is useful for the insertion of the foreign DNA into the recombinant HVT and into the recombinant chimeric viral vaccine comprising a chimera of the short region of the Marek's disease and the long region of turkey herpes virus.

CLONA SUBGENOMICA 721-38. ÍJ. The cosmid 721-38.1J was constructed for the purpose of inserting the gA, gD and gB genes of MDV into the single short region of the HVT and in order to generate the recombinant HVT. The coMido721-38. ÍJ contains the genes gA, gD and gB of the MDV inserted within the Stul site in the US2 gene of HVT converted into a unique HindIII site within the BamHI fragment # 1 of the unique short region of the HVT. This region of the BamH1 # 1 fragment of HVT containing the MDV genes is derived from S-HVT-062. The cosmid 721-38.1J was constructed by a partial restriction digestion with BamHI of DNA S-HVT-062 and was isolated from a fragment of approximately 39,300 base pairs. The cosmid was constructed using standard recombinant DNA techniques (Sambrook, et al., 1989) by binding restriction fragments from the following sources. The vector is a BamHI fragment of approximately 8,200 base pairs? from the cosmid vector pWEl5. The first P1501 / 99 X fragment is a BamHI fragment of approximately 900 base pairs from the repeat region of the HVT genome. The second fragment is a BamHl to Stul subfragment of approximately 15,500 pairs of BamHl # 1 HVT bath. The third fragment is a cassette of approximately 8,400 base walls containing the gA, gD and gB genes of MDV. The fourth fragment is a HindIII to BamHI subfragment of approximately 14,500 base pairs of BamHI # 1 of HVT.

CLONA SUBGENOMICA 722-60. E2. The cosmid 722-60. E2 was constructed for the purpose of inserting the gA, gD and gB genes of MDV and the HN and F genes of NDV into the single short region of the HVT and in order to generate recombinant HVT. The cosmid 722-60. E2 contains the genes gA, gD and gB of MDV and the HN and F genes of NDV inserted within a Stul site in the .gen US2 of HVT converted to a unique HindIII within the BamHI fragment # 1 of the single short region of HVT. All five genes were inserted in the same transcriptional orientation as the US2 gene of HVT. This region of the BamH1 # 1 fragment of HVT containing the MDV and NDV genes was derived from S-HVT-106. The cosmid 722-60. E2 was constructed by a partial restriction digestion with BamHI of S-HVT-106 and a fragment of approximately 46,300 base pairs was isolated. The second cosmid was built using the P1501 / 99MX standard recombinant DNA techniques (Sambrook, et al., 1989) joining the restriction fragments from the following sources. The vector is a BamHI fragment of approximately 6,100 base pairs from cosmid vector pSY626 derived from pHC79 (Bethesda Research Labs, Inc.) and pWE15 (Stratagene, Inc.). The first fragment is a BamHI fragment of approximately 900 base pairs from the repetition region of the HVT genome. The second fragment is a subfragment of BamHl to Stul of approximately 15,500 base pairs of BamHl # 1 of the HVT. The third fragment is a cassette of approximately 15,400 base pairs containing the MDV gA gene (SEQ ID NO: 6), the PRV gX promoter (Lomniczi et al., 1984), the NDV NH gene (SEQ. ID NO: 8), PRX gX polyadenylation site (Lomniczi et al., 1984), the immediate early promoter HCMV (DR Thomsen, et al 1981), the NDV F gene (SEQ ID NO: 10), the HSV polyadenylation TK (McGeochj, et al. ., 1985), the gD gene of MDV, the US3 polyadenylation site of ILTV of approximately 450 base pairs? and elg in MDV gD. The fourth fragment is a Stul to BamHl subfragment of approximately 14,500 base pairs of BamHI # 1 of HVT.

CLONA SUBGENOMICA 739-27.16. The cosmid 739-27.16? E built for the purpose of building an HVT / MDV virus P1501 / 99 Chimeric X containing the HVT genes of the single long region and the MDV type 1 genes of the single short region. The cosmid 739-27.26 contains the complete single short region of MDV type 1. This region contains the entire Smal B fragment and two Smal K fragments. The cosmid 739-27.16 was constructed by a partial restriction digestion with Smal MDV DNA and the isolation of a fragment of approximately 29,000 to 33,000 base pairs. The cosmid was constructed using standard recombinant DNA techniques (Sambrook, et al., 1989) by binding restriction fragments from the following sources. The vector is a BamHI fragment of approximately 8200 base pairs (the ends of which were made blunt with Lenow DNA polymerase) from the cosmid pWEl5. The first fragment is a Smal K fragment of approximately 4050 base pairs from the short internal repeat region of the MDV genome. The second fragment is a Smal B fragment of MDV, of approximately 21,000 base pairs. The third fragment is a Smal K fragment of approximately 3,650 base pairs from the short terminal repeat region of the MDV genome (Fukuchi, et al., 1984, 1985).

CLONA SUBGENOMICA 751-87. 8. Plasmid 751-87. 8? E was built for the purpose of generating recombinant HVT. He P1501 / 99MX plastic 751-87. A8 that contains the chicken myelomonocytic growth factor (cGMF) gene within the polyidicity of the plamid 654-45.1. The cMGF gene uses the immediate early promoter HCMV and the TK polyadenylation signal of HSV-1. The cosmid was constructed using standard recombinant DNA techniques (Sambrook, et al., 1989). The following fragments were inserted into the Syrian Pací of the subgenomic clone HVT, 654-45.1. The first fragment is a PstI to Avall restriction subfragment of approximately 1191 base pairs, from the genomic Xbal E fragment of HCMV (D.R. Thomsen, et al., 1981). The second fragment is a fragment of approximately 640 base pairs that codes for the cMGF gene (58) derived by reverse transcription and polymerase chain reaction (PCR) (Sambrook, et al., 1989) of RNA ISOLATED FROM CELLS CHICKEN SPLICE STIMULATED WITH CONCANAVALINE A. The antisense primer used for reverse transcription and PCR was 5 '-CGCAGGATCCGGGGCGTCAGAGGCGGGCGAGGTG-3' (SEQ ID NO: 19). The homosentide primer used for PCR was 5 '-GAGCGGATCCTGCAGGAGGAGACACAGAGCTG-3' (SEQ ID NO: 20). The cMGF fragment was then subcloned with the HCMV IE promoter using the BamH1 sites generated by the PCR primers. The DNA fragment containing the coding sequence for amino acid 1 to amino acid 201 of the cMGF protein (58) including a P 15 01/9 9IIX guiding sequence of 23 amino acids in the amino terminus and 178 amino acids of the mature cMGF protein. The third fragment is a Smal to Smal restriction subfragment of approximately 784 base pairs of the Q-restriction fragment of BamHI of HSV-1 (McGeoch, et al., 1985).

CLONA SUBGENOMICA 761-07. Al. Plasmid 761-07. Al was constructed for the purpose of generating recombinant HVT. The plasmid 761-07. It contains the chicken interferon gene inserted into the Pací site of plasmid 654-45.1. The chicken interferon gene uses the immediate early promoter HCMV and the TK polyadenylation signal of HSV-1. The cosmid was constructed using standard recombinant DNA techniques (Sambrook, et al., 1989). Subsequently, the fragments were inserted into the Pací site of the subgenomic clone HVT 654-45.1. The first fragment is a PstI to Avall restriction subfragment of approximately 191 base pairs of the genomic Xbal E fragment of HCMV (D.R. Thomsen, et al., 1981). The second fragment is a fragment of approximately 577 base pairs for the chicken interferon gene (59 derived by reverse transcription and polymerase chain reaction (PCR) (Sambrook, et al., 1989) of RNA ISOLATED FROM CELLS BACON STIMULATED BY CONCANAVALINE The antisense primer used for reverse transcription and for PCR P1501 / 99MX was 5 '-TGTAGAGATCTGGCTAAGTGCGCGTGTTGCCTG-3' (SEQ ID NO: 21). The homosentide primer used for PCR was 5 '-TGTACAGATCTCACCATGGCTGTGCCTGCAAGC-3' (SEQ ID NO: 21). The fragment of the chicken interferon gene was subsequently positioned to the HCMV IE promoter using BglII generated by the PCR primers. The DNA fragment contains the coding sequences from amino acid 1 to amino acid 193 of the chicken interferon protein (59) which includes a signal sequence of 31 amino acids in the amino terminus and 162 amino acids of protein-encoding chicken interferon . The third fragment is a Smal to Smal restriction subfragment of approximately 784 base pairs of the Q-restriction fragment of BamHI of HSV-1 (McGeoch, et al., 1985).

VECTOR OF HOMOLOGY 301-07. Y # D1: Plasmid 301-07.Y # D1 was constructed for the purpose of generating the recombinant chimeric HVT / MDV vaccine by expressing a foreign DNA sequence. The lacZ gene of E. Coli was expressed under the control of the gV promoter of PRV. The HVT DNA is an Ascl subclone of cosmid 407-32.1C1 (see Figures 2 and 5). The cosmid was constructed by joining the restriction fragment (Sambrooks, et al., 1989) from the following sources. The vector is an AscI fragment of approximately 2000 base pairs? built from a fragment P1501 / 99MX AatlI to PvuII of 2000 base pairs of pNEBl93 (New England Biolabs, Inc.) blunt ended by Klenow DNA polymerase and inserted Ascl binders. The HVT fragment is an AscI to AscI fragment of approximately 8600 pairs of DNA from genomic HVT DNA. This region includes fragments. BamHI 10 and 21 and approximately 1100 base pairs of fragment 6 and approximately 1300 base pairs of fragment 7. The Xhol site (Nucleotide # 1339-1344; SEQ ID NO: 12) was used for insertion and expression of IO? foreign genes in HVT (see Figure 3B), The foreign DNA inserted into the Xhol site of HVT is as follows: Fragment 1 is a restriction subfragment Salí a BamHl of approximately 413 base pairs? of PRV restriction fragment BamHl, 10 (Lomniczi et al., 1984). Fragment 2 is a restriction fragment of BamHI to PuvII of approximately 3010 base pairs of plasmid pJF751 (Ferrari et al., 1985). Fragment 3 is a restriction subfragment from Ndel to Salí of approximately 754 base pairs? of restriction fragment # 7 BamHl from PRV (Lomniczi et al., 1984). The plasmid 301-07. Y # D1 was used together with S-HVY-145 according to the TRANSFECTION OF DNA TO GENERATE RECOMBINANT VIRUS for the construction of recombinant chimeric HVT / MDV vaccine.

CLONA SUBGENOMICA 852-52.114. The plasmid 852-52.114 P1501 / 99MX? E was constructed for the purpose of generating recombinant chimeric HVT / MDV vaccine expressing a foreign DNA sequence. The genes of glycoprotein D (gD) and glycoprotein I (gl) of infectious lanringotracheitis virus (ILT) were expressed under the control of the gD and gl promoters of the ILT virus. The HVT DNA is an Ascl subclone of cosmid 407-32.1C1 (see Figures 2 and 5). The cosmid was constructed by joining the restriction fragments (Sambrooks, et al., 1989) from the following sources. The vector is an AscI fragment of approximately 2000 base pairs? constructed from an AatlI to PvuII fragment of 2000 base pairs of pNEBl93 (New England Biolabs, Inc.) whose ends were made blunt with Klenow DNA polymerase and ligands were inserted with Ascl. The HVT fragment is an AscI to AscI fragment of approximately 8600 base pairs of the genomic HVT DNA. This region includes the BamHI fragments 10 and 21 and approximately 1100 base pairs? of fragment 6 and approximately 1300 base pairs of fragment 7. The Xhol site (Nucleotide # 1339-1344; SEQ ID NO: 12) was used for the insertion and expression of the foreign genes in HVT. (See Figure 3B). The foreign DNA inserted into the XhoI site of HVT is as follows: the fragment is a SalI to HindIII restriction subfragment of about 3556 base pairs of the ILP Asp718I genomic fragment # 8 (10.6 kb). The plasmid 852-52.114 was used P1501 / 99MX together with other subgenomic clones according to the PROCEDURE FOR GENERATING R.ECOMBINANT HERPESVIRUS FROM SUBGENOMIC TRANSLATING FRAGMENTS, for the construction of recombinant HVT.

CLONA SUBGENOMICA 854-33.6. Plasmid 854-33.6 was constructed for the purpose of generating the recombinant HVT expressing a foreign DNA sequence and generating a recombinant chimeric viral vaccine comprising a chimera from the short region of the Marek's disease virus and the long region of the turkey herpesvirus. which expresses a strange DNA sequence. The hemagglutinin (HN) gene of the disease virus (NDV) is expressed under the control of the gX PRV promoter. The fusion gene (F) of NDV is expressed under the control of the immediate early promoter HCMV. The NDV HN and F genes under the control of their respective promoters are transcribed in the opposite direction of the UL52, UL54 and UL55 genes of HVT in the adjacent HVT genomic DNA. The HVT DNA is derived from cosmid 720-51.3 which is a subclone of cosmid 407-32.1C1 (see Figures 2, 5). The HVT fragment is a fragment of approximately 34,700 base pairs of the genomic HVT DNA. This region includes fragments 11, 7, 8, 21, 6, 18 of BamHI and approximately 1250 base pairs of fragment 13 and approximately 2600 base pairs of fragment 1. The Xhol site (Nucleotide # 1339-1344; SEQ ID NO. .

P1501 / 99MX 12) was used for insertion and expression of foreign genes in HVT. (See Figure 3). The cosmid was constructed by joining the restriction fragments (Sambrooks, et al., 1989) from the following sources. The vector is a Notl fragment of about 8,000 base pairs of pWEl5. The foreign DNA inserted into the Xhol site of HVT is as follows: fragment 1 is a restriction subfragment SalI to BamHl of approximately 413 base pairs of fragment # 10 Ba Hl of PRV (Lomniczi, et al., 1984). Fragment 2 is an Avall-Nael restriction fragment of approximately 1811 base pairs? of the full-length NDV HN cDNA clone (strain Bl). The fragment 3 is a subfragment Ndel to Salí of approximately 750 base pairs of BamHl # 7 of PRV (B. Lomniczi et al.). Fragment 4 is a PstI to Avall restriction subfragment of approximately 1191 base pairs? of the genomic Xbal E fragment of HCMV (D.R. Thomsen, et al., 1981). The fragment 5 e? a restriction fragment of BamHl to P? tl of approximately 1812 base pairs of the full length NDV FD cDNA clone (strain Bl, Reference: WO96 / 05291). The fragment is a Smal to Smal restriction subfragment of approximately 784 base pairs of the Q-restriction fragment of BamHI of HSV-1 (McGeoch, et al., 1985).

P1501 / 99MX VECTOR OF HOMOLOGY 864-74.18: Plasmid 864-74.18 was constructed for the purpose of generating a recombinant chimeric HVT / MDV vaccine expressing a foreign DNA sequence. The lacZ gene of E. Coli is expressed under the control of the gV promoter of PRV. The HVT DNA is an Ascl subclone of cosmid 407-32.1C1 (see Figures 2 and 5). The cosmid was constructed by joining the restriction fragments (Sambrooks, et al., 1989) from the following sources. The vector is an AscI fragment of approximately 2000 base pairs? built from an AatlI to PvuII fragment of 2000 base pairs? from pNEBl93 (New England Biolabs, Inc.) whose ends were made blunt with the Klenow DNA polymerase and Ascl ligands were inserted. The HVT fragment is an AscI to AscI fragment of approximately 8600 base pairs of the genomic HVT DNA. This region includes fragments 10 and 21 of BamHI and approximately 1100 base pairs of fragment 6 and approximately 1300 base pairs of fragment 7. The Xhol site (Nucleotide # 1339-1344; SEQ ID NO: 12) was used for insertion and expression of foreign genes in HVT. (See Figure 3B). The foreign DNA inserted into the Xhol site of HVT is as follows: Fragment 1 is a SalI to HindIII restriction subfragment of approximately 3556 base pairs of the genomic # 8 fragment Asp718l of ILTV (10.6 kb). Fragment 2 is a restriction subfragment Salí a VamlII de P1501 / 99MX approximately 413 base pairs of PRV BamHl restriction fragment 10 (Lomniczi et al., 1984). Fragment 3 is a restriction fragment BamHl to PvuII of approximately 3010 base pairs? of plasmid pJF751 (Ferrari et al., 1985). Fragment 4 is a restriction subfragment Ndel to Salí of approximately 754 base pairs of fragment # 7 of restriction BamHl of PRV (Lomniczi et al., 1984). Plasmid 864-74.18 was used together with S-HVY-145 according to TRANSFECTION OF DNA TO GENERATE RECOMBINANT VIRUS, for the construction of the recombinant chimeric vaccine of HVT / MDV.

SUBONAUTICAL CLONE 867-96. B9 The plasmid 867-96. B9 was built for fine? of generating recombinant HVT expressing a foreign DNA sequence. The E. coli β-galac tosidase gene expressed under the control of a novel chicken anemia virus promoter. The HVT DNA is an AscI subclone of cosmid 407-32.1C1 (Figures 2 and 5). The cosmid was constructed by joining the restriction fragments (Sambrooks, et al., 1989) from the following sources. The vector is an AscI fragment of approximately 2000 base pairs constructed from an AatlI to PvuII fragment of 2000 base pairs of pNEBl93 (New England Biolabs, Inc.) with blunt ends with Klenow DNA polymerase and AscI ligands inserted. The HVT fiagment is an AscI to AscI fragment of P1501 / 99MX approximately 8600 base pairs? of genomic HVT DNA. This region includes fragments ^ 10 and 21 of BamHI and approximately 1100 base pairs of fragment 6 and approximately 1300 base pairs of fragment 7. The Xhol site (Nucleotide # 1339-1344; SEQ ID NO: 12) was used for insertion and expression of foreign genes in HVT (see Figure 3B). The foreign DNA inserted into the Xhol site of HVT is as follows: Fragment 1 is a CAV promoter synthesized by PCR as an EcoRI to BamHI fragment of 386 bp from strain CAV CL-1 (NVSL, Dr DB Snyder, Univ. Maryland, SEQ ID No. 23). Fragment 2 is a restriction fragment BamHl to PvuII of 3001 base pairs? of plasmid pJF751 (Ferrari et al., 1985). Fragment 3 is a restriction subfragment Ndel a Salí of approximately 754 base pairs of restriction fragment # 7 of PRV BamHl (Lomniczi et al., 1984). The plasmid 867-96. B9 together with other subgenomic clones according to the PROCEDURE TO GENERATE HERPES RECOMBINANT VIRUSES FROM TRANSLAPANTS SUBGENOMIC FRAGMENTS for the construction of recombinant HVT.

SUBONAUTICAL CLONE 890-77.10. Plasmid 890-77.10 was constructed for the purpose of generating recombinant HVT expressing the foreign DNA sequence and generating recombinant chimeric viral vaccine comprising a P1501 / 99MX chimera of the short region of the Marek's disease virus and the long region of the turkey herpesvirus expressing a foreign DNA sequence. The lacZ gene of E. coli was expressed under the control of the gV promoter of PRV. The fusion gene (F) of New Castle disease virus (NDV) was expressed under the control of the immediate early promoter HCMV. The HVT DNA is a? Ubclone AscI of cosmid 407-32.1C1 (see Figure 2, 5). The lacZ gene of E. coli and the F gene of NDV under the control of their respective promoters are transcribed in the opposite direction of the UL52, UL54 and UL55 genes of HVT in the adjacent HVT genomic DNA. The cosmid was constructed by joining the restriction fragments (Sambrooks, et al., 1989) from the following sources. The vector is an AscI fragment of approximately 2000 base pairs constructed from a 2000 base pair AatlI to PvuII fragment of pNEB193 (New England Biolabs, Inc.) with blunt ends with Klenow DNA polymerase and inserted AscI ligands. The HVT fragment is an AscI to AscI fragment of approximately 8600 base pairs of the genomic HVT DNA. This region includes fragments 10 and 21 of BamHI and approximately 1100 base pairs of fragment 6 and approximately 1300 base pairs of fragment 7. The Xhol site (Nucleotide # 1339-1344; SEQ ID NO: 12) was used for insertion and expression of foreign genes in HVT (see Figure 3B). Strange DNA P1501 / 99HX inserted into the Xhol site of HVT is as follows: Fragment 1 is a restriction subfragment Exit to BamHl of approximately 413 base pairs? of fragment # 10 BamHl of PRV (Lomniczi, et al., 1984). Fragment 2 is a PvII to BamH1 subfragment of approximately 3006 base pairs of pJF751 (Ferrari, et al.). The fragment 3 is a Ndel subframe to come out of approximately 754 base pairs? from BamHl # 7 (B. Lomniczi et al.). The fragment 4 e? a restriction subfragment from PstI to Avall of approximately 1191 base pairs of the genomic Xbal E fragment of HCMV (D.R. Thomsen, et al., 1981). Fragment 5 is a BamHI to PstI restriction fragment of approximately 1812 base pairs? of the full-length NDV F-cDNA clone (strain Bl, reference: WO 96/05291). Fragment 6 is a Smal to Smal restriction subfragment of approximately 784 base pairs of the Q fragment of BamHI restriction of HSV-1 (McGeoch, et al., 1985).

CLONA SUBGENÓMICA 900-87. H8 Plasmid 900-87. H8 was constructed for the purpose of generating recombinant HVT expressing a foreign DNA sequence and generating a recombinant chimeric viral vaccine comprising a chimera from the short region of the Marek's disease virus and the long region of turkey herpesvirus expressing a sequence of strange DNA. The VP2 gene of infectious bursal disease virus (IBDV) is P 150 1/9 91IX expressed under the control of a chicken anemia virus (CAV) promoter. The VP2 gene of IBDV under the control of the CAV promoter is transcribed in the opposite direction to the UL52, UL54 and UL55 genes of HVT in the adjacent HVT genomic DNA. The HVT DNA is an Ascl subclone of cosmid 407-32.1C1 (see Figures 2, 5). The cosmid was constructed by joining the restriction fragments (Sambrooks, et al., 1989) from the following sources. The vector is an AscI fragment of approximately 2000 base pairs constructed from an AatlI to PvuII fragment of 2000 base pairs of pNEBl93 (New England Biolabs, Inc.) whose ends were blunt-ended by Klenow DNA polymerase and binder inserted. Ascl. The HVT fragment is an A? CI to AscI fragment of approximately 8600 base pairs of the genomic HVT DNA. The region includes the BamHl fragments 10 and 21, and approximately 1100 base pairs of fragment 6 and approximately 1300 base pairs of fragment 7. The Xhol site (Nucleotide # 1339-1344; SEQ ID NO: 12) was used for the insertion and expression of foreign genes in HVT (see Figure 3B). The foreign DNA inserted into the XhoIthium of HVT is as follows: Fragment 1 is the CAV promoter synthesized by PCR as the 386 base pair EcoRI to BamHI fragment of strain CAV CL-1 8NVSL; I KNOW THAT ID NO. ). The BamHl site was altered to a site EcoRV in this construction. Fragment 2 is a P1501 / 99MX HindIII to EcoRI fragment of approximately 1356 base pairs encoding the VP2 gene of IBDV derived from plasmid 779-54.1. Bursvac M large segment RNA from IBDV was first cloned as cDNA into two overlapping fragments using the first strand / second strand cDNA synthesis from Okayama-Berg. These clones were linked with unique restriction site to create plasmid 2.40 / 84 # 3. Plasmid 779-54.1 was created by PCR in two parts using plasmid 2.40 / 84 # as a template. The homosense and antisense primers of the first half were 5'-GCCGGCGGCCGCGGATACGATCGGTCTGACCCCGG-3"and 5'TTGTGTGCACCGCGGAGTACCCC-3 ', respectively.The homosense and antisense primers of the second half were 5 * -TCGCGAATCTATTCCAGGTGCCCC-3', and 5'- CGGA? TTCTCCAATTTGGGATGTTGTAAGGCCGA-3 *, respectively.The two halves were joined at a single SalI site.Fragment 3 is the PKTV polyadenylation signal of ILTV synthesized by PCR as a 144 bp SalI fragment from the USDA USDA sensitization strain The homosense and antisense primers used for PCR were '-GGTGCCACGGTCGACGAGTGAAGGTTAAT-3 '(SEQ ID NO.) And '-AGACCGAGCAGAGTCGACGCGCGAAAG-3' (SEQ ID NO.), Respectively.

CLONA SUBGENÓMICA 928-47.1. The cosmid 928-47.1 was constructed for the purpose of generating recombinant HVT P1501 / 99MX expressing a foreign DNA sequence and generating recombinant chimeric viral vaccine comprising a chimera from the short region of Marek's disease virus and from the long region of turkey herpesvirus expressing for a foreign DNA sequence. The vector is a Notl fragment of about 8000 base pairs pWE15. The viral DNA insert consists of two segments. Fragment 1 is an AscI to Seal fragment of approximately 8.6 kb which contains the binding region between the long repeat region of HVT and the short region of MDV from the HVY-145 of the recombinant chimeric virus. Fragment 2 is a Seal to Notl fragment of 28.8 kb derived from 739-27.16, and contains the remainder of the short MDV region from that clone.

CLONA SUBGENÓMICA 928-58. J2. Plasmid 928-58. J2 was constructed for the purpose of generating recombinant HVT expressing a foreign DNA sequence and generating recombinant chimeric viral vaccine comprising a chimera from the short region of Marek's disease virus and the long region of turkey herpesvirus expressing for a sequence of Strange DNA The lacZ gene of E. coli sa is expressed under the control of a glycoprotein I (gl) promoter of infectious laryngotracheitis virus (ILTV). The lacZ gene of E. coli under the control of the ILTV gl promoter is transcribed in the opposite direction to the genos P1501 / 99MX UL52, UL54, UL55 of HVT in the adjacent HVT genomic DNA. The HVT DNA is an Ascl subclone of cosmid 407-32.1C1 (see Figure 2, 5). The cosmid was constructed by joining restriction fragments (Sambrooks, et al., 1989) from the following sources. The vector is an AscI fragment of approximately 2000 base pairs constructed from an AatlI to PvuII fragment of 2000 base pairs of pNEBl93 (New England Biolabs, Inc.) whose ends are blunt with Klenow DNA polymerase and inserted AscI binders inserted. The HVT fragment is an AscI to AscI fragment of approximately 8600 base pairs of the genomic HVT DNA. This region includes fragments 10 and 21 of BamHI and approximately 1100 base pairs? of fragment 6 and approximately 1300 base pairs of fragment 7. The Xhol site (Nucleotide # 1339-1344; SEQ ID NO: 12) was used for the insertion and expression of the foreign genes in HVT (Figure 3B). The foreign DNA inserted into the Xhol site of the HVT is as follows: Fragment 1 is the gD promoter of ILTV synthesized by PCR as a Pmel to BamHI fragment of 279 base pairs from the USDA sensitizing strain of ILTV. The sense and antisense primers used for PCR were '-CAGTTTAAACTCGGATTCTGACTATTAC-3' (SEQ ID NO.) And '-CGGATCCATGCTTTTCGAACGTCC-3' (SEQ ID NO.), Respectively. (See Figure 8). Fragment 2 is P1501 / 99MX a subfragment PvuII to BamH1 of approximately 3006 base pairs of pJF751 (Ferrari, et al.). Fragment 3 is the ILTV PK polyadenylation signal synthesized by PCR as a 144 bp Sali fragment from the USDA USDA challenge strain. The homosentide and antisense primers used for PCR were 5 '-GGTGCCACGGTCGACGAGTGAAGGTTAAT-3' (SEQ ID NO.) And 5 '-AGACCGAGCAGAGTCGACGCGCGAAAG-3' (SEQ ID NO XX), respectively.

CLONA SUBGENÓMICA 928-58. K7 Plasmid 928-58. K7 was constructed for the purpose of generating recombinant HVT expressing a foreign DNA sequence and generating recombinant chimeric viral vaccine comprising the chimera of the short region of the Marek's disease virus and the long region of the turkey herpesvirus expressing for a sequence of Strange DNA The lacZ gene of E. coli is expressed under the control of a glycoprotein D (gD) promoter of infectious laryngotracheitis virus (ILTV). The lacZ gene of E. coli under the control of the gD promoter ILTV is transcribed in the opposite direction to the UL52, UL54 and UL55 genes of HVT in the genomic DNA of adjacent HVT. The HVT DNA is an Ascl subclone of cosmid 407-32.1C1 (see Figure 2, 5). The cosmid was constructed by joining restriction fragments (Sambrooks, et al., 1989) from the following sources. The vector is an AscI fragment of P1501 / 99MX approximately 2000 base pairs constructed from an AatlI to PvuII fragment of 2000 base pairs of pNEBl93 (New England Biolabs, Inc.) whose ends are blunt with Klenow DNA polymerase and AscI ligands inserted. The HVT fragment is an AscI to AscI fragment of approximately 8600 base pairs of the genomic HVT DNA. This region includes fragments 10 and 21 of BamHI and approximately 1100 base pairs of fragment 6 and approximately 1300 base pairs of fragment 7. The Xhol site (Nucleotide # 1339-1344; SEQ ID NO: 12) was used for the insertion and expression of foreign genes in HVT (Figure 3B). The foreign DNA inserted into the Xhol site of the HVT is as follows: Fragment 1 is the gD promoter of ILTV synthesized by PCR as a Pmel to BamHI fragment of 530 base pairs from the USDA sensitizing strain of ILTV. The homosentide and antisense primers used for reverse transcription and PCR were 5 '-GGTTTAAACAGCTGTACTACAGAGTAAC-3' (SEQ ID NO.) And '-CGGATCCATAGCGTTGCGTACAAAG-3' (SEQ ID NO.), Respectively. (See Figure 7). Fragment 2 is a PvuII to BamH1 subfragment of approximately 3006 base pairs of pJF751 (Ferrari, et al.). Fragment 3 is the PKTV polyadenylation signal of ILTV synthesized by PCR as a 144 bp SalI fragment from the USDA USDA challenge strain. The homosense and antisense primers used P1501 / 99MX for PCR were 5 '-GGTGCCACGGTCGACGAGTGAAGGTTAAT-3' (SEQ ID NO.) And 5'- AGACCGAGCAGAGTCGACGCGCGAAAG-3 ' (SEQ ID NO.), Respectively.

CLONA SUBGENÓMICA 949-01.12. Plasmid 949-01.12 was constructed for the purpose of generating recombinant HVT expressing a foreign DNA sequence and generating recombinant chimeric viral vaccine comprising a chimera from the short region of the Marek's disease virus and the long region of the turkey herpesvirus that expresses a foreign DNA sequence. The VP2 gene of the infectious bursal disease virus (IBDV) is expressed under the control of a glycoprotein I (gl) promoter of infectious laryngotracheitis virus (ILTV). The VP2 gene of IBDV under the control of the ILTV gl promoter is transcribed in the opposite direction to that of the UL52, UL54 and UL55 genes of HVT in the adjacent HVT genomic DNA. The HVT DNA is an Ascl subclone of cosmid 407-32.1C1 (see Figure 2, 5). The cosmid was constructed by joining the restriction fragments (Sambrooks, et al., 1989) from the following entities. The vector is an AscI fragment of approximately 2000 base pairs constructed from an AatlI to PvuII fragment of 2000 base pairs? from pNEBl93 (New England Biolabs, Inc.) whose ends were born blunt with Klenow DNA polymerase and AscI ligands inserted. The HVT fragment is a fragment AscI to AscI of P1501 / 99MX approximately 8600 base pairs of genomic HVT DNA. This region includes fragments of BamHl 10 and 21 and approximately 1100 base pairs of fragment 6 and approximately 1300 base pairs? of fragment 7. The Xhol site (Nucleotide # 1339-1344; SEQ ID NO: 12) was used for the insertion and expression of foreign genes in HVT (Figure 3B). The foreign DNA was inserted into the Xhol site of the HVT in the following manner: Fragment 1 is the ILTV gl promoter synthesized by PCR as a Pmel to BamHI fragment of 279 bp from the USDA sensitizing strain of ILTV. The sense and antisense primers used for PCR were 5'-CAGTTTAAACTCGGATTCTGACTATTAC-3 '(SEQ ID NO.) And 5' -CGGATCCATGCTTTTCGAACGTCC-3 '(SEQ ID NO.), Respectively. (See Figure 8). Fragment 2 is a HindII to EcoRI subfragment of approximately 1356 base pairs which codes for the VP2 gene of IBDV derived from plasmid 779-54.1. The Bursvac M large segment RNA from IBDV? Cl cloned first as cDNA in two overlapping fragments using the first strand / second strand cDNA synthesis from Okayama-Berg. These clone? joined in a unique restriction site to create plasmid 2.40 / 84 # 3. Plasmid 779-54.1 was created by PCR in two parts using plasmid 2.40 / 84 # 3 as a template. The homosentide and antisense primers for the first half were 5 '-GCCGGCGGCCGCGGATACGATCGGTCTGACCCCGG- P1501 / 99MX 3 'and 5' -TTGTGTGCACCGCGGAGTACCCC-3 ', respectively. The homosense and antisense primers of the second half were 5 '-TCGCGAATCTATTCCAGGTGCCCC-3' and 5'-CGGAATTCTCCAATTTGGGATGTTGTAAGGCCGA-3 ', respectively, the two halves were joined at a single SalI site. The fragment 3 is the polyadenylation signal PK of ILTV synthesized by PCR as a Sali fragment of 144 bp from the USDA sensitization strain of ILTV. The sense and antisense primers used for PCR were '-GGTGCCACGGTCGACGAGTGAAGGTTAAT-3 '(SEQ ID NO.) And '-AGACCGAGCAGAGTCGACGCGCGAAAG-3' (SEQ ID NO.), Respectively.

SUBONAUTICAL CLONE 949-19.2. Plasmid 949-19.2 was constructed for the purpose of generating recombinant HVT by expressing a foreign DNA sequence and generating a recombinant chimeric viral vaccine comprising a chimera from the short region of the Marek's disease virus and the long region of the turkey herpesvirus that expresses a foreign DNA sequence. The VP2 gene of the infectious bursal disease virus (IBDV) is expressed under the control of a glycoprotein I (gl) promoter of infectious laryngotracheitis virus (ILTV). The VP2 gene of IBDV under the control of the ILTV gl promoter was transcribed in the same direction as the UL52, UL54 and UL55 genes of HVT in the adjacent HVT genomic DNA. The DNA of P1501 / 99MX HVT was derived from cosmid 720-51.3, which is a subclone of cosmid 407-32.1C1 (see Figure 2.5). The HVT fragment is a fragment of approximately 34700 base pairs of genomic HVT DNA. This region includes the 11, 7, 8, 21, 6, 18 fragments of BamHl and approximately 1250 base pairs of fragment 13 and approximately 2600 base pairs of fragment 1. The Xhol site (Nucleotide # 1339-1344; SEQ ID NO.12) was used for the insertion and expression of foreign genes in HVT (see Figure 3B). The cosmid was constructed by joining the restriction fragments (Sambrooks, et al., 1989) from the following sources. The vector is a Notl fragment of approximately 8,000 base pairs of pWE15. The foreign DNA inserted into the Xhol site of HVT is as follows: Fragment 1 is the ILTV gl promoter synthesized by PCR as a Pmel to BamHI fragment of 279 bp from the USDA USDA sensitization strain. The sense and antisense primers used for PCR were 5'- CAGTTTAAACTCGGATTCTGACTATTAC-3 '(SEQ ID NO.) And '-CGGATCCATGCTTTTCGAACGTCC-3' (SEQ ID NO.), Respectively. (See Figure 8). The BamHl site was then altered to an EcoRV site using a binder. Fragment 2 is an HincII to EcoRI subfragment of approximately 1356 base pairs which codes for the VP2 gene of IBDV derived from plasmid 779-54.1. The large segment RNA Bursvac P1501 / 99MX M from IBDV? Cloned first as cDNA in two overlapping fragments using the first strand / second strand cDNA synthesis from Okayama-Berg. These clones were joined in a single restriction site to create plamid 2.40 / 84 # 3. Plasmid 779-54.1 was created by PCR in two parts using plasmid 2.40 / 84 # 3 as a template. The homosentide and antisense primers for the first half were 5, -GCCGGCGGCCGCGGATACGATCGGTCTGACCCCGG-3 'and 5' -TTGTGTGCACCGCGGAGTACCCC-3 ', respectively. The homosentide and antisense primers for the second half were 5 '-TCGCGAATCTATTCCAGGTGCCCC-3' and 5'-CGGAATTCTCCAATTTGGGATGTTGTAAGGCCGA-3 ', respectively. The two halves were united in a single Salí site. Fragment 3 is the polyadenylation signal PK of ILTV synthesized by PCR as a Sali fragment of 144 bp from the USDA USDA ILTV sensitization strain. The homosentide and antisense primers used for PCR were 5'-GGTGCCACGGTCGACGAGTGAAGGTTAAT-3 '(SEQ ID NO.) And '-AGACCGAGCAGAGTCGACGCGCGAAAG-3' (SEQ ID NO.), Respectively.

CLONE SUBGENOMICA 949-24. GAVE. Plasmid 949-24. Di was constructed for the purpose of generating recombinant HVT expressing a foreign DNA sequence and generating recombinant chimeric viral vaccine comprising the chimera of the short region of the disease virus.

P1501 / 99MX Marek and the long region of the turkey herpesvirus that expresses a foreign DNA sequence. The VP2 gene of infectious bursal disease virus (IBDV) was expressed under the control of the infectious laryngotracheitis virus (ILTV) glycoprotein D (gD) promoter. The VP2 gene of IBDV under the control of the gD promoter of ILTV was transcribed in the opposite direction of the UL52, UL54 and UL55 genes of HVT in the genomic DNA of HVT. The HVT DNA was derived from the cosmid 720-51.3 which is a subclone of the cosmid 407-32.1C1 (see figure 2, 5). The HVT fragment is a fragment of 34,700 base pairs of genomic HVT DNA. This region includes the BamHl fragments 11, 7, 8, 21, 6, 18 and approximately 1250 base pairs of fragment 13 and approximately 2600 base pairs of fragment 1. The Xhol site (Nucleotide # 1339-1344; SEQ ID NO. 12) was used for the insertion and expression of foreign HVT genes. (see Figure 3B). The cosmid was constructed by joining the restriction fragments (Sambrooks, et al., 1989) from the following sources. The vector is a NotW fragment of 8000 base pairs of pWEl5. The foreign DNA inserted into the Xhol site of HVT is as follows: fragment 1 is the gD promoter of ILTV synthesized by PCR as a fragment Pmel to Ba Hl of 530 pb from a USDA strain of ILTV sensitization. The homosentide and antisense primers for 3a reverse transcription and PCR were 5'- P1501 / 99MX GGTTTAAACAGCTGTACTACAGAGTAAC-3 '(SEQ ID NO.) And 5'- CGGATCCATAGCGTTGCGTACAAAG-3' (SEQ ID NO.), Respectively. (See Figure 7). Fragment 2 is an HincII to EcoRI fragment of approximately 1356 base pairs which codes for the VP2 gene of IBDV derived from plasmid 779-54.1. The large segment RNA Bursavac M IBDV was first cloned as cDNA into two overlapping fragments using the first strand / second strand cDNA synthesis of Okayama-Berg. These clone? joined in a unique restriction site to create plasmid 2.40 / 84 # 3. Plasmid 779-54.1 was created by PCR in two parts using plasmid 2.40 / 84 # 3 as a template. The homosense and antisense primers of the first half were 5 '-GCCGGCGGCCGCGGATACGATCGGTCTGACCCCGG-3' and 5'-TTGTGTGCACCGCGGAGTACCCC-3 ', respectively. The homosense and antisense primers of the second half were 5 '-TCGCGAATCTATTCCAGGTGCCCC-3' and 5'-CGGAATTCTCCAATTTGGGATGTTGTAAGGCCGA-3 ', respectively. The two halves were united in a single Salí site. Fragment 3 is the PK and polyadenylation signal ILTV synthesized by PCR as a 144 bp Sali fragment from the USDA USDA challenge strain. The sense and antisense primers used for PCR were '-GGTGCCACGGTCGACGAGTGAAGGTTAAT-3 '(SEQ ID NO.) And '-AGACCGAGCAGAGTCGACGCGCGAAAG-3' (SEQ ID NO.), Respectively.

P1501 / 99MX SUBGENOMIC CLONE 949-24.12. Plasmid 949-24.12 was constructed in order to generate recombinant HVT expressing the foreign DNA sequence and generating the recombinant chimeric viral vaccine comprising a chimera from the short region of the Marek's disease virus and the long region of the turkey herpesvirus expressing a strange DNA sequence. The VP2 gene of infectious bursal disease virus (IBDV) was expressed under the control of the glycoprotein promoter I (gl) of infectious laryngotracheitis virus (ILTV). The VP2 gene of IBDV under the control of the ILTV promoter gl transcribed in the opposite direction of the UL52, UL54 and UL55 genes of HVT in the genomic DNA of HVT. The HVT DNA was derived from the cosmid 720-51.3 which is a subclone of the cosmid 407-32.1C1 (see figure 2, 5). The HVT fragment e? a fragment of 34,700 base pairs of the genomic HVT DNA. This region includes the BamHl fragments 11, 7, 8, 21, 6, 18 and about 1250 pairs of fragment 13 ba and fragment 13 bases and approximately 2600 base pairs. The Xhol site (Nucleotide # 1339-1344; SEQ ID NO.12) was used for the insertion and expression of foreign HVT genes. (see Figure 3B). The cosmid was constructed by joining the restriction fragments (Sambrooks, et al., 1989) from the following sources. The vector is a Notl fragment of 8000 base pairs of pWEl5 .. The P1501 / 99MX. Extraneous DNA inserted into the Xhol site of HVT is as follows: fragment 1 is the ILTV gl promoter synthesized by PCR as a Pmel to B9H1 fragment of 279 bp from a USDA isolate strain of ILTV. The homosentide and antisense primers for PCR were 5 '-CAGTTTAAACTCGGATTCTGACTATTAC-3' (SEQ ID NO.) And 5 '-CGGATCCATGCTTTTCGAACGTCC-3' (SEQ ID NO. ), respectively (See Figure 8). Fragment 2 is an HincII to EcoRI fragment of approximately 1356 base pairs which codes for the VP2 gene of IBDV derived from plasmid 779-54.1. The large segment RNA Bursavac M IBDV was first cloned as cDNA into two overlapping fragments using the first strand / second strand cDNA synthesis of Okayama-Berg. These clones were joined in a single restriction site to create plasmid 2.40 / 84 # 3. Plasmid 779-54.1 was created by PCR in two parts using plasmid 2.40 / 84 # 3 as a template. The homosense and antisense primers of the first half were 5'-GCCGGCGGCCGCGGATACGATCGGTCTGACCCCGG-3 'and 5'-TTGTGTGCACCGCGGAGTACCCC-3', respectively. The homosense and antisense primers of the second half were 5 '-TCGCGAATCTATTCCAGGTGCCCC-3' and 5'-CGGAATTCTCCAATTTGGGATGTTGTAAGGCCGA-3 ', respectively. The two halves were united in a single Salí site. Fragment 3 is the PK and polyadenylation signal ILTV synthesized by PCR as a P1501 / 99HX fragment I left 144 bp from the USDA USDA strain of sensitization. The sense and antisense primers used for PCR were '-GGTGCCACGGTCGACGAAGGTTAAT-3' (SEQ ID NO.) And 5 '-AGACCGAGCAGGACGCGCGAAAG-3' (SEQ ID NO.), Respectively.

VECTOR OF HOMOLOGY 881-23. # 28: Plasmid 881-23. # 28 was constructed in order to generate recombinant chimeric HVT / MDV vaccine expressing a foreign DNA sequence. The lacZ gene of E. coli is expressed under the control of the HSV-1 TL promoter. The Newcastle disease virus (NDV) hemagglutinin (HN) gene is under the control of the PRV gX promoter and the Newcastle disease virus (NDV) fusion (F) gene is under the control of the immediate early promoter of HCMV. The HVT DNA is an Ascl subclone of cosmid 40732.1C1 (see Figures 2 and 5). The cosmid was built by the union of fragment? of restriction (Sambrooks, et al., 1989) from the following sources. The vector is an AscI fragment of approximately 2000 base pairs constructed from an AatlI to PvuII fragment at 2000 base pairs of pNEB193 (New England Biolabs, Inc.) whose ends are blunt with Klenow DNA polymerase and AscI ligands. inserted. The HVT fragment is an AscI to AscI fragment of approximately 8600 base pairs of HVT DNA Genomic P1501 / 99MX. This region includes fragments 10 and 21 of BamHI and approximately 1100 base pairs of fragment 6 and approximately 1300 base pairs of fragment 7. The Xhol site (Nucleotide # 1339-1344; SEQ ID NO: 12) was used for insertion and expression of foreign genes in HVT. (See Figure 3B). The foreign DNA within the Xhol site of the HVT genomic DNA is as follows: fragment 1 is a restriction subfragment Rsal to HaelII of approximately 266 base pairs of the N-restriction fragment BamH1 of HSV-1. Fragment 2 is a BamHi to Ball restriction fragment of approximately 3330 base pairs of plasmid pJF751 (Ferrari et al., 1985). Fragment 3 is a restriction subfragment SalI to BamHl of approximately 412 base pairs of the restriction fragment 10 of BamH1 from pRV (Lomniczi et al., 1984). Fragment 4 is an Avall to Nael restriction fragment of approximately 1811 base pairs? of the full-length NDV HN cDNA clone (strain Bl) The fragment 5 is a restriction subfragment Ndel to Salí of approximately 754 base pairs of the restriction fragment # 7 of PRV BamHl (Lomniczi et al., 1984) .Fragment 6 is a PstI to Avall restriction subfragment of approximately 1191 base pairs of the HCMV genomic Hbal E fragment (DR Thomsen, et al., 1981) .Fragment 7 is a BamHl restriction fragment.

P1501 / 99MX PstI of approximately 1812 base pairs of the full length NDV FD cDNA clone (strain Bl). The fragment 8 is a Smal to Smal restriction subfragment of approximately 784 base pairs of the Q fragment of BamHI restriction of HSV-1 (Mcgeoch, et al., 1985). The plasmid 881-23. # 28 was used together with S-HVY-145 according to THE TRANSFECTION OF DNA TO GENERATE RECOMBINANT VIRUS, for the reconstruction of the recombinant chimeric vaccine of HVT / MDV.

VECTOR OF HOMOLOG AT 883-10-A1: The homology vector 883-10-Al was constructed in order to insert the lacZ marker gene under the control of the novel chicken anemia virus promoter within the avian pustulation virus (FPV) ) in a unique Notl site. The EcoRI genomic fragment of FPV of 1.8 KB was used for the insertion of foreign DNA into the FPV. The Notl / Sfil lingula was inserted into a unique SnaBI site in the FPV fragment. The cassette was built using standard -ADN technique (Maniatis et al., 1982 and Sambrook et al., 1989), binding the restriction fragments from the following sources with the appropriate synthetic DNA sequences. The foreign DNA inserted into the Notl site of FPV is as follows: Fragment 1 is the CAV promoter synthesized by PCR as an EcoRI to BamHI fragment of 386 bp from the P1501 / 99 X strain CAV CL-1 (NVSL, Dr D.B. Snyder, Univ. Maryland; SEQ ID No. 23). Fragment 2 is a BamH1 to PvuII restriction fragment of approximately 3001 base pairs of plasmid pJF751 (Ferrari et al., 1985). The fragment 3 a restriction subfragment Ndel a Salí of approximately 754 base pairs of fragment # 7 of restriction BamHl of PRV (Lomniczi et al., 1984). The recombinant avian pustulation virus was regenerated by the HOMOLOGA RECOMBINATION procedure to generate RECOMBINANT FPV.

TRANSIENT TRANSFER TEST: Chicken embryo fibroblast cells or QT-35 cells at 80-90% confluence in 6-cm dishes were subjected to removal of the growth medium and 5 ml of maintenance medium was added. 10-20 μg of the plasmid DNA was diluted in water and CaCl 2 added at a concentration of 0.25 M and a volume of 600 μl. To this, 600 μl of 2X HEPES buffer (0.28 M NaCl, 50 mM HEPES (N-2-hydroxyethylpiperazine-N "-2-ethanesulfonic acid), 1.5 mM Na2HP04, pH 7.05) was added allowing one minute to incubate at room temperature and then this solution is divided into two aliquots of 600 μl and dropped onto two 6 cm dishes, after 3 hours the solution is removed and 10% glycerol is added in PBS and allowed to stand on the cells for 1 minute. This is removed, the cells are washed once with PBS, and fed back with 5 ml of P1501 / 99MX maintenance medium. After 24-48 hours, the cells are harvested, pelleted at 2.8K in a Sorvall RT7, and resuspended in 500 μl PBS. The samples are subjected to three cycles of freezing and thawing and the cell debris is centrifuged in a microcentrifuge. The duplicate sample consisting of 11 μ of each supernatant is tested in a microtitre plate with 100 μl of ONPG assay solution (0.8 mg / ml orthonitro-phenyl-galactoside, 60 mM Na2HP04, 40 mM NaH2P04, 10 mM KCl, 1 mM MgSO4, 50 mM β-mercaptoethanol, pH 7.5), incubated at 37 ° C and readings taken at 415 nm with a BioRad microplate reader model 450 every fifteen minutes.

PLASMID 388-65.2. Plasmid 388-65.2 was constructed for the purpose of generating an immediate early promoter (IE) HCMV to express a foreign DNA sequence.

The E. coli β-galactosidase gene was expressed under the control of the HCMV IE promoter. The plasmid was constructed by joining the restriction fragments (Sambrooks, et al., 1989) from the following sources. The vector is an AatlI to PvuII fragment of approximately 2000 base pairs of pNEB193 (New England Biolags, Inc.). Fragment 1 is a PstI to Avall subfragment of 1149 base pairs of the Xbal E fragment of the Towne HCMV strain. Fragment 2 is a restriction fragment of BamHl to PvuII from P1501 / 99MX approximately 3001 base pairs of plasmid pJF751 (Ferrari et al., 1985). Fragment 3 is a restriction subfragment Ndel to Salí of approximately 754 base pairs of fragment # 7 of PRV BamHl restriction (Lomniczi et al., 1984) Plasmid 388-65.2 was used according to the TRANSITORY TRANSFER TEST to MEASURE the strength of the CAV promoter.

PLASMIDE 850-25.18. Plasmid 850-25.18 was constructed for the purpose of generating a novel CAV promoter to express a foreign DNA sequence. The E. coli β-galactosidase gene is expressed under the control of the modified chicken anemia virus promoter. The plasmid was constructed by joining the restriction fragments (Sambrooks, et al., 1989) from the following sources. The vector is an AatlI to PvuII fragment of approximately 2000 base pairs of pNEBl93 (New England Biolags, Inc.). Fragment 1 of the CAV promoter synthesized by PCR as an EcoRI to BamHI fragment of 854 bp from the CAV CL-1 strain (NVSL; Dr DB Snyder, Univ. Maryland;) using PCR primers 51-ATCGAATTCCGAGTGGTTACTATTC-3 '(SEQ ID NO 24) and 5'-CGTGGATCCATCTTACAGTCTTATAC-3' (SEQ ID NO 25). Fragment 2 is a restriction fragment of BamHI to PvuII of approximately 3001 base pairs? of plasmid pJF751 (Ferrari et al., 1985). The fragment P1501 / 99MX 3 is a restriction subfragment Ndel a Salí of approximately 754 base pairs of fragment # 7 of restriction BamHl of PRV (Lomniczi et al., 1984) Plasmid 350-25.18 was used according to the TEST TRANSFER TRANSIENT MEASURE the strength of the CAV promoter.

PLASMID 850-69.1. Plasmid 850-69.1 was constructed for the purpose of generating a novel CAV promoter to express a foreign DNA sequence. The E. coli β-galactosidase gene is expressed under the control of the modified chicken anemia virus promoter. The plasmid was constructed by joining the restriction fragments (Sambrooks, et al., 1989) from the following sources. The vector is an AatlI to PvuII fragment of approximately 2000 base pairs of pNEB193 (New England Biolags, Inc.). Fragment 1 is the CAV promoter synthesized by PCR as an EcoRI to BamHI fragment of 854 bp from the CAV strain of CL-1 (NVSL; Dr DB Snyder, Univ. Maryland;) using the PCR primers 5'-ATCGAATTCCGAGTGGTTACTATTC -3 '(SEQ ID NO 24) and 5'-CGTGGATCCATCTTACAGTCTTATAC-3' (SEQ ID NO 25). The HindIII site near the BamHl site is within the CAV apoptin reading frame. The HindIII site was filled to destroy the apoptin reading frame. Fragment 2 is a restriction fragment of BamHl to PvuII of approximately 3001 P1501 / 99MX base pairs of plasmid pJF751 (Ferrari et al., 1985). The fragment 3 is a restriction subfragment Ndel to Salí of approximately 754 base pairs of fragment # 7 of restriction BamHl of PRV (Lomniczi et al., 1984) The plamid 850-69.1 was used according to the TEST OF TRANSIENT TRANSFECTION TO MEASURE the strength of the CAV promoter.

PLASMID 850-80.A2. The plasmid 850-80. A2 was constructed for the purpose of generating a novel CAV promoter to express a foreign DNA sequence. The E. coli β-galactosidase gene is expressed under the control of the modified chicken anemia virus promoter. The plasmid was constructed by joining the restriction fragments (Sambrooks, et al., 1989) from the following sources. The vector is an AatlI to PvuII fragment of approximately 2000 base pairs of pNEB193 (New England Biolags, Inc.). Fragment 1 is the CAV promoter synthesized by PCR as an EcoRI to BamHI fragment of 381 bp (SEQ ID NO.23) from the CAV CL-1 strain (NVSL, Dr DB Snyder, Univ. Maryland; PCR primers 5 '-GTTCGGATCCATCCTCCCGGACCGCCTTG-3' (SEQ ID NO 26) and 5 '-GCGGAAGAGCGCCAATACG-3' (SEQ ID NO 27). The fragment 2 e? a restriction fragment of BamH1 to PvuII of approximately 3001 base pairs of plasmid pJF751 (Ferrari et al., 1985). The fragment 3 is a restriction subfragment Ndel a Salí de P1501 / 99 X approximately 754 base pairs of fragment # 7 of PRV BamHl restriction (Lomniczi et al., 1984) Plasmid 850-80.2 was used according to the TRANSITORY TRANSFER TEST to MEASURE the strength of the CAV promoter.

PLASMID 883-11. TO 5. The plamido 883-11. A5? E constructed for purposes of generating a novel CAV promoter to express a foreign DNA sequence. The E. coli β-galactosidase gene is expressed under the control of the modified chicken anemia virus promoter. The plasmid was constructed by joining the fragment? of restriction (Sambrooks, et al., 1989) from the following sources. The vector is an AatlI to PvuII fragment of approximately 2000 base pairs of pNEBl93 (New England Biolags, Inc.). Fragment 1 is the CAV promoter synthesized by PCR as an EcoRI to BamHI fragment of 381 bp from the CAV CL-1 strain (NVSL, Dr DB Snyder, Univ. Maryland;) using the 5 'PCR primers -GTTCGGATCCATCCACCCGGACCGCCTTG- 3 '(SEQ ID NO 28) and 5'-GCGGAAGAGCGCCAATACG-3' (SEQ ID NO 27). Fragment 2 is a restriction fragment of BamH1 to PvuII of approximately 3001 base pairs of plasmid pJF751 (Ferrari et al., 1985). Fragment 3 is a restriction subfragment Ndel to Salí of approximately 754 base pairs of fragment # 7 of restriction BamHl of PRV (Lomniczi et al., 1984) P1501 / 99MX plasmid 883-11. A5 was used according to the TRANSITORY TRANSFER TEST to MEASURE the strength of the CAV promoter.

VECTOR OF HOMOLOGY 849-69. Al: Plasmid 849-69. To the It was constructed in order to generate recombinant viral vector of porcine rashes expressing a foreign DNA sequence. The lacZ gene of E. coli is expressed under the control of the LP1 promoter. The quail interferon gene type 1 (qIFN-1) is expressed under the control of the LP2EP2 promoter. The ORF qIFN-1 (594 base pairs) was generated by PCR (as described in Example 23B) with compatible ends EcorRI and BamHI to be cloned into the homology vector SPV 752-22.1 (see WO 96/22363). The plasmid 849-69. Al was used together with S-SPV-001 according to THE RECOMBINATION PROCEDURE HOMOLOGO TO GENERATE FPV or RECOMBINANT SPV for the reconstruction of the recombinant viral vector SPV Example 19A S-HVY-145 Recombinant chimeric virus HVT / MDV. S-HVY-145 is a recombinant chimeric virus that contains MDV and HVT genomic sequences that in a vaccine formulation protect against Marek's disease and are produced by the combination of cosmids of the genes that contain MDV genomic DNA that codes for antigens protectors P1501 / 99MX relevant to serotype 1 of virulent MDV and cosmids of genomic DNA of HVT according to the PROCEDURE FOR GENERATING RECOMBINANT HERPESVIRUS FROM SUBGENOMIC TRANSLATING FRAGMENTS. The resulting virus is a vaccine that has the protective immune response for the virulent serotype 1 of MDV and the attenuated growth characteristics of the HVT. In one embodiment, the chimeric virus contains the MDV genes of the single short region and the HVT genes of the single long region and is useful as a vaccine against Marek's disease in chickens. In another embodiment, the chimeric virus contains the MDV genes of the short region and the gene. HVT of the long region and is useful as a vaccine against Marek's disease in chickens. The MDV protective antigens within the single short region (gD, gE and gl) produce a protective immune response against MDV, while IO? Virulence elements present in the single long region of MDV (55, 56, 57) are replaced by unique long sequences attenuating HVT. The result is an attenuated virus vaccine that protects against Marek's disease. Multivalent protection against Marek's disease, infectious laryngotracheitis, infectious bursal disease, Newcastle disease or other poultry pathogen is achieved by inserting the gB, gD and gl genes of ILTV, the VP2 gene of IBDV, the P1501 / 99MX NDV HN and F genes or an antigen gene from a poultry pathogen to an Xhol site converted to a PacI site or to a Notl site in the EcoRl # 9 fragment (BamHl # 10) within the region long unique recombinant HVT / MDV virus (Figures 2 and 5). A cosmid was constructed that contained the entire short MDV region. The genomic DNA of MDV contains several Smal sites in the unique long region and the internal and terminal repetitions of the virus, but no Smal site within the short region of the viru? . The complete short region of MDV was isolated by a partial restriction digestion of the MDV DNA with Smal. The DNA fragment of about 29,000 to 33,000 base pairs was isolated and cloned into a blunt-ended site of the cosmid vector pWEl5. To generate S-HVY-145, a recombinant HVT / MDV chimeric virus, the cosmid containing the short MDV region was combined with the long HVT region containing concomers according to the PROCEDURE FOR GENERATING RECOMBINANT HERPESVIRUS FROM SUBGENOMICAL TRANSLATING FRAGMENTS. The following combination of subgenomic clones and enzymes were used: 407-32.2C3 with Notl, 172-07.BA2 with BamHl, 407-32.5G6 with Notl, 407-32.1C1 with Notl and 739-27.16 with Notl. The resulting virus vaccine provides greater protection against Marek's disease or as P1501 / 99MX a multivalent vaccine against Marek's disease and infectious laryngotracheitis, infectious bursal disease, Newcastle disease or other bird pathogen. This vaccine is superior due to the expression of the MDV genes in the HVT / MDV chimera vaccine that is safer and provides better protection against Marek's disease than the currently available vaccines containing HVT and MDV type 2 (SB-1) or HVT. Second, the expression of MDV glycoprotein genes can be demonstrated in the absence of homologous HVT genes for diagnostic and regulatory purposes. This is useful since antibodies to an MDV glycoprotein will cross-react with the homologous HVT glycoprotein. Finally, a recombinant HVT / MDV virus that contains a single copy of each glycoprotein gene is more stable than a recombinant virus that contains two copies of the homologous glycoprotein gene from HVT and MDV that can be canceled by homologous recombination. In an alternative embodiment, the cosmids containing MDV protective antigen genes from the single long region (gB and gC of MDV) are combined with cosmids having HVT gene sequences from the short and long regions, effectively avoiding the MDV virulence genes at the junction of single long region / internal repeat and at the single long region junction / repeat P1501 / 99 X terminal (55, 56 and 57). Strain SB-1 in a serotype 2 of MDV with attenuated pathogenicity. Vaccination with a combination of live HVT and SB-1 viruses better protects against virulent MDV sensitization than the vaccine with any of the viruses alone. In an alternative embodiment of the present invention a recombinant virus vaccine comprises protective antigen genes of the virulent serotype 2 MDV combined with the attenuating genes of non-virulent MDV serotypes 1 and 3, for example SB-1 and HVT. The genomic DNA corresponding to the single long region is contributed with the SB-1 serotype. The genomic DNA corresponding to the single short region is contributed with serotype HVT. Three major glycoprotein antigens (gB, gA and gD) of MDV serotype 1 are inserted into the single short region of the virus. The recombinant virus is constructed using the subgenomic clone HVT 721-38.1J to reconstruct the single short region. The subgenomic clone 721-38. ÍJ contains an insertion of the gene? gB, gA and gD of MDV. A large molar excess of these clones is cotransfected with a subinfectious dose of the Sb-1 genomic DNA. To determine the appropriate subinfectious dose, transfection of SB-1 is decreased in titer to a dose that no longer provides virus plaques in the cell culture.

P1501 / 99MX This dose contains subgenomic fragments that expand the single long region of SB-1 that recombine with the unique short subgenomic clones of HVT. Therefore, a virus resulting from recombination between the overlapping homologous regions of the subgenomic fragments HVT and SB-1 is highly favored. Alternatively, the SB-1 genomic fragments of the single long region are subcloned into cosmid vectors. A recombinant virus containing the unique long region of SB-1, the unique short region of HVT with genes gB, gA and gD of MDV was produced using the PROCEDURE TO GENERATE RECOMBINANT HERPESVIRUS FROM SUBGENOMIC TRANSLATING FRAGMENTS. This method is also used with a subgenomic HVT clone to insert antigen genes from other avian pathogens, which include, but are not limited to, infectious laryngotracheitis virus, Newcastle disease virus and infectious bursal disease virus.

Example 19B S-HVY-149 S-HVY-149 is a recombinant chimeric virus comprising a chimera from the short region of the Marek's disease virus and the long region of the turkey herpesvirus. S-HVY-149 is a recombinant chimeric viral vaccine that comprises foreign DNA from the glycoprotein D (gD) virus of P1501 / 99 X infectious laryngotracheitis and genetics of glycoprotein I (gl) inserted into an Xhol site in the EcoRl # 9 fragment within the single long region of the chimeric virus. The genes gD and gl of the viru? ILT remain under the control of the gD and gl promoters of the ILT virus. The recombinant chimeric viral vaccine is useful against the sensitization with the virulent Marek's disease virus and against the infectious laryngotracheitis virus. To generate S-HVY-149, the following combination of subgenomic clones and enzymes was used in the PROCEDURE TO GENERATE RECOMBINANT HERPESVIRUS FROM SUBGENOMIC TRANSLATING FRAGMENTS: 407-32.2C3 with Notl, 172-07. BA2 with BamHl, 407-32.5G6 with Notl, 852-52.114 with Notl and 739-277.16 with Notl. S-HVY-149 was purified by plating and plaque purification and purity was tested by BLACK PLATE TEST. S-HVY-149 was 100% pure by the BLACK PLATE ESSAY using convalescent ILT virus antisera.

Example 19C S-HVY-151 S-HVY-151 is a recombinant chimeric comprising a chimera from the short region of the Marek's disease virus and the long region of the turkey herpesvirus. S-HVY-151 is a chimeric viral vaccine P1501 / 99 Recombinant X comprising foreign DNA from the lacZ gene of E. coli inserted into an Xhol site in the EcoRl # 9 fragment within the single long region of the chimeric virus. The lacZ gene of E. coli is under the control of the gV promoter of PRV. The recombinant chimeric viral vaccine is useful against sensitization with the virulent Marek's disease virus. S-HVY-151 was derived from S-HVY-145. This was achieved using the homology vector 301-07. AND # D1 and the S-HVY-145 virus in the TRANSFECTION OF DNA TO GENERATE RECOMBINANT VIRUSES, inside the primary chicken embryo fibroblast cells (CEF). A blue virus obtained from the transfection storage material is purified by successive plate purifications using the BLUOGAL SCREENING PROCEDURE FOR RECOMBINANT HERPESVIRUS. S-HVY-151 is useful as a vaccine against sensitization with the Marek's disease virus.

Example 19D S-HVY-152 S-HVY-152 is a recombinant chimeric virus comprising a chimera from the short region of the Marek's disease virus and the long region of the turkey herpesvirus. S-HVY-152 is a recombinant chimeric viral vaccine comprising foreign DNA from the lacZ gene of E. coli and the genes of P1501 / 99MX glycoprotein D (gD) of infectious laryngotracheitis virus (ILT) and glycoprotein I (gl) at an Xhol site in the EcoRl fragment # 9 within the single long region of the chimeric virus. The lacZ gene of E. coli is under the control of the gV promoter of PRV. The gD and gl genes of the ILT virus are under the control of the gD and gl promoters of the ILT virus, respectively. S-HVY-152 was derived from S-HVY-145. This is achieved by using homology vectors 864-74, 18 and S-HVY-145 viruses in the DNA TRANSFECTION PROCEDURE FOR GENERATING RECOMBINANT VIRUSES within primary cells of chicken embryo fibroblast (CEF). S-HVY-152 was tested for purity by BLACK PLATE TEST using convalescent ILT virus antisera and rabbit anti-β-galactosidase antisera and purified by successive plaque purifications and BLACK PLATE TEST. S-HVY-152 is useful as a vaccine against sensitization with Marek's disease virus and infectious laryngotracheitis virus.

Example 19E S-HVY-153 S-HVY-153 is a recombinant chimeric virus comprising a chimera from the short region of the Marek's disease virus and the long region of the P1501 / 99MX turkey herpesvirus. S-HVY-153 is a recombinant chimeric viral vaccine comprising foreign DNA from the lacZ gene of E. coli and the genes of hemagglutinin (HN) and fu-tion (F) of Newcastle disease virus (NDV) inserted within of the Xhol site in the EcoRl fragment # 9 within the single long region of the chimeric virus. The lacZ gene of E. coli is under the control of the HSV-1 TK promoter. The NDV HN gene is under the control of the PRV gX promoter and the F NDV gene is under the control of the intermediate early HCMV promoter. S-HVY-153 was derived from S-HVY-145. This was achieved using the homology vector 881-23. # 28 and S-HVY-145 virus in the TRANSFECTION OF DNA TO GENERATE RECOMBINANT VIRUSES within primary cells of chicken embryo fibroblast (CEF). A red plaque virus obtained from the transfection storage material was purified by successive plate purifications using the BLUOGAL TAMIZED 0 CPRG FOR RECOMBINANT HERPESVIRUS procedure and selecting the blue or red plates. S-HVY-153 is tested for purity by BLACK PLATE TEST using anti-β-galactosidase and anti-NDV antibodies S-HVY-153 is useful as a vaccine against sensitization with Marek's disease virus and disease virus of Newcastle.

P1501 / 99 X Example 20 Recombinant HVT expressing chicken myelomonocyte growth factor (cMGF), chicken interferon (cIFN) or quail interferon (qlFN) is useful as a vaccine against the Marek's disease virus and is also useful for strengthen the immune response against other diseases of poultry. Chicken myelomonocyte growth factor (cMGF) is related to G-CSF and interleukin-6 protein (58) and Type 1 chicken interferon (cIFN) that is homologous to mammalian type 1 interferon (59). When used in combination with the vaccines described in the previous examples, S-HVT-144 or HVT expressing cIFN are useful for providing improved cell-mediated immunity or mucosal or humoral immunity against viruses that cause avian diseases, including enunciatively: Marek's disease virus, Newcastle disease virus, infectious laryngotracheitis virus, infectious bronchitis virus, infectious bursal disease virus. Recombinant HVTs expressing cMGF or cIFN are useful to provide improved immunity against avian disease caused by the organisms described in Example 15.

P1501 / 99 X 14 * 7 Example 20A S-HVT-144 S-HVT-144 is a recombinant turkey herpesvirus containing the chicken myelomonocyte growth factor (cMGF) gene inserted into an Xhol site converted to a PacI site in the EcoRl fragment # 9 within the unique long region of the HVT. The cMGF gene is in the transcriptional orientation opposite to the open reading frame (ORF A) within the EcoRl fragment # 9 of the HVT genome (Figure 4; SEQ ID NOs: 12 and 13). The cMGF gene is expressed from a human cytomegalovirus immediate early promoter. S-HVT-144 is useful as a vaccine in poultry against Marek's disease. S-HVT-144 was constructed according to the PROCEDURE TO GENERATE RECOMBINANT HERPESVIRUS FROM SUBGENOMICAL TRANSFER FRAGMENTS. The following combination of subgenomic clones and enzymes was used: 407-32.2C3 with Notl, 172-07. BA2 with BamHl, 407-32.5G6 with Notl, 672-07. C40 with Notl, 672-01. A40 with Notl, 751-87. A8 with Ase I, 415-09. BAl with BamHl.

Example 20B recombinant HVT expressing chicken interferon. A herpesviru? of recombinant turkey that P1501 / 99MX contains the Type 1 chicken interferon gene (cIFN) inserted into an Xhol site converted to a PacI site in the EcoRl # 9 fragment within the single long region of HVT. The cIFN gene is expressed from a human cytomegalovirus immediate early promoter. Recombinant HVT expressing cIFN is useful as a vaccine in poultry against Marek's disease. The recombinant HVT expressing cIFN is constructed according to the PROCEDURE FOR GENERATING RECOMBINANT HERPESVIRUS FROM SUBGENOMIC TRANSLATING FRAGMENTS. The following combination of subgenomic clones and enzymes was used: 407-32.2C3 with Notl, 172-07. BA2 with BamHl, 407-32.5G6 with Notl, 672-07. C40 with Notl, 672-01. A40 with Notl, 761-07. Al with Ase I, 415-09. BAl with BamHl. Recombinant HVT expressing avian cytokines is combined with HVT that expresses genes for avian disease antigens to improve the immune response. Additional cytokines that are expressed in HVT and have immune stimulatory effects include, but are not limited to: beta transformation growth factor, family epidermal growth factor, fibroblast growth factors, hepatocyte growth factor, similar growth factors to insulin, nerve growth factor B, platelet-derived growth factor, factor of P1501 / 99MX vascular endothelial growth, interleukin 1, IL-1 receptor antagonist, interleukin 2, interleukin 3, interleukin 4, interleukin 5, interleukin 6, soluble receptor IL-6, interleukin 7, interleukin 8, interleukin 9, interleukin 10, interleukin 11, interleukin 12, interleukin 13, angiogenin, chemokines, colony stimulating factors, granulocyte-macrophage colony stimulating factors, eri tropoietin, interferon, interferon gamma, leukemia inhibitory factor, oncostatin M, pleiotrofin, leukocyte protease inhibitor secretory, hemocytoblast factor, tumor necrosis factors and soluble TNF receptors. These cytokines come from avian species or other animals, including man, cattle, horses, cats, dogs or porcionos.

Example 20C Recombinant HVT expressing genes of Marek's disease virus and chicken interferon gene. A turkey recombinant herpesvirus contains the type 1 chicken interferon gene (cIFN) inserted in an Xhol site converted to a PacI site in the EcoRl fragment # 9 within the single long region of the HVT and also contains the genes gA, gD and gB of MDV inserted into a unique Stul site converted into the HindIII site in the US2 gene of the HVT. The cIFN gene is expressed from a promoter Immediate early P1501 / 99MX human cytomegalovirus. The MDV genes are expressed from endogenous MDV promoters. The recombinant HVT expressing cIFN and gA, gB and gD of MDV is useful as a vaccine to improve the immune response in poultry against Marek's disease. The recombinant HVT expressing the MDV genes and the cIFN gene is constructed according to the PROCEDURE FOR GENERATING RECOMBINANT HERPESVIRUS FROM SUBGENOMICAL TRANSLATING FRAGMENTS. The following combinations of subgenomic clones and enzymes are used: 407-32.2C3 with Notl, 172-07. BA2 with BamHl, 407-32.5G6 with Notl, 672-07. C40 with Notl, 672-01. A40 with Notl, 761-07. Al with Ase I, 721-38. ÍJ.

Example 20D Recombinant HVT expressing Marek's disease virus genes, Newcastle disease virus genes and chicken interferon gene. A recombinant turkey herpesvirus recombinant contains the type 1 chicken interferon gene (cIFN) within the Xhol site converted to a PacI site in the EcoRl fragment # 9 within the single long region of the HVT and also contains the genes gA, gD and gB of MDV and to the HN and F genes of NDV inserted in a single Stul site converted to a HindIII site in the US2 gene of HVT. The cIFN gene is expressed from an immediate early promoter of P1501 / 99MX human cytomegalovirus. The -MDV genes are expressed from endogenous MDV promoters. The NDV gene is used to control the gV promoter of PRV and the F gene of NDV is under the control of the immediate early promoter HCMV. The recombinant HVT expressing cIFN and gA, gB and gD of MDV is useful as a vaccine to improve the immune response in livestock birds against Marek's disease and Newcastle disease. The recombinant HVT expressing the MDV genes, NDV genes and cIFN is constructed according to the PROCEDURE FOR GENERATING RECOMBINANT HERPESVIRUS FROM SUBGENOMIC TRANSLATING FRAGMENTS. The following combination of subgenomic clones and enzymes was used: 407-32.2C3 with Notl, 172-07. BA2 with BamHl, 407-32.5G6 with Notl, 672-07. C40 with Notl, 672-01. A40 with Notl, 761-07. Al with Ase I, 722-60. E2.

Example 20E Recombinant HVT expressing genes of Marek's disease virus and chicken myelomonocytic growth factor gene. A recombinant turkey herpesvirus contains the chicken myelomonocyte growth factor (cMGF) gene within an Xhol site converted to a PacI site in the EcoRl # 9 fragment within the single long region of the HVT and also contains the gA genes, gD and gB of MDV inserted into a single Stul site converted to the HindIII site in the US2 gene of P1501 / 99 X HVT. The cMGF gene is expressed from a human cytomegalovirus immediate early promoter. The MDV genes are expressed from endogenous MDV promoters. Recombinant HVT expressing cMGF and gA, gB and gD of MDV is useful as a vaccine with an improved immune response in poultry against Marek's disease. The recombinant HVT expressing the cMGF gene and the MDV genes is constructed according to the PROCEDURE FOR GENERATING RECOMBINANT HERPESVIRUS FROM SUBGENOMIC TRANSLATING FRAGMENTS. The following combination of subgenomic clones and enzymes is used: 407-32.2C3 with Notl, 172-07. BA2 with BamHl, 407-32.5G6 with Notl, 672-07. C40 with Notl, 672-01. A40 with Notl, 751-87. A8 with Ase I, 721-38.1J.

A 20F recombinant HVT expressing Marek's disease virus genes, Newcastle disease virus genes and chicken myelomonocytic growth factor gene. A recombinant turkey herpesvirus contains the chicken myelomonocyte growth factor (cGMF) gene inserted in an Xhol site converted to a Pací site in the EcoRl fragment # 9 within the single long region of the HVT and also contains the gA genes, gD and gB of the MDV and the NDV HN and F genes inserted into the single converted Stul site P1501 / 99MX in the HindIII site of the US2 HVT gene. The cGMF gene is expressed from a human cytomegalovirus immediate early promoter. The MDV genes are expressed from endogenous MDV promoters. The NDV HN gene is under the control of the gV promoter of PRV and the F gene of NDV is under the control of the immediate early promoter HCMV. Recombinant HVT expressing the genes cIFN and gA, gB and gD of MDV is useful as a vaccine to improve the immune response in livestock birds against Marek's disease and Newcastle disease. The recombinant HVT that expresses the MDV genes, NDV genes and cGMF gene is constructed according to the PROCEDURE FOR GENERATING RECOMBINANT HERPESVIRUS FROM SUBGENOMICAL TRANSLATING FRAGMENTS. The following combination of subgenomic clones and enzymes was used: 407-32.2C3 with Notl, 172-07. BA2 with BamHl, 407-32.5G6 with Notl, 672-07. C40 with Notl, 672-01. A40 with Notl, 751-87. A8, 722-60. E2.

She uses recombinant turkey herpesvirus that expresses antigens from microorganisms that cause the disease. Herpesviru? of recombinant turkey (HVT) and useful for expressing antigens from microorganisms that cause the disease of animals as well as avian species. The HVT Recombinant P1501 / 99MX is used as a vaccine in animals, including, but not limited to, humans, equines, cattle, swine, canines and felines. Recombinant HVT is useful as a vaccine against equine diseases when foreign antigens of disease or disease organisms are expressed in the HVT vector, including but not limited to: equine influenza, equine herpesvirus-1 and equine herpesvirus-4. Recombinant HVT is useful as a vaccine against bovine diseases when the foreign antigens of the following diseases or organisms that cause them are expressed in the HVT vector, including, but not limited to: bovine herpesvirus type 1, bovine viral diarrhea virus, virus bovine respiratory syncytial virus, bovine parainfluenza virus. Recombinant HVT is useful as a vaccine against swine diseases when the foreign antigens of the following diseases or disease-causing organisms are expressed in the vector of HVT, including but not limited to: pseudorabies virus, porcine reproductive and respiratory syndrome (PRRS / SIRS), cholera virus in pigs, swine influenza virus, porcine parvovirus, porcine rotavirus. Recombinant HVT is useful as a vaccine against feline and canine diseases when the foreign antigens of the following diseases or organisms causing diseases P1501 / 99 X are expressed in the HVT vector, including, but not limited to: herpesviru? feline, feline leukemia virus, feline immunodeficiency virus and Dirofilaria immitis (heartworm). Disease-causing microorganisms in dogs include, but are not limited to: canine herpes virus, canine disorder, canine adenovirus type 1 (hepatitis), adenovirus type 2 (respiratory disease), parainfluenza, Leptospira canicola, icterohemorrhage, parvovirus, coronavirus, Borrelia burgdorferi, herpesvirus canine, Bordetella bronchiseptic, Dirofilaria immi tis (heartworm) and rabies virus.

Eg 22 Human vaccines that use herpesviru? of recombinant turkey as a vector. Recombinant turkey herpesvirus (HVT) is useful as a vaccine against human diseases. For example, human influenza is a rapidly evolving virus whose viral neutralizing epitopes change rapidly. A useful recombinant HVT vaccine is one in which influenza neutralizing epitopes change rapidly to protect against new strains of influenza. The human influenza HA and NA genes are cloned using polymerase chain reaction within the recombinant HVT. Recombinant HVT is useful P1501 / 99 X as a vaccine against other human diseases when the foreign antigens from the following diseases or disease-causing organisms are expressed in the vector VHT: core and surface antigens of hepatitis B virus, hepatitis C virus, human immunodeficiency, herpes simplex-1 virus, herpes simplex-2 virus, human cytomegalovirus, Epstein-Barr virus, Varicella-Zoster virus, human herpesvirus 6, human herpesvirus 7, human influenza, measles virus, hantaan virus, pneumonia, rhinovirus, poliovirus, human respiratory syncytial virus, retrovirus, human T cell leukemia virus, rabies virus, mumps virus, malaria (Plasmodium falciparum), Bordetella pertussis, Diphtheria, Rickettsia prowazekii, Borrelia bergdorferi, tetanus toxoid, antigens of malignant tumor. Recombinant HVT expressing human cytokines is combined with HVT that expresses genes for human disease antigens in order to improve the immune response. Additional cytokines, including but not limited to: transforming growth factor beta, family of epidermal growth factor, fibroblast growth factors, hepatocyte growth factor, insulin-like growth factors, growth factor B nerve, factor P1501 / 99MX of platelet-derived growth, vascular endothelial growth factor, interleukin 1, IL-1 receptor antagonist, interleukin 2, interleukin 3, interleukin 4, interleukin 5, interleukin 6, soluble receptor IL-6, interleukin 7, interleukin 8 , interleukin 9, interleukin 10, interleukin 11, interleukin 12, interleukin 13, angiogenin, chemokines, colony-stimulating factors, granulocyte-macrophage colony stimulating factors, heru tropoietin, interferon, gamma interferon, leukemia inhibitory factor, oncostatin M, Pleiotrofin, inhibitors of secretory leukocyte proteases, hemocytoblast factor, tumor necrosis factors, and soluble TNF receptors from humans and other animals are expressed in HVT and have immune stimulatory effects.

Example 23A Improved production of recombinant turkey herpesvirus vaccine. Cytokines, for example interferons and interleukins, inhibit the replication of viruses in cell culture and in animals. The inhibition of production of interleukin production or cellular interferon improves the growth of recombinant HVT in cell cultures. Chicken Interferon Type 1 (cIFN) P1501 / 99 X expressed from a recombinant porcine pustulation vector was added to chicken embryo fibroblast (CEF) cell cultures and infected with S-HVT-012 expressing β-galactosidase. The cIFN added to the cell culture medium reduced both β-galactosidase expression and the S-HVT-012 titer in a dose-dependent manner. This result indicates that the growth of HVT is limited by the exogenous addition of chicken interferon. Several strategies are used to improve the growth of HVT in CEF cells by removing or inactivating chicken interferon activity in CEF cells. In one embodiment, a chicken interferon neutralizing antibody is added to the culture medium to inhibit chicken interferon activity and enhance the growth of recombinant HVT in CEF cell culture. Anti-cIFN s * e antibody is derived from mouse or rabbit sera from animals injected with chicken interferon protein, preferably cIFN comes from a recombinant porcine pustulation virus expressing chicken interferon. Poxviruses or poxivirus secrete cytokine inhibitory proteins as a strategy of immune evasion. One type of immune evasion mechanism of poxivirus involves the soluble receptor of poxivirus for interleukins, interferon or P1501 / 99MX tumor necrosis factors that inactivates cytokines and allows viral replication (60). In one embodiment of the invention, avian pustulation virus is useful as a source of inhibition proteins of chicken interferon and other immune evasion proteins. Conditioned media from CEF cell cultures infected with FPV are added to CEF cells infected with HVT to inhibit interferon activity and increase the titer of HVT. In another embodiment, the recombinant chicken interferon inhibitory protein or another poxivirus immune evading protein is expressed in a vector in combination with an HVT vaccine composition to increase the HVT titer. Chicken embryo fibroblast cells have been genetically engineered to express foreign genes (61). In another embodiment, an interferon-negative CEF cell line is constructed by introducing a vector that expresses a gene that * codes for an antisense RNA for, chicken interferon within the CEF cell line. The recombinant HVT grown in an interferon-negative CEF cell line demonstrates the improved virus titers compared to the culture of HVT in a CEF cell line that produces interferon. In another embodiment, a CEF cell line positive for chicken myelomonocytic growth factor (cMGF) P1501 / 99 X e build by the introduction of a vector that expresses the cMGF gene within the CEF cells. The recombinant HVT cultured in a CEF cMGF-positive cell line demonstrates better virus titers compared to the HVT cultured in the negative CEF cMGF cell line. The recombinant HVT of the present invention is useful as a vaccine against Marek's disease and against other diseases outlined in the previous examples. A higher efficiency in the growth of recombinant HVT in CEF cells is useful in the production of the vaccine.

EXAMPLE 23B Cloning of the Quail Interfinger, type 1 ^ Southern blot analysis showed that a fragment of the 32P-labeled chicken interferon gene 1 was hybridized to a 5.0 kb BamHl genomic DNA fragment from the quail cell line QT- 35 Based on this information, the PCR method was used with primers homologous to chicken interferon, type 1, to clone the quail interferon gene, type 1. (PCR primers were: 5'-TGTACAGATCTCACCATGGCTGTGCCTGCAAGC-3 '(SEQ ID NO.29) from the 5 'end of the quail IFN gene: 5'-GGCGAATTCGGCTAAGTGCGCGTGTTG-3' (SEQ ID NO: 30) "from the 3 'end of the quail IFN gene.

P1501 / 99 X DNA by genomic PCR * QT-35 of 594 bp blunt ends was generated by * PCR and inserted into a single Smal site at the multiple cloning site of plasmid pBluescript II KS (Strategene, La Jolla, CA). The resulting plasmid is: 832-71. Bll. The DNA and the amino acid sequences were analyzed and determined to complete the open reading frame of quail interferon, type 1 (SEQ ID No. 31 and 32), based on the homology sequence and on the conservation of the genetic motifs Significant structural differences between three avian interferon genes type 1, published, for example, of duck, turkey and chicken. The OFN of quail IFN-1 contains 594 nucleotides encoding 198 amino acids, including a translation start codon, ATG and an amber translation stop signal, TAG. The sequence contains 6 cysteine residues that are conserved motifs among other avian IFN genes type 1. The amino acid sequence predicts a hydrophobic signal sequence of 31 amino acids and two putative N-glycosylation sites. The homology of the amino acid sequence (Jotun Hein Method, DNASTAR MegAlign Program) of the quail IFN-1 in comparison with the IFN-1 of chicken, turkey and duck is 82.0%, 76.0% and 49.0%, respectively. The Kyte-Doolittle protein hydrophobicity plots show the structure of quail IFN-1 protein and the characteristics of P1501 / 99MX cargo that are similar to chicken, turkey and duck IFN-1 proteins. A plasmid pSY832 -71. bll containing the quail DNA has been deposited on February 21, 1997 according to the Budapest Treaty on the International Deposit of Microorganisms for the Purpose of Patent Procedure before the Patent Crop Agency of the American Type Culture Collection, 12301 Parkiawn Drive, Rockville, Maryland 20852 USA with accession number ATCC 97892. The expression plasmid 832-71. Bll is useful as a vaccine against avian disease in avian species such as chickens, ducks, quail, turkeys, gallins, guinea fowl and others. Quail IFN-1 improves an immune response against disease-provoking microorganisms when it is delivered in a viral vector alone or in combination with other antigens that cause avaric disease. The expression plasmid 832-71. Bll is useful as a vaccine to improve weight gain in avian species.

EXAMPLE 23C The expression plasmid for the antisense quail interferon: 866-79.2 A tronic expression plasmid bicis, pIRESlneo, (Clontech, Palo Alto, CA) was used as the cloning vector. This cassette of expression P1501 / 99 X contains the immediate early major protein of human cytomegalovirus (CMV) / enhancer, followed by a multiple cloning site (MCS); a synthetic intron; and the internal ribosome entry site (IRES) of the encephalomyocarditis virus, followed by the neomycin phosphotransferase gene with a polyadenylation signal of bovine growth hormone downstream. The antisense for the open DNA reading frame of quail interferon, type 1, was inserted into a unique BamHl site downstream of MCS promoter CMV and the transcription initiation site of pIRESrzeo. The resulting expression plasmid pIRESneo containing the quaternary interferon type 1 was 866-79.2. Cell line stably transformed QT-35 with plasmid 866-79.2 of quaternary interferon type 1. As it does not easily obtain a chicken cell line stably transformed to elaborate cell lines less chicken interferon, a QP-35 cell line transformed with a quail IFN-1 antisense expression plasmid was designed to make new and improved cell substrates for the growth of avian viruses, such as HVT. The stably transformed cell line QT-35 and the vector expressing the antisense cDNA to quaternary interferon type 1 was used.

P1501 / 99MX The QT-35 cells were transfected with a plasmid containing antisense DNA for quail IFN type 1 and a selectable marker gel, neomycin phosphotransferase. Plasmid DNA, 866-79.2, was precipitated in the presence of regulator HEPES and calcium, mixed with growth medium QT-35 and se. incubated with monolayers of QT-35 cells at 89 ° C. After 5 to 6 hours, the cells were challenged with 15% glycerol for 3 minutes, washed with PBS and fed with growth medium. The cells were allowed to form confluent layers and then they were trypsinized and plated on 6 cm dishes at subconfluent concentrations. After the cells had been bound and acclimated to the plastic, the medium was replaced with culture medium containing 400 μg / ml G418. G418-resistant cells were allowed to form colonies in the presence of 400-800μg / ml of G418 for 3 to 3 weeks or at least until nontransfected and non-G418-resistant QT-35 cells had died. Clones resistant to G418 were trypsinized and plated on 10cm culture dishes. The clones were subcultured and expanded as required once a week in a medium containing 800 μg / ml of G418. The QT-35 clones were selected based on their growth stability in the presence of P1501 / 99MX 800μg / ml of G418. The Southern blot analysis of genomic DNA from clones resistant to G418 and from the original QT-35 cells was carried out. A DNA probe of neomycin phosphotransferase was labeled with digoxigenin dUTP (Boeringer Mannheim) which was used to identify the presence of the gene construct stably integrated into the genomic DNA. Cell clones resistant to G418 and positive for the neomycin gene by the southern blot were considered with full expression of the plasmid, 866-79.2 stably integrated into the chromosomal genomic cell DNA. The improved QT-35 cell line transformed with a quail IFN-1 antisense expression plasmid is useful for the production of avian viruses as per example: HVT, chimeric viral vector HVT / MDV, avian pustulation virus, avipox virus and other viruses. The growth of avian viruses at higher titers in the improved QT-35 cell line was due to the inhibition of interferon production in the cell line.

S-SPV-129 S-SPV-129 is a recombinant porcine pustulation virus that contains 2 genes, the quail interferon gene type 1 (qIFN-1) and the lacZ gene, inserted in a single Accl site (within the ORF) P1501 / 99MX 01L) within the subfragment HindIII to BglII of the HindIII M genomic fragment of SPV. The qIFN-1 gene is under the control of the late promoter 2 early promoter 2 (LP2EP2), and the lacZ gene is under the control of the late promoter 1 (LP1). S-SPV-129 was created by the HOMOLOGA RECOMBINATION PROCEDURE TO GENERATE PV *** RECOMBINANT SPV between S-SPV-001 and the homology vector 849069-A1 in the ESK-4 cell line (See WO 96/22363) . The transfection storage material was screened by RECOMBINANT SPV SCREENING EXPRESSING ENZYMATIC MARKERS GENES. The final result of blue plate purification was the recombinant virus designated S-SPV-129. This virus was assessed for ß-galactosidase expression, purity, insert stability by multiple subcultures monitored by blue plate analysis as described in the materials and methods section. After the initial three rounds of purification, all the plates observed were blue indicating that the virus was pure, stable and expressed the marker gene. To verify the biological activity of quail IFN-1, supernatants of ESK-4 cells infected with S-SPV-129 were harvested and tested for ability to inhibit infection with vesicular stomatitis virus (VSV) in CEF cells. .

P1501 / 99MX S-SPV-129 is useful as a vaccine against avian diseases - in avian species, for example, chickens, ducks, quail, turkey, chicken, guinea hen and others. Quail IFN-1 improves an immune response against microorganisms that cause the disease when it is delivered in a viral vector alone or in combination with other antigens that cause avian disease. S-SPV-129 is useful as a vaccine to improve weight gain in avian species. The expression of quail IFN-1 in S-SPV-129 or a eukaryotic expression plasmid for example, pIRESlneo, (Clontech, Palo Alto, CA) (see example 23C above) is useful when it is injected into the germ line chickens or other avian species to produce transgenic chickens or other avian species. Transgenic chickens for qIFN-1 demonstrate improved gain in weight and growth as well as better resistance to disease.

E 24 Test? of transient expression comparing a promoter of novel chicken anemia virus (CAV) with the immediate early promoter of HMCV. Chicken anemia virus (CAV) is a 2300-nucleotide single-stranded DNA virus that contains a P1501 / 99MX main open reading frame (51 kdal capsid) and several smaller ORFs. CAV infects chicken to lymphoid cells and produces an RNA transcript of 2100 nucleotides from a different promoter region (References 67-72). Four different CAV promoter constructs expressing β-galactosidase have been assessed to determine activity in the transient assays. These promoter constructs are plasmids 850-25.18, 850-69.1, 850-80.2 and 883-11. A5, everything? IO? which use different CAV promoters to boost the expression of β-galac tosidase. Several transient assays in CEF and other cell lines have been made with these constructs and plasmid 388-65.2, which uses the HCMV-IE promoter to express β-galactosidase. Plasmid 850-25.18 contains a 854 bp version of the CAV promoter that extends through the first two translational beginnings for ORFs 1 and 2 up to the translational start for ORF 3, the CAV capsid protein gene. This promoter would include a coding sequence functional for the apoptin gene (ORF 2). Plasmid 850-69.1 contains a novel CAV promoter that is similar to 850-25.18, except that a Hind III site within the apoptin reading frame has been filled to destroy the apoptin reading frame, but only near the 3 'end. .

P1501 / 99IIX This creates a 858 base pair version of the CAV promoter. Plasmid 883-11. 5 contains a promoter CAV that starts at the same site upstream as the two previous promoters, but is only 381 bp long, extending towards the transductional start of 0RF1. Plasmid 850-80.2 contains a CAV promoter (SEQ ID NO 23) which is similar to 883-11. A5, but position-3 relative to the transductional start has been altered from nucleotide T to nucleotide A (Nucleotide 377 of SEQ ID NO 23), which is closest to a consensus ribosone entry site, as described for other promoters eukaryotic by Kozak. The results? of the transient transfection assay show that plasmid 850-80.2 of CAV promoter expresses high levels of β-galactosidase comparable with the levels observed with plasmid 388-65.2, which contains the highly active HMCV IE promoter. The CAV promoter plasmid 850-80.2, which has a nucleotide change from T to A at the proposed ribosome entry site expresses higher transient levels of, for example, β-galactosidase than the CAV promoter plasmids, 850-25.18, 850-69.1 and 883-11. 5 (see Figure 6). The results of transient transfection assays were observed in both cells of P1501 / 99MX chicken embryo fibroblast as in QT-35 cells (quail). The CAV promoter in plasmid 850-80.2 is useful as a vaccine to express high levels of viral or bacterial antigens, cytokines, immune modulating proteins and cell surface receptors or cytostatic cycloses making it an excellent promoter for use in recombinant viruses. The CAV promoter in plasmid 850-80.2 was used to make recombinant S-HVT-148 and S-FPV-106.

Use 25 Recombinant HAVT and recombinant FPV expressing foreign DNA from a chicken anemia virus promoter.

S-HVT-148 S-HVT-148 is a recombinant turkey herpesvirus containing the E. coli β-galactosidase gene inserted into the Xhol site in the EcoRl # 9 fragment within the unique long region of HVT. This plasmid contains the β-galactosidase gene from E. coli under the control of a chicken anemia virus (CAV) promoter. The CAV promoter is a 381 PV fragment containing the CAV promoter sequence up to the first CAV ORF, where the 3-position changes from T to A (Nucleotide 377 of SEQ IDE NO 23).

P1501 / 99MX The S-HVT-148 was constructed according to the PROCEDURE TO GENERATE = RECOMBINANT HERPESVIRUS FROM SUBGENOMICAL TRANSLATING FRAGMENTS The following combination of subgenomic clones and enzymes was used: 407-32.2C3 with Notl, 407-32.5G6 with Notl, 172-07, BA2 with BamHl, 415-09 BAl with BamHl, 672-01, A40 with Notl, 672-07, C40 with Notl and the homology vector 867-96, B9 not cut, HVT-148. is a pure virus and expresses ß-galactosidase as tested by ONPG and blue plate assays.S-HVT-148 is useful as a vaccine in poultry against Marek's disease.Another foreign DNA of interest is inserted under control of the CAV promoter to be used as a vaccine in poultry The CAV promoter is useful in HVT, the HVT / MDV chimeric viral vaccine and other herpesviruses and during the infection of foreign DNA from disease causing microorganisms, it is useful as a vaccine in canines, felines, bovines, pigs, equines and human species S-FPV-106 S-FPV-106 is a recombinant avian pustulation virus that contains the E. coli β-galactosidase gene inserted into a unique SnaBI site (converted to a NotI site) in the EcoRI fragment genomic FPV of 2.8 KB. S-FPV-106 contains the E. coli β-galactosidase gene under the P1501 / 99MX control of a novel promoter of chicken anemia virus (CAV). The CAV promoter is a 381 bp fragment containing the CAV promoter sequence up to the first CAV ORF, where the 3-position changes from T to A (Nucleotide 377 of SEQ ID NO 23). S-FPV-106? E created by RECOMBINATION PROCEDURE HOMOLOGA TO GENERATE RECOMBINANT FPV between 883-10. l and the homology vector S-FPV-001. (See WO 94/19015). The transfection storage material was screened by the RECOMBINANT FPV SCREENING METHOD EXPRESSING ENZYMATIC MARKERS GENES. The final result of blue plate purification was the recombinant virus designated S-FPV-106. This virus was analyzed for ß-galactosidase expression, purity and insert stability by multiple subcultures monitored by the blue plate assay, as described in the materials and methods section. After the initial three rounds of purification, all the plates observed were blue indicating that the virus was pure, stable and expressed the marker gene. S-FPV-106 is useful as a vaccine in poultry against avian pustulation disease. Another "interesting" foreign DNA is inserted under the control of the CAV promoter to be used as a vaccine in poultry.The CAV promoter is useful in FPV, porcine pustulation virus *, pustulation virus in raccoons and other poxviruses as a P1501 / 99MX vaccine and with the insertion of foreign DNA from the microorganisms that cause diseases, it is useful as a vaccine in canines, felines, bovines, swine, equines and human species.

Example 26 S-HVY-148 S-HVY-148 is a recombinant chimeric viral vaccine comprising a chimera from the short region of the Marek's disease virus and the long region of the turkey herpesvirus. S-HVT-148 is a recombinant chimeric viral vaccine comprising foreign DNA from the hemagglutinin (HN) genes of Newcastle disease virus (NDV) and fusion (F) inserted into the Xhol site in the EcoRl fragment. # 9 within the unique long region of the chimeric virus. The NDV HN gene is under the control of the PRV gX promoter and the NDV F gene is under the control of the HCMV immediate early promoter. The NDV HN and F genes under the control of their respective promoters are transcribed in opposite directions to those of the HCVT UL52, UL54 and UL55 genes in adjacent HVT genomic DNA. To generate S-HVY-148, the following combination of subgenomic clones and enzymes were used in the PROCEDURE TO GENERATE RECOMBINANT HERPESVIRUS FROM SUBGENOMICAL TRANSFER FRAGMENTS: 407-32.2C3 with Notl, 172-07.BA2 with BamHl, 407-32.5 G6 with Notl, 854-33.6 CON P1501 / 99MX Sfil, and 739-27.16 with Notl. S-HVY-148 was purified by plating and purifying on plate and testing the purity by means of the BLACK PLATE TEST. S-HVY-148 was 100% pure by the BLACK PLATE TEST using anti-NDV-F monoclonal antiserum (# 3-lG5) and anti-NDV-HN monoclonal antiserum (# 15c4). To confirm the expression of the NDV HN and F gene products, the cells were infected with S-HVY-148 and the samples of the infected cell lysates were subjected to SDS-polyacrylamide gel electrophoresis. The gel was transferred and analyzed using the WESTERN TRANSFER PROCEDURE. The NDV HN and F antisera were used to detect the expression of the NDV HN and F protein. The lysate of the infected S-HVY-148 cells exhibited a band in the expected size for native HN and F of NDV. The recombinant chimeric viral vaccine, S-HVY-148, is useful for protection against sensitization with virulent Marek's disease virus and Newcastle disease virus. The recombinant virus S-HVY-148 is useful for the production of recombinant protein, HN and F of NDV, for diagnostic analysis or as a vaccine.

Example 27 S-HVY-154 S-HVY-154 is a chimeric viral vaccine Recombinant P1501 / 99MX comprising a chimera from the short region of the Marek's disease virus and the long region of the turkey herpesvirus. S-HVY-154 is a recombinant chimeric viral vaccine comprising foreign DNA from the fusion (F) gene of Newcastle disease virus (NDV) and the lacZ gene of E. coli inserted into the Xhol site in the fragment EcoRl # 9 within the unique long region of the chimeric virus. The F gene of NDV is under the control of the immediate early promoter HCMV, and the lacZ gene of E. coli is under the control of the PRX gX promoter. The lacZ gene of E. coli and the F gene of NDV under the control of their respective promoters are transcribed in the opposite direction of the UL52, UL54 and UL55 genes of HVT in the adjacent HVT genomic DNA. The S-HVY-154 was derived from S-HVY-145. This was achieved using the homology vector 890-77.10 and the virus S-HVY-145 in the method of TRANSFECTION OF DNA TO GENERATE RECOMBINANT VIRUS, in primary chicken embryo fibroblast cells (CEF). A blue virus obtained from the transfection storage material is purified by successive plate purifications using the CPRG SCREENING FOR RECOMBINANT HERPESVIRUS method. To confirm the expression of the NDV HN and F gene products, cells were infected with S-HVY-154 and samples of the cell lysates were subjected to electrophoresis in SDS-polyacrylamide gel. He P 15 01 / 99IIX gel was transferred and analyzed using the WESTERN TRANSFER PROCEDURE. The antisera for HN and F of NDV were used to detect expression of the HN and F protein of NDV. The lysate of the infected S-HVY-154 cells exhibited a band in the expected size for native NDV HN_ and F. S-HVY-154 is useful as a vaccine against sensitization with virulent Marek's disease virus and Newcastle disease virus. The recombinant virus "S-HVY-154 is useful for the production of recombinant protein, NDV F, for diagnostic analysis or as a vaccine.

E ng 28 S-HVT-149 S-HVT-149 is a recombinant turkey herpesvirus vaccine comprising foreign DNA from the VP2 gene of infectious bursal disease virus (IBDV) inserted into the Xhol site in the EcoRl fragment # 9 within the single long region of HVT. The VP2 gene of IBDV is expressed under the control of a chicken anemia virus (CAV) promoter. The VP2 gene of IBDV under the control of the CAV promoter is transcribed in the opposite direction of the UL52, UL54 and UL 55 genes of HVT in the adjacent HVT genomic DNA. To generate S-HVT-149, the following P1501 / 99MX combination of clones and subgenomic enzymes were used in the PARA PROCEDURE. GENERATE RECOMBINANT HERPESVIRUS FROM SUBGENOMICAL TRANSFER FRAGMENTS: 407-32.2C3 with Notl, 172-07.BA2 with BamHl, 407-32.5G6 with Notl, 672-01. C40 with Notl, 900-87. H8 with Notl, 672-01. A40 with Not I and 415-09. BAl with BamHl. S-HVT-149 was purified by plating and purifying on plate and its purity was tested by the BLACK PLATE TEST. S-HVT-149 was 100% pure by the BLACK PLATE ESSAY using anti-IBDV antisera. To confirm the expression of the VP2 gene product of IBDV, the cells were infected with S-HVT-149 and the samples of the infected cell lysates were subjected to SDS-polyacrylamide gel electrophoresis. The gel was transferred and analyzed using the WESTERN TRANSFER PROCEDURE. The antiserum? for VP2 of IBDV were used to detect the expression of the VP2 protein of IBDV. The lysate from the infected S-HVT-149 cells exhibited a band in the expected size for VP2 of IBDV gene VP2 of 'iBDV gene VP2 of IBDV gene VP2 of IBDV gene VP2 of native IBDV. The recombinant viral vaccine, S-HVT-149, is useful for protection against sensitization with the Marek's disease virus and the infectious bursal disease virus. The recombinant virus S-HVT-149 P15 01/9 9IIX is useful for the production of recombinant protein, VP2 of IBDV, for diagnostic analysis or as an S-HVT-149 illustrates the utility of the CAV promoter for expressing foreign DNA in a recombinant virus.

Example 29 S-HVT-154 S-HVT-149 is a recombinant turkey herpesvirus vaccine comprising foreign DNA from the VP2 gene of infectious bursal disease virus (IBLV) inserted into the Xhol site in the EcoRl fragment # 9 within of the unique long region of HVT. The VP2 gene of IBDV is expressed under the control of a chicken anemia virus (CAV) promoter. The VP2 gene of IBDV under the control of the CAV promoter transcribes in the opposite direction of the UL52, UL54 and UL55 genes of HVT in the adjacent HVT genomic DNA. To generate S-HVT-149, the following combination of subgenomic clones and enzymes were used in the PROCEDURE TO GENERATE RECOMBINANT HERPESVIRUS FROM SUBGENOMIC TRANSLATING FRAGMENTS: 407-32.2C3 with Notl, 172-07. BA2 with BamHl, 407-32.5G6 with Notl, 672-01. C40 with Notl, 900-87. H8 with Notl, 672-01.A40 with Not I and 415-09. BAl with BamHl. S-HVT-149? E purified by plating and plaque purification and? E tested to determine purity by the BLACK PLATE TRIAL. The S-HVT-149 was 100% pure by the BLACK PLATE TEST P1501 / 99MX using anti-IBDV antisera. The recombinant protein, VP2 of IBDV, produced by S-HVT-149 was shown by the WESTERN TRANSFER ANALYSIS as similar in size to the VP2 of native IBDV. S-HVT-154 was purified by plating and plaque purification and was tested for purity by BLACK PLATE ANALYSIS. S-HVT-154 was 100% by BLACK PLATE ANALYSIS using anti-IBDV antisera. The recombinant viral vaccine, S-HVT-154, is useful for protection against sensitization with Marek's disease virus and infectious bursal disease virus. The recombinant virus S-HVT-154 is useful for the production of recombinant protein, VP2 of IBDV for diagnostic analysis or as a vaccine. The recombinant virus S-HVT-154 illustrates the utility of ILTV gl promoter for expressing foreign DNA in a recombinant virus.

Example 30 S-HVT-155 S-HVT-155 e a recombinant turkey herpesvirus vaccine comprising the VP2 gene of infectious bursal disease virus (IBDV) inserted in an Xhol site in the EcoRl fragment # 9 within the region long unique HVT. The VP2 gene of IBDV is expressed under the control of a glycoprotein I (gl) promoter of infectious laryngotracheitis virus (ILTV). The VP2 gene of IBDV under the P1501 / 99MX control of the ILTV gl promoter is transcribed in the same direction as the UL52, UL54 and UL55 genes of HVT in the adjacent HVT genomic DNA. To generate S-HVT-155, the following combination of clones and subgenomic enzymes was used in the PROCEDURE TO GENERATE RECOMBINANT HERPESVIRUS FROM SUBGENOMIC TRANSLATING FRAGMENTS: 407-32.2C3 with Notl, 172-07. BA2 with BamHl, 407-32.5G6 with Notl, 949-19.2 with I-Scel and 415-09. BAl with BamHl. The S-HVT-155 was purified by plating and plaque purification and its purity was tested by the BLACK PLATE TEST. The S-HVT-155 was 100% pure by the BLACK PLATE TEST using anti-IBDV antisera. The recombinant viral vaccine, S-HVT-155, is useful for protection against sensitization with the Marek's disease virus and the infectious bursal disease virus. The recombinant virus S-HVT-155 is useful for the production of recombinant protein, VP2 of IBDV, for the diagnostic test or as a vaccine. The recombinant virus S-HVT-155 illustrates the utility of the ILTV gl promoter for expressing foreign DNA in a recombinant virus.

Example 31 S-HVT-156 S-HVT-156 is a recombinant turkey herpesvirus vaccine comprising foreign DNA P1501 / 99MX from the VP2 gene of the infectious bursal disease virus (IBDV) inserted into the Xhol site in the EcoRl fragment # 9 within the single long region of HVT. The VP2 gene of IBDV is expressed under the control of a glycoprotein D promoter (gD) of infectious laryngotracheitis virus (ILTV). The VP2 gene of IBDV under the control of the gD promoter of ILTV is transcribed in the opposite direction from the UL52 genes, UL54 and UL55 of HVT in the adjacent HVT genomic DNA. To generate S-HVT-156, the following combination of subgenomic clones and enzymes was used in the PROCEDURE TO GENERATE RECOMBINANT HERPESVIRUS FROM SUBGENOMICAL TRANSFER FRAGMENTS: 407-32.2C3 with Notl, 172-07.BA2 with BamHl, 407- 32.5G6 with Notl, 949-24. Say with I-Scel and 415-09. BAl with BamHl. The S-HVT-156? E was purified by plating and plaque purification and its purity was tested by the BLACK PLATE TEST. The S-HVT-156 was 100% pure by the BLACK PLATE ESSAY using antiserum? anti-IBDV. The recombinant viral vaccine, S-HVT-156, e? useful for protection against the susceptibility to Marek's disease virus and the infectious bursal disease virus. The recombinant virus S-HVT-156 is useful for the production of recombinant protein, VP2 of IBDV, for the assay of P1501 / 99 X diagnosis or as a vaccine. The recombinant virus S-HVT-156 illustrates the utility of the gD promoter of ILTV to express foreign DNA in a recombinant virus.

Example 32 S-HVY-159 S-HVY-159 is a recombinant chimeric viral vaccine comprising a chimera from the short region of the Marek's disease virus and the long region of the turkey herpesvirus. S-HVY-159 is a recombinant chimeric viral vaccine comprising foreign DNA from the VP2 gene of infectious bursal disease virus (IBDV) inserted into the Xhol site in the EcoRl # 9 fragment within the single long region of the chimeric virus . The VP2 gene of IBDV is expressed under the control of a glycoprotein I (gl) promoter of infectious laryngotracheitis virus (ILTV). The VP2 gene of IBDV under the control of the ILTV gl promoter is transcribed in the same direction as the gene. UL52, UL54 and UL55 of HVT in the adjacent HVT genomic DNA. To generate S-HVY-159, the following combination of subgenomic clones and enzymes was used in the PROCEDURE TO GENERATE RECOMBINANT HERPESVIRUS FROM SUBGENOMICAL TRANSFER FRAGMENTS: 407-32.2C3 with Notl, 172-07.BA2 with BamHl, 407- 32.5G6 with Notl, 949-19.2 with I-Scel, and 928-47.1 with Notl.

P1501 / 99MX The S-HVY-159 was purified by plating and purifying the plate as well as by testing the purity by means of the BLACK PLATE TEST. The S-HVY-159 was 100% pure by the BLACK PLATE TEST using anti-IBDV antisera. The recombinant viral vaccine, S-HVY-159, is useful for protection against sensitization with the Marek's disease virus and the infectious bursal disease virus. The recombinant virus S-HVY-159 is useful for the production of recombinant protein, VP2 of IBDV, for the diagnostic assay or as a vaccine. The recombinant virus S-HVY-159 illustrates the utility of the ILTV gl promoter for expressing foreign DNA in a recombinant virus.

Example 33 S-HVY-160 The S-HVY-160 e? a recombinant chimeric viral vaccine comprising a chimera from the short region of the Marek's disease virus and the long region of the turkey herpesvirus. S-HVY-160 is a recombinant chimeric viral vaccine comprising foreign DNA from the VP2 gene of infectious bursal disease virus (IBDV) inserted into an Xhol site in the EcoRl # 9 fragment within the single long region of the virus chimerical. The VP2 gene of IBDV is expressed under the control of a glycoprotein I (gl) promoter of infectious laryngotracheitis virus (ILTV). The VP2 gene of IBDV under the P1501 / 99MX promoter control of ILTV? E transcribed in the opposite direction that the UL52, UL54 and UL55 genes of HVT in the genomic DNA of adjacent HVT. To generate S-HVY-160, the following combination of clones and subgenomic enzymes were used in the PROCEDURE TO GENERATE RECOMBINANT HERPESVIRUS FROM SUBGENOMICAL TRANSFER FRAGMENTS: 407-32.2C3 with Notl, 172-07.BA2 with BamHl, 407- 32.5G6 with Notl, 949-24.12 with I-Scel, and 928-47.1 with Notl. The S-HVY-160 was purified by plating and purifying the plate as well as by testing the purity by means of the BLACK PLATE TEST. The S-HVY-160 was 100% pure by the BLACK PLATE TEST using anti-IBDV antisera. The recombinant viral vaccine, S-HVY-160, is useful for protection against sensitization with the Marek's disease virus and the infectious bursal disease virus. The recombinant virus S-HVY-160 is useful for the production of recombinant protein, VP2 of IBDV, for the diagnostic assay or as a vaccine. The recombinant virus S-HVY-160 illustrates the utility of the ILTV gl promoter for expressing foreign DNA in a recombinant virus.

Example 34 S-HVT-150 S-HVT-150 is a recombinant turkey herpesvirus vaccine comprising foreign DNA to P1501 / 99MX from the lacZ gene of E. coli inserted into an Xhol site in the EcoRl fragment # 9 within the single long region of HVT. The lacZ gene of E. coli is expressed under the control of a glycoprotein I (gl) promoter of infectious laryngotracheitis virus (ILTV). The lacZ gene of E. coli under the control of the ILTV gl promoter is transcribed in the opposite direction to the UL52 genes, UL54 and UL55 of HVT in the adjacent HVT genomic DNA. To generate S-HVT-150, the following combination of clones and subgenomic enzymes were used in the PROCEDURE TO GENERATE RECOMBINANT HERPESVIRUS FROM SUBGENOMICAL TRANSFER FRAGMENTS: 407-32.2C3 with Notl, 172-07.BA2 with BamHl, 407-32.5 G6 with Notl, 672-07. C40 with Notl, 928-58. J2 with Notl, 672-01. A40 with Not I and 415-09. BAl with Ba Hl. The S-HVT-150 was purified by plating and purifying the plate and testing the purity by means of the BLACK PLATE TEST. The S-HVT-150 was 100% pure by the BLACK PLATE TEST using antiserum? of anti-β-galactosidase. The recombinant viral vaccine, S-HVT-150 e? useful for protection against? in? ization with the Marek's disease virus. The recombinant virus S-HVT-150 is useful for diagnostic testing or as a vaccine. The recombinant virus S-HVT-150 illustrates the utility of the gl ILTV promoter for P1501 / 99 X express foreign DNA in a recombinant virus.

Example 35 S-HVT-151 S-HVT-151 is a recombinant turkey herpesvirus vaccine comprising foreign DNA from the E. coli lacZ gene inserted into an Xhol site in the EcoRl # 9 fragment within the region long unique HVT. The lacZ gene of E. coli is expressed under the control of a glycoprotein D (gD) promoter of infectious laryngotracheitis virus (ILTV). The lacZ gene of E. coli under the control of the gD promoter of ILTV is transcribed in the opposite direction to the UL52, UL54 and UL55 genes of HVT in the genomic DNA of adjacent HVT. To generate S-HVT-151, the following combination of clones and subgenomic enzymes were used in the PROCEDURE TO GENERATE RECOMBINANT HERPESVIRUS FROM SUBGENOMICAL TRANSFER FRAGMENTS: 407-32.2C3 with Notl, 172-07.BA2 with BamHl, 407-32.5 G6 with Notl, 672-07. C40 with Notl, 928-58. K7 with Notl, 672-01. A40 with Not I and 415-09. BAl with BamHl. S-HVT-151 was purified by plating and plaque purification and tested for purity by the BLACK PLATE TEST. The S-HVT-151 was 100% pure by the BLACK PLATE TEST using anti-β-galactosidase antisera. The recombinant viral vaccine, S-HVT-151, e? P1501 / 99 X useful for protection against sensitization with the Marek's disease virus. The recombinant virus S-HVT-151 is useful for diagnostic testing or as a vaccine. The recombinant virus S-t HVT-151 illustrates the utility of the gD promoter of ILTV to express foreign DNA in a recombinant virus.

P1501 / 99MX EFFECTS 1. Buckmaster et al., J. Gen. Virol. 69: 2033, 1988. 2. FA. Ferrari et al., Journal of Bacteriology t 161, 556-562, 1985. 3. U. Gubler and B.J Hoffman, Gene 25, 263-269. 4. D. Hanahan, Molecular Biology 166, 557-580, 1983. 5. P.J. Hudson et al., Nucleic Acid Research 14, 5001-5012, 1986. 6. T. Igarashi et al., International Herpesvirus Workshop, Abstract No. 17, Ann Arbor, Michigan, August 1985. 7. T. Ihara et al. ., Virus Genes 3, 127-140, 1989. 8. MA Innis et al., PCR Protocols A Guide to Methods and Applications, 84-91, Academic Press, Inc., San Diego, 1990. 9. RJ Isfort et al., 9th International Herpesvirus Workshop, Abstract No. 146, Seattle, Washington, August 1984. 10. M.N. Jagadish et al., J. of Virol. 62, 1084-1087, 1988. 11. Kawai and Nishizawa Mol. and Cell Bio. 4, 1172-1174, 1984. 12. B. Lomniczi et al., Journal of Virology 49, 970-979 1984. 13. Maniatis et al., Molecular Cloning, Cold Spring Harbor Laboratory, New York, 1982.

P1S01 / 99MX 14. D.J. McGeoch et al., Journal of Molecular Biology 181, 1-13, 1985. 15. S.L. McKnight and R. Kingsbury, Science 217, 316-324, 1982. 16. L.J.N. ROS? et al., Journal of General Virology 70, 1789-1804, 1989. 17. L.J.N. Ross et al., Journal of General Virology 72, 949-954, 1991. 18. J. Sambrook et al., Molecular Cloning A Laboratory Manual Second Edition, Cold Spring Harbor Press, 1989. 19. M. Zijil et al., Journal of Virology 62, 2191-2195, 1988. 20. Maniatis et al. Intervirology 16, 201-217, 1981. 21. S.L. Mansour et al., Proc. Nati Acad. Sci. USA 82, 1359-1363, 1985. 22. C. Thummel et al., Cell 33, 455-464, 1983. 23. D. Scolnick, Cell 24, 135-143, 1981. ' 24. C. Thummel et al., Cell 23, 825-836, 1981. 25Y. Haj-Ahmed and F.L. Graham, J. of Virology 57, 267-274, 1986. 26. M. Mackett et al., Proc. Nati Acad. Sci. USA 79, 7415-7419, 1982. 27. D. Panicali and E. Paoletti, Proc. Nati Acad. Sci. USA 79, 4927-4931, 1982. 28. E. Paoletti et al., Proc. Nati Acad. Sci. USA 81, 193-197,1984 29. G.L. Smith et al., Nature 302, 490-495, 1983.

P1501 / 99MX 30. J.H. Gillespie et al., J. Clin. Microbiology 23, 283-288, 1986. 31. D. Panicali et al., Proc. Nati Acad. Sci. USA 80, 5364-5368, 1983. 32. G.L. Smith et al., Proc. Nati Acad. Sci, USA 80, 7155-7159, 1983. 33. G.L. Smith et al., Science 224, 397-399, 1984. 34. M. Mackett et al., Science 227, 433-435, 1985.

. IS. Moccarski et al., Cell 22, 243-255, 1980. 40 L.E. Post and B. Roizman, Cell 25, 227-232, 1981. 41. K.L. Poffenberger et al., Proc. Nati Acad. Sci. USA 80, 2690-2694, 1981. 42. M.G. Gibson and P.G. Spear, Journal of Virology 48, 396-404, 1983. 43. G.T. -Y. Lee et al., Proc. Nati Acad. Sci. USA 79, 6612-6616, 1982. 44. M.-F. Shih et al., Proc. Nati Acad. Sci. USA 81, 5867-5870, 1984. 45. R. Desrosiers et al., Ninth Annual Herpesvirus Meeting, Seattle, Abstract # 280, 1984. 46. M. Arsenakis and B. Roizman, in "The High Technology Route to Virus Vaccines" , American Society for Microbiology, Washington DC, 1985 (Proceedings of the First Annual Southwest Foundation for Riamedical Research International Symposium, Houston, Texas, 8-10 November 1984). 47. YOU. Post et al., Tenth International P150 ** / 99MX Herpesviru? Workshop, Ann Arbor, August 1985. 48. S.B. Mohanty and S.K. Dutta, Veterinary Virology, Lea and Febiger, pubs. , Philadelphia, 1981. í 49. A.M. Griffin, Journal of General Virology 72, 393-398, 1991. 50. D.R. Thomsen et al., Gene 16, 207-217, 1981. 51. Carpenter, D.E. and Misra, V. Journal of General Virology 72 3077-3084 (1991). 52. Kibenge, F.S., Jackwood, D.J., Mercado, C.C., Journal of General Virology 71 569-577 (1990). 53. Fulcuchi et al., J. Virologu 51 102-109, 1984. 54. Fukuchi et al., J. Virology 53 994-997, 1985. 55. Ross, N., et al., Virus Genes 7 33-51, 1993. 56. Maotani, KA, et al., J. Virology 58: 657-659, 1986. 57. Ross, LJN, et al., J General Virology 64: 2785-2790, 1983. 58. A. Leutz, et al., EMBO Journal 8: 175-182 (1989). 59. M.J. Sekellick, et al., Journal of Inferieron Research 14: 71-79 (1994). 60. G.L. Smith, Journal of General Virology 74, 1725-1740 (1993). 61. B. Schumacher, et al., Virology 203, 144-148 (1994). 62. Dxgby and Lowenthal, Journal of Interferon and Cytokine Research 15: 933-938 (1995).

P1501 / 99MX 63. Lowenthal, et al., Journal of Interferon and Cytokine Research 15: 939-945 (1995). 64. Schultz, et al., Virology 212: 641-649 (1995). 65. Sekellick, et al., WO 95/11302; Univ. Conn. 66. Lowenthal, et al., WO 96/27666; CSIRO. 67. Noteborn, M.H.M., et al J. Virol. 65 (6), 3131-31 (1991). 68. Noteborn, et al., U.S. 5,491,073; Aesculaap, N.V. 69. Noteborn, et al., WO 95/03414; Aesculaap, N.V. 70. Schat, et al., WO 96/01116; Cornell Research Fdn. 71. Schreir, et al., EP 533 294 Al; Akzo, N.V. 72. Sondermeijer, et al., EP 483 911 A2; Akzo, N.V.

P1501 / 99MX SEQUENCE LISTS (1) GENERAL INFORMATION: (i) APPLICANT: Cochran, Mark D Wild, Martha A Barbara J. Winslow f (ii) TITLE OF THE INVENTION: Recombinant Chimeric Viruses and Uses of Them (iii) NUMBER OF SEQUENCES: 32 (iv) ADDRESS FOR CORRESPONDENCE: (A) RECIPIENT: John P. White (B) STREET: 1185 Avenue of the Americas (C) CITY: New York (D) STATE: New York (E) ) COUNTRY: USA (F) POSTAL CODE: 10036 (v) FORM FOR COMPUTER READING: (A) TYPE OF MEDIUM: flexible disk (B) COMPUTER: IBM PC compatible (C) OPERATING SYSTEM: PC-DOS / MS- DOS (D) SOFTWARE: Patentln Reeléase # 1.0, Version # 1.25 (vi) DATA OF THIS APPLICATION (A) NUMBER OF APPLICATION: (B) DATE OF PRESENTATION: (C) CLASSIFICATION: (viii) INFORMATION OF THE APPORTER / AGENT ( A) NAME: White, John P (B) REGISTRATION NUMBER: 28,678 (ix) INFORMATION FOR TELECOMMUNICATION (A) TELEPHONE: (212) 278-0400 (B) TELEFAX: (212) 391-0526 (C) TELEX: 422523 P1501 / 99MX (2) INFORMATION FOR SEQ ID NO: 1: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 5426 base pairs (B) TYPE: nucleic acid (C) TYPE OF CHAIN: double (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: DNA (genomic) (iii) HYPOTHETICAL: NO (iv) ANTI-SENSE: NO (ix) CHARACTERISTICS: (A) NAME / KEY: CDS (B) LOCATION: 73..1182 (D) OTHER INFORMATION: / product = 'HVT UL42 (ix) CHARACTERISTICS: (A) NAME / KEY: CDS (B) LOCATION: 1306..2574 (D) OTHER INFORMATION: / product ***** "HVT UL43" (ix) FEATURES: (A) NAME / KEY: CDS (B) LOCATION: 2790..4259 (D) OTHER INFORMATION: / product = "HVT gA" (ix) CHARACTERISTICS: (A) NAME / KEY: CDS (B) ) LOCATION: 4701..5339 (D) OTHER INFORMATION: / product = "HVT UL45" (xi) DESCRIPTION OF THE SEQUENCE: SÉQ ID NO: 1: P1501 / 99MX GGATCCGAGC TTCTACTATA CAACGCGGAC GATAATTTTG TCCACCCCAT CGGTGTTCGA 60 GAAAGGGTTT TT ATG ATG GCA GGA ATA ACT GTC GCA TGT GAC CAC ACT 108 Het Met Wing Gly lie Thr Val Wing Cys Asp Hi = Thr 1 5 10 156 Wing Gly Glu Wing His Thr Pro Glu Asp Met Gln Ly = Lys Trp Arg He 15 20 25 ATA TTG GCA GGG GAA AAA TTC ATG ACT ATA TCG GCA TCG TTG AA? TCG 204? Le Leu Ala Gly Glu Lys Phe Het Thr He Ser Wing Ser Leu Lys Ser 30 35 40 ATC GTC AGT TGT AAA AAC CCC CTT CTC ACG TTT GGC GCA GAT GGG 252 lie Val Ser Cys Val Lys Asn Pro Leu Leu Thr Phe Gly Wing Asp Gly 45 50 55 60 CTC ATT GTA CA GGT ACT GTC TGC GGA CAG CGC ATT TTT GTT CCA ATC 300 Leu He Val Gln Gly Thr Val Cys Gly Gln Arg He Phe Val Pro He 65 70 * 75 GAC CGT GAT TCC TTC AGC GAA TAT GAA TGG CAT GGG CCA ACT GCG ATG 348 Asp Arg Aap Ser Phe Ser Glu Tyr Glu Trp His Gly Pro Thr Wing Met 80 85 90 TTT CT? GCA TTA ACT G? TCC AGA CGC ACT CTT TTA GAT GCA TTC? AA 396 Phe Leu Wing Leu Thr A = p Ser Arq Arg Thr Leu Leu Asp Wing Phe Lys 95 '100 105 TGT G? A AAG AGA AGG GCA ATT GAC GTC TCC TTT ACC TTC GCG GGA GAG 444 Cya Glu Lys Arg Arg Ala? Le Aap Val Ser Phe Thr Phe Wing Gly Glu 110 115 120 CCT CCA TGT AGG CAT TTA ATC CAA GCC GTC ACA TAC ATG ACC G? C GGT 492 Pro Pro Cys Arg His Leu He Gln Wing Val Thr Tyr Met Thr Asp Gly 125 130 - 135 140 GGT TCA GTA TCG AAT ACA ATC ATT AAA TAT GAG CTC TGG AAT GCG TCT 540 Gly Ser Val Ser Asn Thr He He Lys Tyr Glu Leu Trp Asn Wing Ser 145 150 155 ACA ATT TTC CCC CAA AAA ACT CCC GAT GTT ACC TTT TCT CTA AAC AAA 5BB Thr He Phe Pro Gln Lys Thr Pro Asp Val Thr Phe Ser Leu Asn Lys 160 165 no CAÁ CAÁ TTG AAC AAA ATA TTG GCC GTC GCT TCA AAA CTG CAA C? C GAA 636 Gln Gln Leu Asn Lys He Leu Wing Val Wing Ser Lys Leu Gln His Glu 175 180 185 GAA CTT GTA TTC TCT TTA AAA CCT GAA GGA GGG TTC TAC GTA GGA ACG 684 Glu Leu Val Phe Ser Leu Lys Pro Glu Gly Gly Phe Tyr Val Gly Thr 190 195 200 GTT GTT ACT GTT ATA AGT TTC GAA GTA GAT GGG ACT GCC ATG ACT CAI} 73: Val Cys Thr Val He Ser Phe Glu Val Asp Gly Thr Ala Met Thr Gl? 205 210 215 220 TAT CCT TAC AAC CCT CCA ACC TCG GCT ACC CTA GCT CTC GT? GT? GC? 700 Tyr Pro Tyr Asn Pro Pro Thr Be Wing Thr Leu Wing Leu Val Val Wing 225 230 235 TGC AGA AAG AAG AAG GCG AAT AAA AAC ACT ATT TTA ACG GCC T? T GGA 828 Cys Arg Lys Lys Lys Wing Asn Lys Asn Thr He Leu Thr Wing Tyr Gly 240 245 250 AGT GGT AAA CCC TTT TGT GTT GCA TTG GAA GAT ACT AGT GCA TTT AGA 876 Ser Gly Lys Pro Phe Cys Val Wing Leu Glu Asp Thr Ser Wing Phe Arg 255 260 265 AAT ATC GTC AAT AAA ATC AAG GCG GGT ACG TCG GGA GTT GAT CTG GGG 924 Asn He Val Asn Lys He Lys Wing Gly Thr Ser Gly Val Asp Leu Gly 270 275 280 TTT TAT ACA ACT TGC GAT CCG CCG ATG CTA TGT ATT CGC CCA CAC GCA 972 Phe Tyr Thr Thr Cys Asp Pro Pro Met Leu Cys He Arg Pro His Wing 285 290 295 300 TTT GGA AGT CCT ACC GCA TTC CTG TTT TGT AAC ACA GAC TGT ATG ACA 1020 Phe Gly Ser Pro Thr Wing Phe Leu Phe Cys Asn Thr Asp Cys Met Thr 305 310 315 ATA TAT GAA CTG GAA GAA GTA AGC GCC GTT GAT GGT GCA ATC CGA GCA 1068 He Tyr Glu Leu Glu Glu Val Ser Wing Val Asp Gly Wing He Arg Wing 320 325 330 AAA CGC ATC AAC GAA TAT TTC CCA ACA GTA TC G CAG GCT ACT TCC AAG 1116 Lys Arg He Asn Glu Tyr Phe Pro Thr Val Ser Gln Wing Thr Ser Lys 335 340 345 AAG AGA AAA CAG TCG CCG CCC CCT ATC GAA AGA GAA AGG AAA ACC ACC 1164 Lys Arg Lys Gln Ser Pro Pro Pro He Glu Arg Glu Arg Lys Thr Thr 350 355 360 AGA GCG GAT ACC CAA TAAAATGCCA GACAAACCCG GCATCCTGGT TAGAGGGCAG 1219 Arg Wing Asp Thr Gln 365 370 GTGGGCTGGG CCAACCTTCA CGGGCGTCCG ACAGATCGGT GACACTCATA CGTTA? CTAA 1279 ACGCCGGCAG CTTTGCAGAA GAAAAT ATG CCT TCC GGA GCC AGC TCG AGT CCT 1332 Met Pro Ser Gly Wing Ser Ser Pro 1 5 CCA CCA GCT TAT ACA TCT GCA GCT CCG CTT GAG ACT TAT AAC AGC TGG 1380 Pro Pro Wing Tyr Thr Wing Wing Pro Leu Glu Thr Tyr Asn Ser Trp 10 15 20 25 CTA AGT GCC TTT TCA TGC GCA TAT CCC CA TGC ACT GCG GGA AGA GGA 142E P15Q1 / 99MX Leu Ser Wing Phe Ser Cys Wing Tyr Pro Gln Cvs Thr Wing Gly Arg Gly 30 35 40 CAT CGA CAA AAT GGC AAG AAG TGT ATA CGG TGT ATA GTG ATC AGT GTA 1476 His Arg Gln A = n Gly Lys Lys Cys He Arg cys He Val He Ser Val 45 50 55 TGT TCC TTA GTG TGC ATC GCT GCA CAT TTA GCT GTT ACC GTG TCG GGA 1524 Cys Sor Leu Val Cys He Wing Wing His Leu Wing Val Thr Val Ser Gly 60 65 70 j GTG GCA TTA ATT CCG CTT ATC GAT CAA AAC AGA GCT TAC GGA AAC TGT 15 * 72 Val Ala Leu He Pro Leu He Asp Gln Asn Arg Ala Tyr Gly Asn Cys 75 80 85 ACG GTA TGT GTA ATT GT GGA TTC ATC GCT ACG TTT GCT GCA CGA CTT 1620 Thr val Cys Val He Wing Gly Phe He Wing Thr Phe Wing Wing Arg Leu 90 SS 100 105 ACG ATA AGA CTT TCG GAA ACG CTT ATG CTA GTG GGC AAG CCG GCG CAG 1668 Thr He Arg Leu Ser Glu Thr Leu Met Leu Val Gly Lys Pro Wing Gln 110 115 120 TTT ATA TTT GCT ATA ATC GCT TCC GTT GCG GAA ACA CTG ATC AAT AAC 1716 Phe He Phe Wing He He Wing Ser Val Wing Glu Thr Leu He Asn Asn 125 130 135 GAG GCG CTT GCC ATC AGT AAT ACT ACT TAC AAA ACT GCA TTG CGA ATA 1764 Glu Ala Leu Ala He Ser Asn Thr Thr Tyr Lys Thr Ala Leu Arg He 140 145 150 ATC G? A GTA AC? TCT TTG GCG TGT TTT GTT ATG CTC GGG GCA ATA ATT 1812 He Glu Val Thr Ser Leu Ala Cys Ph? Val Met Leu Gly Wing Xle He 155 160 165 ACA TCC CAC AAC TAT GTC TGC ATT TCA ACG GCA GGG GAC TTG ACT TGG 1860 Thr Ser His Asn Tyr Val Cys He Ser Thr Wing Gly Asp Leu Thr Trp 170 175 180 185 AAG GGC GGG? TT TTT CAT GCT TAC CAC GGA ACA TTA CTC GGT ATA ACA 1908 Lys Gly Gly He Phe His Ala Tyr His Gly Thr Leu Leu Gly He Thr 190 195 200 ATA CCA AAC ATA CAC CCA ATC CCT CTC GCG GGG TTT CTT GCA GTC TAT 1956 He Pro Asn He His Pro He Pro Leu Wing Gly Phe Leu Wing Val Tyr 205 210 215 ACA ATA TTG GCT ATA AAT ATC GCT AGA GAT GCA AGC GCT ACA TTA TTA 2004 Thr He Leu Wing He Asn He Wing Arg Asp Wing Ser Wing Thr Leu Leu 220 225 230 TCC ACT TGC TAT TAT CGC AAT TGC CGC GAG AGG ACT ATA CTT CGC CCT 2052 Ser Thr Cys Tyr Tyr Arg Asn Cys Arg Glu Arg Thr He Leu Arg Pro - 235 240 245 TCT CGT CTC GG? CAT GGT TAC ACA ATC CCT TCT CCC GGT GCC GAT ATG 2100 Ser? Rg Leu Gly His Gly Tyr Thr He Pro Ser Pro Gly Ala Asp Met. 250 255 260 265 CTT T? T G ?? G ?? G? C GTA TAT AGT TTT GAC GCA GCT AAA GGC C? T T? T 214B Leu Tyr Glu Glu? Sp Val Tyr Ser Phe? Sp Ala? The Lys Gly Hrs Tyr 270 275 280 99MX TCG TCA ATA TTT CTA TGT TAT GCC ATG GGG CTT ACA ACA GCC CTG ATT 2196 Ser Ser've Phe Leu Cys Tyr Ala Met Gly Leu Thr Thr Pro Leu He 285290295 ATT GCG CTC CAT AAA TAT ATG GCG GGC ATT AAA AAT TCG TCA GAT TGG 2244 He Ala Leu His Lys Tyr Met Ala Gly He Lys Asn Ser Ser Asp Trp 300 305 310 ACT GCT ACA TTA CAA GGC ATG TAC GGG CTT GTC TTG GGA TCG CTA TCG 2292 Thr Ala Thr Leu Gln Gly Met Tyr Gly Leu Val Leu Gly Ser Leu Ser 315 320 325 TCA CTA TGT ATT CCA TCC AGC AAC AAC GAT GCC CTA ATT CGT CCC ATT 2340 Ser Leu Cys He Pro Ser Ser Asn Asn Asp Ala Leu He Arg Pro He 33"335 340 345 CA ATT TTG ATA TTG ATA ATC GGT GCA CTG GCC ATT GCA TTG GCT GGA 2388 Gln He Leu He Leu He He Gly Ala Leu Ala He Ala Leu Ala Gly 350 355 360 TGT GGT CAA ATT ATA GGG CCT ACA TTA TTT GCC GCG AGT GCT GCT GCG 2436 Cys Gly G n He He Gly Pro Thr Leu Phe Wing Wing Ser Wing Wing 365 370 375 ATG TCA TGT TTT ACA TGT ATC AAT ATT CGC GCT ACT AAT AAG GGT GTC 2484 Met Ser Cys Phe Thr Cys He Asn He Arg Ala Thr Asn Lys Gly Val 3B0 385 390 AAC AAA TTG GCA GCA GCC AGT GTC GTG AAA TCT GTA CTG GGC TTC ATT 2532 Asn Lys Leu Ala Ala Ala Ser Val Val Lys Ser Val Leu Gly Phe He 395 400 405 ATT TCC GGG ATG CTT ACT TGC GTG CTA TTA CCA CTA TCG TGATAGATCG 2581 He Be Gly Met Leu Thr Cys Val Leu Leu Pro Leu Ser 410 415 420 TCGGTCTGCG CATCGCCCAT GCTGGCGGAA CGCTCTTTCG AACCGTGA? TA ??? CTTTGT 26 1 ATCTACTAAA CAATAACTTT GTGTTTTATT GAGCGGTCGA AAACAATGAG GAGCTGCAAT 2701 TTAAAGCTAA CCGCATACGC CGGGCGGGTA AAGACCATTT TATACCATAT TACGCATCTA 2761 TCGAAACTTG TTCGAGAACC GCAAGTAT ATG GTT TCC AAC ATG CGC * GTT CTA 2813 Met Val Ser Asn Met Arg Val Leu 1 May CGC GTA CTG CGC CTG ACG GGA TGG GTG GGC ATA TTT CTA GTT CTG TCT 2861 Arg Val Leu Arg Leu Thr Gly Trp Val Gly He Phe Leu Val Leu Ser 10 15 20 TTA CAG CAA ACC TCT TGT GCC GGA TTG CCC CAT AAC GTC GAT ACC CAT 2909 Leu Gln Gln Thr Ser Cys Wing Gly Leu Pro His Asn Val Asp Thr His 30 35 40 CAT ATC CTA ACT TTC AAC CCT TCT CCC ATT TCG GCC GAT GGC GTT CCT 2957 His He Leu Thr Phe Asn Pro Ser Pro He Ser Wing Asp Gly Val Pro 45 50 55 TTG TCA GAG GTG CCC AAT TCG CCT ACG ACC GAA TTA TCT ACA ACT GTC 3005 Leu Ser Glu Val Pro Asn Ser Pro Thr Thr Glu Leu Ser Thr Thr Val 60 • 65 70 11X GCC ACC AAG ACA GCT GTA CCG ACG ACT GAA AGC ACT AGT TCC TCC GAA 3053 Wing Thr Lys Thr Wing Val pro Thr Thr Glu Ser Thr Ser Ser Glu 75 80 85 GCG CAC CGC AAC TCT TCT CAC AAA ATA CCT GAT ATA ATC TGC GAC CGA 3101 Wing His Arg Asn Ser His Lys He Pro Asp He lie Cys Asp Arg 90 95 100 GAA GAA GTA TTC GTC CTT AAC AAT ACA GGA AGA ATT TTG TGT GAC 3149 Glu Glu Val Phe Val Phe Leu Asn Asn Thr Gly Arg He Leu Cy = A $ p 105 110 115 mess CTT ATA GTC GAC CCC CCT TCA GAC GAT GAA TGG TCC A? C TTC GCT CTT 3197 Leu He Val Asp Pro Pro Ser Asp Asp Glu Trp Ser A3t? Phe Ala Leu 125 - 130 135 GAC GTC ACTC TTC AAT CCA ATC GAA TAC CAC GCC AAC GAA AAG AAT GTA 3245 Asp Val Thr Phe Asn Pro He Glu Tyr Hia Wing Asn Glu Lys Asn Val 140 145 150 GAG GTT GCC CGA GTG GCC GGT CTA TAC GGA GTA CCG GGG TCG GAT TAT 3293 Glu Val Wing Arg Val Wing Gly Leu Tyr Gly Val Pro Gly Ser Asp Tyr 155 160 165 GCA TAC CCT AGG AAA TCG GAA TTA ATA TCC TCC ATT CGA CGG GAT CCC 3341 Wing Tyr Pro Arg Lys Ser Glu Leu Be Ser Be Arg Arg Arp Asp Pro 170 175 * * 1B0 CAG GGT TCT TTC TGG ACT AGT CCT ACA CCC CGT GGA AAT AAA TAT TTC 3389 Gln Gly Ser Phe Trp Thr Ser Pro Thr Pro Arg Gly Asn Lys Tyr Phe 185 190 195 200 ATA TGG ATT AAT AAA ACA ATG C? C ACC ATG GGC GTG GAA GTT AGA AAT 3437 He Trp He Asn Lys Thr Met Hrs Thr Met Gly Val Glu Val Arg Asn 205 210 215 GTC GAC T? C AAA GAC AAC GGC TAC TTT CA GTG ATA CTG CGT GAT AGA 3485 Val Asp Tyr Lys Asp Asn Gly Tyr Phe Gln Val He Leu Arg A = p Arg 220 225 230 TTT AAT CGC CCA TTG GTA GAA AAA CAT ATT TAC ATG CGT GTG TGC CA 3533 Phe Asn Arg Pro Leu Val Glu Lys His He Tyr Met Arg Val Cys Gln 235 240 245 CGA CCC GCA TCC GTG GAT GTA TTG GCC CCT CCA GTT CTC AGC GGA GAA 3581 Arg Pro Ala Ser Val Asp Val Leu Pro Pro Val Leu Ser Gly Glu 250 255 260 AAC TAC AAA GCA TCT TGC ATC GTT AGA CAT TTT TAT CCC CCG GGA TCT 3629 Asn Tyr Lys Wing Ser Cys He Val Arg His Phe Tyr Pro Pro Gly Ser 265 270 275 280 GTC TAC GTA TCT TGG AGA CGT AAC GOA AAC ATT GCC ACA CCC CGC AAG 3677 Val Tyr Val Ser Trp Arg Arg Asn Gly? Sn He Wing Thr Pro Arg Lys 285 290 295 G? C CGT G? C GGG AGT TTT TGG TGG TTC GAA TCT GGC CGC GGG GCC ACA 3725 Asp Arg Asp Gly Ser Phe Trp Trp Phe Glu Ser Gly Arg Gly Wing Thr 300 305 310 CTA GTA TCC ACA ATA ACC CTC GGA AAC TCT GGA CTC GAA TCT CCT CCA 3773 Leu Val Ser Thr He Thr Leu Gly Asn Ser Gly Leu Glu Ser Pro Pro 315 320 325 X AAG GTT TCC TGC TTG GTA GCG TGG AGG CAA GGC GAT ATG ATA AGC ACA 3821 Lys Val Ser Cys Leu Val Wing Trp Arg Gln Gly Asp Met He Ser Thr 330 335 340 TCG AAT GCT ACA GCT GTA CCG ACG GTA TAT TAT CAC CCC CGT ATC TCT 3869 Ser Asn Ala Thr Ala Val Pro Thr Val Tyr Tyr His Pro Arg He Ser 345 350 355 360 CTG GCA TTT AAA GAT GGG TAT GCA ATA TGT ACT ATA GAA TGT GTT fc 3917 Leu Wing Phe Lys Asp Gly Tyr Wing He Cys Thr He Glu Cys Val Pro 365 370 375 TCT GGG ATT ACT GTG AGG TGG TTA GTT CAT GAT GAA CCC CAG CCT AAC 3965 Ser Gly He Thr Val Arg Trp Leu Val His Asp Glu Pro Gln Pro Asn 380 385 390 ACA ACT TAT GAT ACT G "G GTT ACA GGT CTC TGC AGG ACC ATC GAT CGT 4013 Thr Thr Tyr Asp Thr i il Val Thr Gly Leu Cys Arg Thr He Asp Arg 395 400 405 TAT AGA AAT CTC GCC AGT CGG ATT CCA GTC CAG GAC AAC TGG GCG AAA 4061 Tyr Arg Asn Leu Wing Ser Arg He Pro Val Gln Asp Asn Trp Wing Lys 410 415 * 420 ACG AAG TAT ACG TGC AGA CTA ATT GGA TAT CCG TTC GAC GTG GAT AGA 4109 Thr Lys Tyr Thr Cys Arg Leu He Gly Tyr Pro Phe Asp Val Asp Arg 425 430 435 440 TTT CAA AAT TCC TAG TAT TAT TAT GAT GCA ACG CCG TCG GCA AGA GGA ATG 157 Phe Gln Asn Ser Glu Tyr Tyr Asp Ala Thr Pro Ser Wing Arg Gly Met 445 450 455 CCG ATG ATT GTA ACA ATT ACG GCC GTT CTA GGA CTG GCC TTG TTT TTA 4205 Pro Met He Val Thr He Thr Wing Val Leu Gly Leu Wing Leu Phe Leu 460 465 470 GGT ATT GGT ATC ATT ATC ACA GCC CTA TGC TTT TAC CTA CCG GGG CGG 4253 Gly He Gly He He Thr Wing Leu Cys Phe Tyr Leu Pro Gly Arg 475 480 485 AAT TAAGATTAAC CATCGTATGT GATATAAAAA TTATTAAGTG TTATAACCGA 4306 Asn 490 TCGCATTCTT CTGTTTCGAT TCACAATAAA TAAAATGGTA TTGTAATCAG CACCATCGCA 4366 TTGTTTCGTA GATGACTCAT GTTCAGTCCG CGTGATGTCA AAAATACGTA TTTTTGGTAT 4426 CACGCAGCGG CCAAAATGCC CATTATGTTA TTTTTACTCC AAACGCGGTA TTTAAAACAT 4 486 CGGGACGTAC ATCATGTGGC GCACGTTAAT CGTATACGGT GCCGCTACAT TAAAA? TCGC 454 6 AAGTCTCCGA ATATCAAGCT CACGGCCAAA ACGTCGGTAA TAATCTTACG CATCGAATGT 4606 GAT? CGGATA CCGTACAATC GCTGAGTAGA TTTCCTATAT AGTTACTCAG TAGTGATACA 4 666 CAATCACAAA ATCGCTGGGG TATATCATAT AAGA ATG ATG TCG CCC ACC CCT 47 18 Met Met Ser Pro Thr Pro 1 5 GAA GAT GAT CGC GAT CTC GTT GTG GTT CGT GGA CGT CTC CGA ATG ATG 4766 Glu Asp Asp Arg Asp Leu Val Val Val Arg Gly Arg Leu Arg Met Met MX 10 15 20 GAT AGC GGC ACG GAA ACA GAT AGA GAG CAGE CGA CAT CCA CGT ACG ACT 4814 Asp Ser Gly Thr Glu Thr Asp Arg Glu Gln Arg His Pro Ar? Thr Thr 25 30 35 TGG CGA TCG ATC TGT TGT GGG TGT ACG ATA GGA ATG GTA TTT ACC ATA • 4862 Trp Arg Ser He Cys Cys Gly Cys Thr Ha Gly Met Val Phe Thr ls 40 A m 50 TTC GTT CTC GTA GCG GCA GTA TTG TTG GGA TCA CTA TTC ACT GTT TCA 4910 Pho Val Leu Val Ala Ala Val Leu Leu Gly Ser Leu Phe Thr Val Ser 55 60 65 70 TAC ATG GCC ATG GAA TCG GGA ACA TGT CCC GAT GAA TGG ATT GGT TTG 4958 Tyr Me Wing Met Glu Ser Gly Thr Cys Pro Asp Glu Trp He Gly Leu 75 80 85 GGT TAT AGT TGC ATG GC GTG GCC GGG AAA AAT GCA ACT GAT CTT GAG 5006 Gly Tyr Ser Cys Met Arg Val Wing Gly Lys Asn Wing Thr A = p L'eu Glu 90 95 100 GCG TTG GAT ACA TGT GCT CGG CAT AAC AGC AAA CTT ATT GAC TTC GCA 5054 Wing Leu Asp Thr Cys Wing Arg His Asn Ser Lys Leu He Asp Phe Wing 105 110 115 AAC GCC AAA GTT CTG GTT GAA GCT ATC GCC CCA TTC GGT GTG CCA AAT 5102 Abn Wing Lys Val Leu Val Glu Wing Wing Pro Pho Gly Val Pro Asn 120 125 130 GCA GCA TAT GGG GAA GTC TTC CGG TTA AGG GAC AGC AAA ACC ACG TGT 5150 Wing Wing Tyr Gly Glu Val Phe Arg Leu Arg Asp Ser Lys Thr Thr Cys 135 140 145 150 ATA CG.ñ CCT ACC ATG GGA GGA CCC GTG TCG GCA GAC TGT CCT GT? ACA 5198? Le Arg Pro Thr Met Gly Gly Pro Val Ser Wing Aso Cys Pro Val Thr 155 160 165 TGT ACC GTT ATA TGT CAG CGA CCC AGG CCT CTA AGT ACC ATG TCT TCC 5246 Cy = Thr Val He Cys Gln Arg ro Arg Pro Leu Ser Thr Met Ser Ser 170 175 180 ATC ATT AGA GAT GCC CGC GTG TAT CTT CAT TTA GAA CGA CGC GAT TAT 5294 He He Arg A = p Wing Arg Val Tyr Leu His Leu Glu Arg Arg Asp Tyr 185 190 195 TAT GAA GTC TAC GCC TCT GTC CTC TCT AAT GCG ATG AGT AAA TAAAAACGCA 5346 Tyr Glu Val Tyr Wing Ser Val Leu Ser Asn Wing Met Ser Lys 200 205 210 CCTCTAACGG TTACTGTGTT TATTATCCAA TCACACCATA GACATTATTA CAATAATATG 5406 GATCTTTATT TCATATAATG 5426 (2) INFORMATION FOR SEQ ID NO: 2: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 369 amino acids (B) TYPE: amino acids P 15 0 1/9 91IX (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: protein (xi) DESCRIPTION OF SEQUENCE: SEQ ID NO: 2 Mot Mot? Gly He Thr Val Wing Cys Asp His Thr Wing Gly Glu Wing 1 5 10 15 His Thr Pro Glu Asp Met Gln Lys Lys Trp Arg He He Leu Wing Gly 20 25 30 Glu Lys Phe Met Thr He Ser Wing Ser Leu Lys Ser He Val Ser Cys 35 40 45 Val Lys Asn Pro Leu Leu Thr * Phe Gly Ala? sp Gly Leu He Val Gln 50 55 60 Gly Thr Val Cys Gly Gln Arg He Phe Val Pro He Asp Arg Asp Ser 65"70 75 80 Phe Ser Glu Tyr Glu Trp His Gly Pro Thr Wing Met Phe Leu Ala Leu 85 90 95 Thr Asp Ser Arg Arg Thr Leu Leu Asp Wing Phe Lys Cys Glu Lys Arg 100 105 110. Arg Afai He Asp Val Ser Phe Thr Phe Wing Gly Glu Pro Pro Cys Arg 115 120 125 His Leu He Gln Wing Val Thr Tyr Met 'Thr Asp Gly Gly Ser Val Ser 130 135 140 Asn Thr He He Lys Tyr Glu Leu Trp Asn Wing Be Thr He Phe Pro 145 150 155 160 Gln Lys Thr Pro Asp Val Thr Phe Ser Leu Asn Lys Gln Gln Leu Asn 165 170 175 Lys He Leu Alk Val Wing Ser Lys Leu Gln His Glu Glu Leu Val Phe 180 185 190 Ser Leu Lys Pro Glu Gly Gly Phe Tyr Val Gly Thr Val Cys Thr Val 195 200 205 He Ser Phe Glu Val Asp Gly Thr Ala Met Thr Gln Tyr Pro Tyr Asn 210 215 220 Pro Pro Thr Be Ala Thr Leu Ala Leu Val Val Ala Cys Arg Lys Lys 225 230 235 240 Lys Ala Asn Lys Asn Thr He Leu Thr Ala Tyr Gly Ser Gly Lys Pro 245 250 255 Phe Cys Val Wing Leu Glu Asp Thr Ser Wing Phe Arg Asn He Val Asn 260 265 270 Lys He Lys Wing Gly Thr Ser Gly Val Asp Leu Gly Phe Tyr Thr Thr 275 280 285 Cys Asp Pro Pro Met Leu Cys He Arg Pro His Wing Phe Gly Ser Pro 290 295 300 Thr Wing Phe Leu Phe Cys Asn Thr Asp Cys Met Thr He Tyr Glu Leu 9MX 305 310 315 320 Glu Glu Val Ser Wing Val Asp Gly Wing He Arg Wing Lys Arg H Asn 325 330 335 Glu Tyr Phe Pro Thr Val Ser Gln Wing Thr Sar Lys Lys Arg Lys Gln 340 345 350. be Pro Pro Pro He Glu Arg Glu Arg Lys Thr Thr Arg Wing Asp Thr 35 = 360 365 Gln (2) INFORMATION FOR SEQ ID NO: 3: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 422 amino acids (B) TYPE: amino acids (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: protein (xi) DESCRIPTION OF THE SEQUENCE: SEQ ID NO: 3 Mee Pro Ser Gly Wing Being Ser Pro Pro Pro Wing Tyr Thr Being Wing 1 5 10 15 Wing Pro Leu Glu Thr Tyr Asn Ser Trp Leu Ser Wing Phe Ser Cys Wing 20 25 30 Tyr Pro Gln Cys Thr Wing Gly Arg Gly His Arg Gln Asn Gly Lys Lys 35 40 45 Cys He Arg Cys He Val He Ser Val Cys Ser Leu Val Cyß He Wing 50 55 60 Wing His Leu Wing Val Thr Val Ser Gly Val Wing Leu He Pro Leu He 65 70 75 80 Asp Gln Asn Arg Wing Tyr Gly Asn Cys Thr Val Cys Val He Wing Gly 85 90 95 Glu Thr Leu Mee Leu Val Gly Lys Pro Wing Gln Phe He Phe Wing He He Wing 115 120 125 Ser Val Wing Glu Thr Leu He Asn Asn Glu Ala Leu Ala He Ser? Sn 130 135"140 Thr Thr Tyr Lys Thr Ala Leu Arg He He Glu Val Thr Ser Leu Ala 145 150 155 160 Cys Phe val Met Leu Gly Wing He He Thr Ser His A = n Tyr Val Cys 165 170 175 He Ser Thr Wing Gly Asp Leu Thr Trp Lys Gly Gly He Phe His Wing 180 185 190 P1501 / 9911X Tyr His Gly Thr Leu Leu Gly He Thr He Pro Asn He His Pro He 195 200 205 Pro Leu Wing Gly Phe Leu Wing Val Tyr Thr He Leu Wing He Asn He 210 215 220 Wing Arg Asp Wing Being Wing Thr Leu Leu Ser Thr Cys Tyr Tyr Arg Asn 225 230 235 240 Cys Arg Glu Arg Thr He Leu Arg Pro Ser Arg Leu Gly His Gly Tyr. 245 250 255 * Thr He Pro Ser Pro Gly Wing Asp Met Leu Tyr Glu Glu Asp Val Tyr 260 265 270 Ser Phe Asp Wing Wing Lys Gly His Tyr Ser Being He Phe Leu Cys Tyr 275 280 285 Wing Met Gly Leu Thr Thr Pro Leu He He Wing Leu His Lys Tyr Met 290 295 300 Wing Gly He Lys Asn Ser Ser Asp Trp Thr Wing Thr Leu Gln Gly Met 305 310 315 320 Tyr Gly Leu Val Leu Gly Ser Leu Be Ser Leu Cys He Pro Ser Ser 325 330 335 Asn Asn Asp Ala Leu He Arg Pro He Gln He Leu He Leu He He 340 345 350 Gly Ala Leu Ala He Ala Leu Ala Gly Cys Gly Gln He He Gly Pro 355 360 365 Thr Leu Phe Wing Wing Being Wing Wing Met Being Cys Phe Thr Cys He 370 375 380 Asn He Arg Wing Thr Asn Lys Gly Val Asn Lys Leu Wing Wing Wing Ser 385 390 395 400 Val Val Lys Ser Val Leu Gly Phe He He Ser Gly Met Leu Thr Cys. 405 410 415 Val Leu Leu Pro Leu Ser 420 (2) INFORMATION FOR SEQ ID NO: 4: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 489 amino acids (B) TYPE: amino acids (D) TOPOLOGY: linear (ii) TI PO OF MOLECULE: protein (xi) ) DESCRI PC OF THE SECUENC IA: SEQ ID NO: 4 Met Val Be Asn Met Arg Val Leu Arg Val Leu Arg. Leu Thr Gl and Trp 1 5? O 15 Val Gly He Phe Leu Val Leu Ser Leu Gln Gln Thr Ser Cys Ala Gly P1501 / 99MX 20 25 30 Leu Pro Hi = Aan Val Asp Thr His His He Leu Thr Phe Asn Pro Ser 35 40 45 Pro He Ser Wing Asp Gly Val Pro Leu Ser Glu Val Pro Asn Pro Pro 50 55 60 Thr Thr Glu Leu Ser Thr Thr Val Wing Thr Lys Thr Wing Val Pro Thr 65 70 75 80 Thr Glu Ser Thr Ser Ser Ser Glu Ala His Arg Asn Ser Ser His Lys 85 90 95 He Pro Asp He He Cys Asp Arg Glu Glu Val Phe Val Phe Leu Asn 100 105 110 Asn thr Gly Arg He Leu Cys Asp Leu He Val Asp Pro Pro Ser Asp 115 120 125 Asp Glu Trp Ser Asn Phe Ala Leu Asp Val Thr Phe ? sn Pro He Glu 130 135 140 Tyr His Wing Asn Glu Lys Asn Val Glu Val Wing Arg Val Wing Gly Leu 145 150 155 160 Tyr Gly Val Pro Gly Ser Asp Tyr Wing Tyr Pro Arg Lys Ser Glu Leu 165 170 175 I Be Being Arg Arg Asp Pro Gln Gly Being Phe Trp Thr Ser Pro t, 180 185 190 Thr Pro Arg Gly Asn Lys Tyr Phe He Trp lie Asn Lys Thr Het His 195 200 205 Thr Met Gly Val Glu Val Arg Asn Val Asp Tyr Lys Asp Asn Gly Tyr 210 215 t. 220 Phe Gln al He Leu Arg Asp Arg Phe Asn Arg Pro Leu Val Glu Lys 225 230 235 240 His He Tyr Met Arg Val Cys Gln Arg Pro Wing Ser Val Asp Val Leu 245 250 255 Wing Pro Pro Val Leu Ser Gly Glu Asn Tyr Lys Wing Ser Cys He Val 260 265 270 Arg His Phe Tyr Pro Pro Gly Ser Val Tyr Val Ser Trp Arg Arg Asn 275 280 285 Gly Asn He Wing Thr Pro Arg Lys Asp Arg Asp Gly be Phe Trp Trp 290 295 - 300 Phe Glu Ser Gly Arg Gly Wing Thr Leu Vil Ser Thr He Thr Leu Gly 305 310 315 320 Asn Ser Gly Leu Glu Pro Pro Pro Lys Val Ser Cys Leu Val Wing Trp 325 330 335 Arg Gln Gly Asp Met He Ser Ser Thr Ser Asn Ala Thr Ala Val Pro Thr 340 345 350 Val Tyr Tyr Hia Pro Arg He Ser Leu Ala Phe Lys A = p Gly Tyr Ala 9MX 355 360 365 He Cys Thr He Glu Cys Val Pro Ser Gly He Thr Val Arg Trp Leu 370 375 380 Val His Asp Glu Pro Gln Pro Asn Thr Thr Tyr Asp Thr Val Val Thr 385 390 395 400 r Gly Leu Cys Arg Thr He Asp Arg Tyr Arg Asn Leu Wing Ser Arg He 405 410 415 Pro Val Gln Asp Asn Trp Wing Lys Thr Lys Tyr Thr Cys Arg Leu He 420 425 430 Gly Tyr Pro Phe Asp Val Asp Arg Phe Gln Asn Ser Glu Tyr Tyr Asp 435 440 445 Wing Thr Pro Wing Wing Arg Gly Met Pro Met He Val Thr He Thr Wing 450 455 460 Val Leu Gly Leu Wing Leu Phe Leu Gly He Gly He He He Thr Wing 465 470 475 480 Leu Cys Phe Tyr Leu Pro Gly Arg Asn 485 (2) INFORMATION FOR SEQ ID NO: 5 (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 212 amino acids (B) TYPE: amino acids (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: protein (xi) DESCRIPTION OF THE SEQUENCE: SEQ ID NO: 5 Met Met Ser Pro Thr Pro Glu Asp Asp Arg Asp Leu Val Val Val Arg 1 5 10 15 Gly Arg Leu Arg Met Met Asp Ser Gly Thr Glu Thr Asp Arg Glu Gln 20 25 30 Arg His Pro Arg Thr Thr Trp Arg Ser He Cys Cys Gly Cys Thr He 35 40 45 Gly Met Val Phe Thr He Phe Val Leu Val Ala Wing Val Leu Leu Gly 50 55 60 Ser Leu Phe Thr Val Ser Tyr Met Wing Met Glu Ser Gly Thr Cys Pro 65"70 75 80 Asp Glu Trp He Gly Leu Gly Tyr Ser Cys Met Arg Val Wing Gly Lys 85 90 95 Asn Wing Thr Asp Leu Glu Wing Leu Asp Thr Cys Wing Arg His Asn Ser 100 105 110 Lys Leu He Asp Phe Wing Asn Wing Lys Val Leu Val Glu Ala He Ala 115 120 125 P1501 / 99MX Pro Phe Gly Val Pro Asn Wing Wing Tyr Gly Glu Val Phe Arg Leu Ring 130 135 140 Asp Ser Lys Thr Thr Cys He Axg pro Thr Met Gly Gly Pro Val Ser 145 150 155 i60 Wing A = p Cys Pro Val Thr Cys Thr Val He Cys Gln Arg Pro Ara Pra 165 170 175 Leu Ser Thr Met Ser Ser He He Arg Asp Ala Arg Val Tyr Leu His 180 185 190 Leu Glu Arg Arg Asp Tyr Tyr Glu Val Tyr Ala Ser Val Leu Ser Asn 195 200 205 Ala Met Ser Lys 210 (2) INFORMATION FOR SEQ ID NO: 6: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 1506 base pairs (B) TYPE: nucleic acid (C) CHAIN TYPE: double (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: DNA (genomic) (iii) HYPOTHETICAL: NO (iv) ANTI-SENSE: NO (ix) CHARACTERISTICS: (A) NAME / KEY: CDS (B) LOCATION: 1..1506 (xi) DESCRIPTION OF THE SEQUENCE: SEQ ID NO: 6 ATG CTC ACG CCG CGT GTG TTA CGA GCT TTG GGG TGG ACT GGA CTC TTT 48 Met Leu Thr Pro Arg Val Leu Arg Ala Leu Gly Trp Thr Gly Leu Phe 1 5 10 '15 TTT TTG CTT TTA TCT CCG AGC AAC GTC CTA GGA GCC AGC CTT AGC CGG 96 Ph? Leu Leu Leu Ser Pro As Asn Val Leu Gly Wing Ser Leu Ser Arg 20 25 30 GAT CTC GAA ACA CCC CCA TTT CTA TCC TTT GAT CCA TCC AAC ATT TCA 144 Asp Leu Glu Thr Pro Pro Phe Leu Ser Phe Asp Pro Ser Asn He Ser 35 40 45? TT? AC GGC GCG CCT TT? ACT G? G GT? CCT C? T GC? CCT TCC AC? GA? 1D2 He? Sn Gly? The Pro Leu Thr Glu Val Pro His Wing Pro Ser Thr Glu 50 55 60 AGT GTG TCA ACA AAT TCG GAA AGT ACC AAT GAA CAT ACC ATA ACA G ?? 240 Ser Val Ser Thr Asn Sor Glu Ser Thr Asn Glu Hxs Thr He Thr Glu 65 70 75 80 ACG ACG GGC AAG AAC GCA TAC ATC CAC AAC AAT GCG TCT ACG GAC AAG 288 Thr Thr Gly Lys Asn Wing Tyr He Hie Asn Asn Wing Being Thr Asp Lys P1501 / 99IIX 85 90 95 CA.A.T? T GCG AAC GAC ACT CAT AAA ACG CCC AAT ATA CTC TGC GAT ACG 336 Gln Asn Wing Asn Asp Thr His Lys Thr Pro Asn He Leu Cys Asp Thr 100 105 110 GAA GAA GTT TTT GTT TTC CTT AAC GAA ACG GGA AGA TTT GTT TGT ACT 384 Glu Glu Val Phe Val Phe Leu Asn Glu Thr Gly Arg Phe Val Cys Thr 115 120 125 í CTC AAA GTC GAC CCC CCC TCG GAT AGT GAA TGG TCC AAC TTT GTT CTA 432 Leu Lys Val Asp Pro Pro Ser Asp Ser Glu Trp Ser Asn Phe Val Leu 130 135 140 GAT CTG ATC TTT AAC CCA ATT GAA TAC CAC GCC AAC GAA AAG AAT GTG 480 hr--} Leu He Phe Asn Pro He Glu Tyr His Wing Asn Glu Lys Asn Val J15 150 155 160 GAA GCG GCG CGT ATC GCT GGT CTC TAT GGA GTC CCC GGA TCA GAC TAT 528 Glu Ala Ala Arg He Ala Gly Leu Tyr Gly Val Pro Gly Ser Asp Tyr 165 170 175 GCA TAC CCA CGT CA T TCT GAA TTA ATT TCT TCG ATT CGA CGA GAT CCC 576 Tyr wing Pro Arg Gln Ser Glu Leu Be Ser Be Arg Arg Asp Pro 180 185 190 CAG GGC ACA TTT TGG ACG AGC CCA TCA CCT CAT GGA AAC AAG TAC TTC 624 Gln Gly Thr Phe Trp Thr Ser Pro Pro Pro His Gly Asn Lys Tyr Phe 195 200 205 ATA TGG ATA AAC AAA ACA ACC AAT ACG ATG GGC GTG GAA ATT AGA AAT 672 He Trp He Asn Lys Thr Thr Asn Thr Met Gly Val Glu He Arg Asn 210 215 220 GTA GAT TAT GCT GAT AAT GGC TAC ATG CAÁ GTC ATT ATG CGT GAC CAT 720 Val Asp Tyr Wing Asp Asn Gly Tyr Met Gln Val He Met Arg Asp His 225 230 235 240 TTT AAT CGG CCT TTA ATA GAT AAA CAT ATT TAC ATA CGT GTG TGT CAA 768 Phe Asn Arg Pro Leu He Asp Lys His He Tyr He Arg Val Cys Gln 245 250 255 CGA CCT GCA TCA GTG GAT GTA CTG GCC CCT CCA GTC CTC AGC GGA GAA 816 Arg Pro Wing Ser Val Asp Val Leu Wing Pro Pro Val Leu Ser Gly Glu 260 265 270 AAT TAC AAG GCA TCT TGT ATC GTT AGA CAC TTT TAT CCC CCT GGA TCT 864 Asn Tyr Lys Wing Ser Cys He Val Arg His Phe Tyr Pro Pro Gly Ser 275 280 285 GTC TAT GTA TCT TGG AGA CAG AAT GGA AAC ATT GCA ACT CCT CGG AAA 912 Val Tyr Val Ser Trp Arg Gln Asn Gly Asn He Wing Thr Pro Arg Lys 290 295 300 GAT CGC GAT GGA AGT TTT TGG TGG TTC GAA TCT GGT AGA GGA GCT ACG 960 Asp Arg Asp Gly Ser Phe Trp Trp Phe Glu Ser Gly Arg Gly Ala Thr 305 310 315 320 TTG GTT TCT ACA ATA ACA TTG GGA AAT TCA GGA ATT GAT TTC CCC CCC 1008 Leu Val Ser Thr He Thr Leu Gly Asn Ser Gly He Asp Phe Pro Pro 325 330 335 AAA ATA TCT TGT CTG GTT GCC TGG AAG CAG GGT GAT ATG ATC AGC ACG 1056 MX Lys He Ser Cys Leu Val Wing Trp Lys Gln Gly Asp Mat He Ser Thr 340 345 350 ACG AAT GCC ACA GCT ATC CCG ACG GTA TAT CAT CAT CCC CGT TTA TCC 1104 Thr Asn Ala Thr Ala He Pro Thr Val Tyr His His Pro Arg Lau Ser 355 360 365 CTG GCT TTT AAA GAT GGG TAT GCA ATA TGT ACT ATA GAA TGT GTC CCC 1152 Leu Ala Phe Lys Asp Gly Tyr Ala He Cys Thr He Glu Cys Val Pro 370 375 380 TCT GAG ATT ACT GTA CGG TGG TTA GTA CAT GAT GAA GCG CAG CCT AA¿ 1200 Ser Glu He Thr Val Arg Trp Leu Val His Asp Glu Ala Gln Pro Asn 385 390 395 400 ACA ACT TAT AAT ACT GTG GTT ACA GGT CTC T < "-C CGG ACC ATC GAT CGC 1248 Thr Thr Tyr Asn Thr Val Val Thr Gly Leu Cvs Arg Thr He Asp Arg 405 410 '415 CAT AGA AAT CTC CTC AGC CGC ATT CCA GTA TGG GAC AAT TGG ACG AAA 1296 His Arg Asn Leu Leu Ser Arg He Pro Val Trp Asp Asn Trp Thr Lys 420 425 430 ACA AAA TAT ACG TGC AGA CTC ATA GGC TAC CCC TTC GAT GAA GAT AAA 1344 Thr Lys Tyr Thr Cys Arg Leu He Gly yr Pro Phe Asp Glu Asp Lys 435 440 445 TT CAA GAT TCG GAA TAT TAC GAT GCA ACT CCA TCT GCA AGA GGA ACA 1392 Phe Gln Asp Ser Glu Tyr Tyr Asp Ala Thr Pro Ser Wing Arg Gly Thr 450 455 460 CCC ATG GTT ATT ACG GTT ACG GCA GTT TTG GGA TTG GCT GTA ATT TTA 1440 Pro Met Val He Thr Val Thr Ala Val Leu Gly Leu Ala val He Leu 465 470 475 480 GGG ATG GGG ATA ATC ATG ACT GCC CTA TGT TTA TAC AAC TCC ACA CGA 1488 Gly Met Gly He Met Met Thr Ala Leu Cys Leu Tyr Asn Ser Thr Arg 4T5 490 49S AAA AAT ATT CGA TTA TAA 1S06 Lys Asn He Arg Leu 500 (2) INFORMATION FOR SEQ ID NO: 7: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 501 amin oacids (B) TYPE: amino acids (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: protein (xi) DESCRI PC OF SECUENC IA: SEQ ID NO: 7: Met Leu Thr Pro Arg Val Leu Arg Ala eu Gly Trp Thr Gly Leu Phe 1 5 10 15 Phe Leu Leu Leu Ser Pro Ser Asn Val Leu Gly Wing Ser Leu Ser Arg 20 25 30 Asp Leu Glu Thr Pro Pro Phe Leu Ser Phe Asp Pro Ser Aan He Ser P1501 / 99HX 35 40 45 He Asn Gly Ala Pro Leu Thr 'Glu Val Pro His Ala Pro Ser Thr Glu 50 55 60 Ser Val Ser Thr Asn Ser Glu Ser Thr Asn Glu His Thr He Thr Glu 65 70 75 80 Thr Thr Gly Lys Asn Wing Tyr He His Asn Asn Wing Being Thr Asp Lys »85 90 • 95 Gln Asn Wing Asn Asp Thr His Lys Thr Pro Asn He Leu Cys Asp Thr 100 105 110 Glu Glu Val Phe Val Phe Leu Asn Glu Thr Gly Arg Phe Val Cys Thr 115 120 125 Leu Lys Val Asp Pro V co Ser Asp Ser Glu Trp Ser Asn Phe Val Leu 130 135 140 Asp Leu He Phe Asn Pro He Glu Tyr His Wing Asn Glu Lys Asn Val 145 150 155 160 Glu Ala Ala Arg He Ala Gly Leu Tyr Gly Val Pro Gly Ser Asp Tyr 165 170 175 Wing Tyr Pro Arg Gln Ser Glu Leu He Be Ser Arg Arg Asp Pro 180 185 190 Gln Gly Thr Phe Trp Thr Ser Pro Pro Pro His Gly Asn Lys Tyr Phe 195 200 205 He Trp He A3n Lys Thr Thr Asn Thr Met Gly Val Glu He Arg Asn 210 215 220 Val Asp Tyr Wing Asp Asn Gly Tyr Met Gln Val He Met Arg Asp His 225 230 235 240 Phe Asn Arg Pro Leu He Asp Lys His He Tyr He Arg Val Cys Gln 245 250 255 Arg Pro Ala Val Ser Val Asu Ala Pro Pro Val Leu Ser Gly Glu 260 265 270 Asn Tyr Lys Ala Ser Cys He Val Arg His Phe Tyr Pro Pro Gly Ser 275 280 285 Val Tyr Val Ser Trp Arg Gln Asn Gly Asn He Ala Thr Pro Arg Lys 290 295 300 A3p Arg Asp Gly Ser Phß Trp Trp Phe Glu Ser Gly Arg Gly Ala Thr 305 310 315 320 Leu Val Ser Thr He Thr Leu Gly Asn Ser Gly He Asp Phe Pro Pro 325 330 335 Lys He Ser Cys Leu Val Wing Trp Lys Gln Gly Asp Met He Ser Thr 340 345 350 Thr Asn Wing Thr Wing He Pro Thr Val Tyr His His Pro Arg Leu Ser 355 360 365 Leu Wing Phe Lya A3p Gly Tyr Wing He Cya Thr He Glu Cys Val Pro 99MX 370 375 380 Ser Glu He Thr Val Arg Trp Leu Val His Asp Glu Wing Gln Pro Asn 385 390 395 400 Thr Thr Tyr Asn Thr Val Val Thr Gly Leu Cya Arg Thr He Asp Arg 405 '410 415 His Arg Asn Leu Leu Ser Arg He Pro Val Trp Asp Asn Trp Thr Lys 420 425 «30 Thr Lys Tyr Thr Cys Arg Leu He Gly Tyr Pro Phe Asp Glu Asp Lys 435 440 445 Phe Gln Asp Ser Glu Tyr Tyr Asp Ala Tfrr Pro Ser Ala Arg Gly Thr 450 455 460 Pro Met Val? Le Thr Val Thr. Ala Val Leu Gly Leu Ala Val He L * ÍUU 465 470 475 480 Gly Met Gly He Met Met Thr Ala Leu Cys Leu Tyr Asn Ser Thr Arg 485 490 495 Lys Asn He Arg Leu 500 (2) INFORMATION FOR SEQ ID NO: 8: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 1734 base pairs (B) TYPE: nucleic acid (C) TYPE OF CHAIN: double (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: DNA (genomic) (iii) HYPOTHETICAL: NO (iv) ANTI-SENSE: NO (ix) CHARACTERISTICS: (A) NAME / KEY CDS (B) LOCATION: 1..1734 (xi) DESCRIPTION OF THE SEQUENCE: SEQ ID NO: 8 AiG GAC CGC GCC GTT AGC CAA GTT GCG TTA GAG AAT GAT GAA AGA GAG 48 Met Asp Arg Ala Val Ser Gln Val Ala LEU Glu Asn Asp Glu Arg Glu 1 5 - 10 15 GCA AAA AAT ACA TGG CGC TTG ATA TTC CGG ATT GCA ATC TTA TTC TTA 96 Wing Lys Asn Thr Trp Arg Leu He Phe Arg He Wing He Leu Phe Leu 20 25 30 ACA GTA GTG ACC TTG GCT ATA TCT GTA GCC TCC CTT TTA TAT AGC ATG 144 Thr Val Val Thr Leu Wing He Ser Val Wing Ser Leu Leu Tyr Ser Met 35 40 45 GGG GCT AGC ACA CCT AGC GAT CTT GTA GGC ATA CCG ACT AGG ATT TCC 192 P1501 / 99MX Gly Wing Ser Thr Pro Ser Asp Leu Val Gly He Pro Thr Arg He Ser 50 55 60 AGG GCA GAA GAA AAG ATT ACA TCT ACA CTT GGT TCC AAT CAA GAT GTA 240 Arg Wing Glu Glu Lys He Thr Ser Thr Leu Gly Ser Asn Gln Asp Val 65 70 75 80 GTA GAT AGG ATA TAT AAG CAG GTG GCC CTT GAG TCT CCA TTG GCA TTG 288 Val Asp Arg He Tyr Lys Gln Val Ala Leu Glu Ser Pro Leu Ala Leu 85 90 95 TTA AAT ACT GAG ACC ACA ATT ATG AAC GCA ATA ACA TCT CTC TCT TAT 336 Leu Asn Thr Glu Thr He Met Asn Wing He Thr Ser Leu Ser Tyr 100 105 110 CAG ATT AAT GGA GCT GCA AAC AAC AGC GGG TGG GGG GCA CCT AT *** 1 CAT 384 Gln He Asn Gly Ala Ala Asn Asn Ser Gly Trp Gly Wing Pro He His 115 120 125 GAC CCA GAT TAT ATA GGG GGG ATA GGC AAA GAA CTC ATT GTA GAT GAT 432 Asp Pro Asp Tyr He Gly Gly He Gly Lys Glu Leu He Val Asp Asp 130 135 140 GCT AGT GAT GTC ACA TCA TTC TAT CCC TCT GCA TTT CAA GAA CAT CTG 480 Wing Ser Asp Val Thr Ser Phe Tyr Pro Ser Wing Phe Gln Glu His Leu 145 150 155 160 AAT TTT ATC CCG GCG CCT ACT ACA GGA TCA GGT TGC ACT CGA ATA CCC 528 Asn Phe He Pro Wing Pro Thr Thr Gly Ser Gly Cys Thr Arg He Pro 165 170 175 TCA TTT GAC ATG AGT GCT ACC CAT TAC TGC TAC ACC CAT AAT GTA ATA 576 Ser Phe Asp Met Ser Wing Thr Hia Tyr Cys Tyr Thr His Aßn Val He 180 185 190 TTG TCT GGA TGC AGA GAT CAC TCA CAC TCA CAT CAG TAT TTA GCA CTT 624 Leu Ser Gly Cys Arg Asp His Ser His Ser His Gln Tyr Leu Ala Leu 195 200 205 GGT GTG CTC CGG ACA TCT GCA ACA GGG AGG GTA TTC TTT TCT ACT CTG 672 Gly Val Leu Arg Thr Ser Ala 'Thr Gly Arg Val Phe Phe Be Thr Leu 210 215 220 CGT TCC ATC AAC CTG GAC GAC ACC CAA AAT CGG AAG TCT TGC AGT GTG 720 Arg Ser He Asn Leu Asp Asp Thr Gln Asn Arg Lys Ser Cys Ser Val 225 230 235 240 AGT GCA ACT CCC CTG GGT TGT GAT ATG CTG TGG TCG AAA GCC ACG GAG 768 Be Ala Thr Pro Leu Gly Cys Asp Met Leu Cys Ser Lys Ala Thr Glu 245 250 255 ACA GAG GAA GAA GAT TAT AAC TCA GCT GTC CCT ACG CGG ATG GTA CAT 816 Thr Glu Glu Glu Asp Tyr Asn Ser Wing Val Pro Thr Arg Met Val His 260 265 270 GGG AGG TTA GGG TTC GAC GGC CAA TAT CAC GAA AAG GAC CTA GAT GTC 864 Gly Arg Leu Gly Phe Asp Gly Gln Tyr His Glu Lys Asp Leu Asp Val 275 280 285 ACA TTA TTC GGG GAC TGG GTG GCC AAC TAC CCA GGA GTA GGG GGT 912 Thr Thr Leu Phe Gly Asp Trp Val Wing Asn Tyr Pro Gly Val Gly Gly 290 295 300 TTT ATT GAC AGC CGC GTG TGG TTC TCA GTC TGA GGG TTA 960 Gly Ser Phe He Asp Ser Arg Val Trp Ph * Ser Val Tyr Gly Gly Leu 305 310 315 320 AAA CCC AAT ACA CCC AGT GAC ACT GTA CAG GAA GGG AAA TAT GTG ATA 1008 Lys Pro Asn Thr Pro Ser Asp Thr Val Gln Glu Gly Lys Tyr Val He 325 330 335 TAC AAG CGA TAC AAT GAC ACA TGC CCA GAT GAG CA CA GAC TAC CAG ATT ÍL056 Tyr Lys Are; Tyr Asn A = p. Thr Cys Pro Asp Glu Gln As Tyr Gln He 340 345 350 CGA ATG GCC AAG TCT TCG TAT AAG CCT GGA CGG TTT GGT GGG AAA CGC 1104 Arg Met Wing Lys Ser Ser Tyr Lys Pro Gly Arg Phe Gly Gly Lys Arg 355 360 365 ATA CAG CAG GCT ATC T? A TCT ATC AAA GTG TCA ACA TCC TTA GGC GAA 1152 He Gln Gln Ala Ha Leu Ser He Lys Val Ser Thr Ser Leu Gly Glu 370 375 380 GAC CCG GTA CTG ACT GTA CCG CCC AAC ACA GTC ACA CTC A? G GGG GCC 1200 Asp Pro Val Leu Thr Val Pro Pro Asn Thr Val Thr Leu Met Gly Wing 385 390 395 400 GAA GGC AGA ATT CTC ACA GTA GGG ACA TCC CAT TTC TTG TAT CAG CGA 1248, Glu Gly Arg He Leu Thr Val Gly Thr Ser His Phe Leu Tyr Gln Arg 405 410 415 GGG TCA TCA TTC TTC TCT CCC GCG TTA TTA TAT CCT ATG ACA GTC AGC 1296 Gly Ser Ser Tyr Phe Ser Pro Wing Leu Leu Tyr Pro Met Thr Val Ser 420 425 430 AAC AAA ACA GCC ACT CTT CAT AGT CCT TAT ACA TTC AAT GCC TTC ACT 1344 Asn Lys Thr Wing Thr Leu His Ser Pro Tyr . Thr Phe Asn Ala Phe Thr 435 440 445 CGG CCA GGT AGT ATC CCT TGC CAG GCT TCA GCA AGA TGC CCC AAC TCA 1392 Arg Pro Gly Ser II * Pro Cys Gln Wing Ser Wing Arg Cys Pro Aßn Ser 450 455 460 TGT GTT ACT GGA GTC TAT ACA GAT CCA TAT CCC ATC CTA TTC TAT AGA 1440 Cys Val Thr Gly Val Tyr Thr Asp Pro Tyr Pro Lau He Phß Tyr Arg 465 470 475 4B0 AAC CAC ACC TTG CGA GGG GTA TTC GGG ACA ATG CTT GAT GGT GAA CAA 1488 have Hid Thr Leu Arg Gly Val Phe Gly Thr Met Leu Asp Gly Glu Gln 485 490 495 GCA AGA CTT AAC CCT GCG TCT GCA GTA TTC GAT AGC ACA TCC CGC AGT 1536 Ala Arg Leu Asn Pro Ala Sar Ala Val Phß Asp Ser Thr Ser Arg Ser 500 50 * 5 510 CGC ATA ACT CGA GTG AGT TCA AGC AGC ATC AAA GCA GCA TAC ACA ACA 1584 Arg He Thr Arg Val Sar Ser Ser He Lys Ala Ala Tyr Thr Thr .515 520 525 TCA ACT TGT TTT AAA GTG GTC AAG ACC AAT AAG ACC TAT TGT CTC AGC 1632 Ser Thr Cys Phe Lys Val Val Lys Thr Asn Lys Thr? Yr Cys Leu Ser 530 535 540 ATT GCT GAA ATA TCT AAT ACT CTC TTC GGA GAA TTC AGA ATC ßTC CCG 1680 / 99MX He Wing Glu He Ser Asn Thr Leu Phe Gly Glu Phe Arg He Val Pro 545 550 555 560 TTA CTA GTT GAG ATC CTC AAA GAT GAC GGG GTT AGA GAA GCC AGG TCT 1728 Leu Leu Val Glu He Leu Lys Asp Asp Gly Val Arg Glu Ala Arg Ser 565 570 575 GGC TAG_1734_Gly (2) INFORMATION FOR SEQ ID NO: 9: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 577 amino acids (B) TYPE: amino acids (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: protein (xi) DESCRIPTION OF THE SEQUENCE: SEQ ID NO: 9 Met Asp Arg Ala Val Ser Gln Val Ala Leu Glu Asn Asp Glu Arg Glu 1 5"10 15 Ala Lys Asn Thr Trp Arg Leu He Phe Arg He Ala He Leu Phe Leu 20 25 30 Thr Val Val Thr Leu Wing He Ser Val Wing Ser Leu Leu Tyr Ser Met 35 40 45 Gly Wing Ser Thr Pro Ser Asp Leu Val Gly He Pro Thr Arg He Ser 50 55 60 Arg Wing Glu Glu Lys He Thr Ser Thr Leu Gly Ser Asn Gln Asp Val 65 70 75 80 Val Asp Arg He Tyr Lys Gln Val Wing Leu Glu Ser Pro Leu Wing Leu 85 90 95 Leu Asn Thr Glu Thr Thr He Met Asn Wing He Thr Ser Leu Ser Tyr 100 105 110 Gln He Asn Gly Wing Wing Asn Asn Ser Gly Trp Gly Wing Pro He His 115 120 125 Asp Pro Asp Tyr He Gly Gly He Gly Lys Glu Leu He Val Asp Asp 130 135 140 Wing Being Asp Val Thr Ser Phe Tyr Pro Being Wing Phe Gln Glu His Leu 145 150 155 160 Asn Phe He Pro Pro Wing Thr Thr Gly Ser Gly Cys Thr Arg He Pro 165 170 175 Ser Phe Asp Met Ser Wing Thr His Tyr CyS Tyr Thr His Asn Val He 180 185 190 Leu Ser Gly Cys Arg Asp His Ser His Ser His Gln Tyr Leu Ala Leu 195 200 205 Gly Val Leu Arg Thr Ser Ala Thr Gly Arg Val Phe Phe Ser Thr Leu P1501 / 99MX 210 215 220 Arg Ser He Asn Leu Asp Asp Thr Gln Asn Arg Lys Ser Cys Ser Val 225 230 235 240 Be Wing Thr Pro Leu Gly Cys Asp Mss Leu Cys Ser Lys Wing Thr Glu 245 250 255 Thr Glu Glu Glu Asp Tyr Asn Be Wing Val Pro Thr Arg Met Val Hi 260 265 270 Gly Arg Leu Gly Phe A = p Gly Gln Tyr His Glü Lys Asp Leu Asp Val 275 280 285 Thr Thr Leu Phe Gly Asp Trp Val Wing Asn Tyr Pro Gly Val Gly Gly 290 295 300 Gly Ser Phe He Asp Ser Arg Val Trp Phe Ser Val? Yr Gly Gly Leu 305 310 315 320 Lys Pro Asn Thr Pro Ser Asp Thr Val Gln Glu Gly Lys Tyr Val He 325 330 335 Tyr Lys Arg Tyr Asn Asp Thr Cyß Pro Asp Glu Glp Asp Tyr Gln He 340 345 350 Arg Met Wing Lys Ser Ser Tyr Lys Pro Gly Arg Phe Gly Gly Lys Arg 355 360 365 He Gln Gln Ala He Leu Ser He Lys Val Ser Thr Ser Leu Gly Glu 370 375 360 Asp Pro Val Leu Thr Val Pro Pro Asn Thr Val Thr Leu Met Gly Wing 385 390 395 400 Glu Gly Arg He Leu Thr Val Gly Thr Ser His Phe Leu Tyr Gln Arg 405 410 415 Gly Ser Ser Tyr Phe Ser Pro Wing Leu Leu Tyr Pro Met Thr Val Ser 420 425 430 Asn Lys Thr Wing Thr Leu His Ser Pro Tyr Thr Phe Asn Wing Phe Thr 435 440 445 Arg Pro Gly Ser He Pro Cys Gln Wing Be Al ' to Arg Cya Pro Asn Ser 450 455 460 Cys Val Thr Gly val Tyr Thr Asp Pro Tyr Pro Leu He Phe Tyr Arg 465 470 475 480 Asn His Thr Leu Arg Gly Val Phß Gly Thr Mßt Leu Aap GXy Glu Gln 485 - 490 495 Wing Arg Leu Asn Pro Wing Wing Val Phß Asp Ser Thr Ser Arg Sat 500 505 510 Arg He Thr Arg Val Ser Ser Ser Be He Lys Wing Wing Tyr Thr Thr 515 520 525 Ser Thr Cys Phe Lys Val Val Lys Thr Asn Lys Thr Tyr Cya Leu Ser 530 535 540 99MX He Ala Glu He Ser Asn Thr Leu Phe Gly Glu Phe Arg He Val Pro 545 550 555 560 Leu Leu Val Glu He Leu Lys Asp Asp Gly Val Arg Glu Ala Arg Ser 565 570 575 Gly (2) INFORMATION FOR SEQ ID NO: 10: * (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 1662 base pairs (B) TYPE: nucleic acid (C) TI PO CHAIN: double (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: DNA (genomic) (iii) HYPOTHETICAL: NO (iv) ANTI-SENSE: NO (ix) CHARACTERISTICS: (A) NAME / KEY: CDS (B) LOCATION: 1..1662 (xi) DESCRIPTION OF THE SEQUENCE : SEQ ID NO: 10: ATG GGC TCC AGA CCT TCT ACC AAG AAC CCA GCA CCT ATG ATG CTG ACT 48 Met Gly Ser Arg Pro Ser Thr Lys Asn Pro Pro Wing Met Met Leu Thr 1 5 10 15 ATC CGG GTC GCG CTG GTA CTG AGT TGC ATC TGT CCG GCA AAC TCC ATT 96 He Arg Val Ala Leu Val Leu Ser Cys He Cys Pro Wing Asn Ser He 20 25 30 GAT GGC AGG CCT CTT GCA GCT GCA GGA ATT GTG GTT ACA GGA GAC AAA 144 Asp Gly Arg Pro Leu Wing Wing Wing Gly He Val Val Thr Gly Asp Lys 35 40 45 GCA GTC AAC ATA TAC ACC TCA TCC CAG ACA GGA TCA ATC ATA GTT AAG 192 Wing Val Asn He Tyr Thr Ser Ser Gln Thr Gly Ser He He Val Val Lys 50 55 60 CTC CTC CCG AAT CTG CCA AAG GAT AAG GAG GCA TGT GCG AAA GCC CCC 240 Leu Leu Pro Asn Leu Pro Lys Asp Lys Glu Ala Cys Ala Lys Ala Pro 65 70 75 80 TTG GAT GAC TAC AAC AGG ACA TTG ACC ACT TTG CTC ACC CCC CTT GGT 288 Leu Asp Ala Tyr Asn Arg Thr Leu Thr Thr Leu Leu Thr Pro Leu Gly 85 90 95 GAC TCT ATC CGT AGG ATA CAA GAG TCT GTG ACT ACA TCT GGA GGG GGG 336 Asp Ser He Arg Arg He Gln Glu Ser Val Thr Thr Ser Gly Gly Gly 100 105 110 AGA CAG GGG CGC CTT ATA GGC GCC ATT ATT GGC GGT GTG GCT CTT GGG 384 Arg Gln Gly Arg Leu He Gly Wing He He Gly Gly Val Wing Leu Gly P1501 / 99MX 115 120 125 GTT GCA ACT GCC GCA CAA ATTA GCG GCC GCT GCT CTG ATA CA CA GCC 432 Val Wing Thr Wing Wing Gln He Thr Wing Wing Wing Wing Leu He Gln Wing 130 135 140 AAA CAA AAT GCT GCC AAC ATC CTC CGA CTT AAA GAG AGC ATT GCC GCA 480 Lys Gln Asn Wing Wing Asn He Leu Arg Leu Lys Glu be He Wing Wing 145 150 155 160 ACC AAT GAG GCT GTG CAT GAG GTC ACT GAC GGA TTA TCG CAA CTA GCA 528 Thr Asn Glu Wing Val His Glu v »l Thr Asp Gly Leu Ser Gln Leu Wing 165 170 175 GTG GCA GTT GGG AAG ATO CAG CAG TTC GTT AAT GAC CAA TTT AAT AAA 576 Val Wing Val Gly Lys Mßt Gln Gln Phen Val Asn Asp Gln Phe Asn Lys 180 185 190 ACA GCT CAG GAA TTA GAC TGC ATC AAA ATT GCA CAG CAA GTT GGT GTA 624 Thr Wing Gln Glu Leu Asp Cys He Lya He Wing Gln Gln Val Gly Val 195 200 205 GAG CTC AAC CTG TAC CTA ACC GAA TCG ACT ACA GTA TTC GGA CCA CAA 672 Glü Leu Asn Leu Tyr Leu Thr Glu Ser Thr Thr Val Phe Gly Pro Gln 210 215 220 ATC ACT TCA CCT GCC TTA AAC AAG CTG ACT ATT CAG GCA CTT TAC ART 720 Ilß Thr Ser Pro Ala Leu Asn Lys Leu Thr He Gln Ala Leu Tyr Asn 225 230 235 240 CTA GCT GGT GGG AAT ATG GAT TAC TTA TTG ACT AAG TTA GG? ATA GGG 768 Leu Wing Gly Gly Asn Met Asp Tyr Leu Leu Thr Lys Leu Gly He Gly 245"250 255 AAC AAT CAA CTC AGC TCA TTA ATC GGT AGC GGC TTA ATC ACC GGT AAC 816 Asn Asn Gln Leu Ser Ser Leu He Gly Ser Gly Leu He Thr Gly Asn 260 265 270 CCT ATT CTA TAC GAC TCA CAG ACT CA CTC TTG GGT ATA CAG GTA ACT 864 Pro lia Leu Tyr Asp Ser Gln Thr Gln Leu Leu Gly He G n Val Thr 275 280 265 CTA CCT TCA GTC GGG AAC CTA? AT AAT ATG CGT GCC ACC TAC TTG GAA 912 Leu Pro Ser Val Gly Aan Leu Asn Asn Mßt Ar? Ala Thr Tyr Leu Glu 290 295 300 ACC TTA TCC GTA AGC ACA ACC AGG GGA TTT GCC TCG GCA CTT GTC CCA 960 Thr Leu Ser to Being Thr Thr Arg Gly Phe Ala Be Ala Leu Val Pro 305 310 315 320 AAA GTG GTG ACA CGG GTC GGT TCT GTG ATA GAA GAA CTT GAC ACC TCA 1008 Lys Val Val Thr Arg Val Gly Ser Val He Glu Glu Leu Aap Thr SER 325 ~ 330 335 TAC TGT ATA GAA ACT GAC TTA GAT TTA TAT TGT ACA AGA ATA GTA ACG 1056 Tyr Cys He Glu Thr Asp Leu Asp Leu Tyr Cys Thr Arg He Val Thr 340 345 350 TTC CCT ATG TCC CCT GGT ATT TAC TCC TGC TTG AGC GGC AAT ACA TCG 1104 Phe Pro Met Ser Pro Gly He Tyr Ser Cys Leu Ser Gly Asn Thr Ser 355 360 365 1 / 99MX GCC TGT ATG TAC TCA AAG ACC GAA GGC GCA CTT ACT ACA CCA TAT ATG 1152 Cys Wing Met Tyr Ser Lys Thr Glu Gly Wing Leu Thr Thr Pro Tyr Met 370 375 380 ACT ATC AAA GGC TCA GTC ATC GCT AAC TGC AAG ATG ACA ACA TGT AGA 1200 Thr He Lys Gly Ser Val He Ala'Asn Cys Lys Met Thr Thr Cys Arg 385 390 395 400 TGT GTA AAC CCC CCG GGT ATC ATA TCG CAA AAC TAT GGA GAA GCC GTG 1248 Cys Val Asn Pro Pro Gly He He Ser Gln Asn Tyr Gly Glu * Wing Val 405 410 415 TCT CTA ATA GAT AAA CAA TCA TGC AAT GTT TTA TCC TTA GGC GGG ATAf 1296 Ser Leu He Asp Lys Gln Ser Cys Asn Val Leu Ser Leu Gly Gly He 420 425 430 AC * TTA AGG CTC AGT GGG GAA TTC GAT GTA ACT TAT CAG AAG AAT ATC 1344 Thr Leu Arg Leu Ser Gly Glu Phe Asp Val Thr Tyr Gln Lys Asn He 435 440 445 TCA ATA CAAT GAT TCT CAÁ GTA ATA ATA ACA GGC AAT CTT GAT ATC TCA 1392 Ser He Gln Asp Ser Gln Val He He Thr Gly Asn Leu Asp He Ser 450 455 460 ACT GAG CTT GGG AAT GTC AAC AAC TCG ATC AGT AAT GCC TTG AAT AAG 1440 Thr Glu Leu Gly Asn Val Asn Asn Ser Asn Asn Ala Leu Asn Lys 465 470 475 480 TTA GAG GAA AGC AAA AGA CTA GAC AAA GTC AAT GTC AAA CTG ACC 1488 Leu Glu Glu Ser Asn Arg Lys Leu Asp Lys Val Asn Val Lys Leu T. hr 485 490 495 AGC ACA TCT GCT CTC ATT ACC TAT ATC GTT TTG ACT ATC ATA TCT CTT 1536 Ser Thr Be Ala Leu He Thr Tyr He Val Leu Thr He He Ser Ser Leu 500 505 510 GTT TTT GGT ATA CTT AGC CTG ATT CTA GCA TGC TAC CTA ATG TAC AAG 1584 Val Phe Gly He Leu Ser Leu He Leu Wing Cys Tyr Leu Met Tyr Lys 515 520 525 CAÁ AAG GCG CAÁ CAÁ AAG ACC TTA TTA TGG CTT GGG AAT AAT ACC CTA 1632 Gln Lys Ala Gln Gln Lys Thr Leu Leu Trp Leu Gly Asn Asn Thr Leu 530 535 540 GAT CAG ATG AGA GCC ACT AAA ATG TGA 1662 Asp Gln Met Arg Ala Thr Thr Lys Met 545 550 (2) INFORMATION FOR SEQ ID NO: 11: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 553 amino acids (B) TI PO: amino acids (D) TOPOLOGY: linear (ii) TI PO OF MOLECULE: protein ( xi) DESCRIPTION OF SECTION IA: SEQ ID NO: 11: Met Gly Be Arg Pro Be Thr Lys Asn Pro Pro Wing Met Met Leu Thr 1 5 *? O 15 P1501 / 99MX He Arg Val Wing Leu Val Leu Ser Cys He Cys Pro Wing Asn Ser He 20 25 30 Asp Gly Arg Pro Leu Wing Wing Wing Gly He Val Val Thr Gly Asp Lys 35 40 45 Wing Val Asn He Tyr Thr Ser Ser Gln Thr Gly Ser He He Val Lys 50 55 60 t Leu Leu Pro Asn Leu Pro Lys Asp Lys Glu Ala Cys Ala Lys Ala Pro 65 70 75 80 Leu Asp Ala Tyr Asn Arg Thr Leu Thr Thr Leu Leu Thr Pro Leu Gly 85 90 95 Asp Being He Arg Arg He Gln Glu Being Val Thr Thr Being Gly Gly Gly 100 105 110 Arg Gln Gly Arg Leu He Gly 'Wing He He Gly Gly Val Wing Ala Leu Gly 115 120 125 val Wing Thr Wing Wing Gln He Thr Wing Wing Wing Wing Leu He Gln Wing 130 135 140 Lys Gln Asn Wing Wing Asn He Leu Arg Leu Lys Glu Wing Be Wing Wing 145 150 155 160 Thr Asn Glu Wing Val His Glu Val Thr Asp Gly Leu Ser Gln Leu Wing 165 170. 175 Val Wing Val Gly Lys Met Gln Gln Phe Val Asn Aap Gln Phe Asn Lys 180 185 190 Thr Wing Gln Glu Leu Asp Cys He Lys He Wing Gln Gln Val Gly Val 195 200 205 Glu Leu Asn Leu Tyr Leu Thr Glu Ser Thr Thr Val Phe Gly Pro Gln 210 215 220 He Thr Ser Pro Ala Leu Asn 'Lys Leu Thr He Glp Ala Leu Tyr Asn 225 230 235 240 Leu Ala Gly Gly Asn Met Asp Tyr Leu Leu Thr Lys Leu Gly He Gly 245 250- 255 Asn Asn Gln Leu Be Ser Leu He Gly Be Gly Leu He Thr Gly Asn 260 265 270 Pro He Leu Tyr Asp Ser Gih Thr Gln Leu Leu Gly He Gln Val Thr 275 2B0 285 Leu Pro Ser Val Gly Asn Leu Asn Asn Met Arg Ala Thr Tyr Leu Glu 290 295"300 Thr Leu Ser Val Ser Thr Thr Arg Gly Phe Wing Ser Wing Leu Val Pro 305 310 315 320 Lys Val Val Thr Arg Val Gly Ser Val He Glu Glu Leu Asp Thr Ser 325 330 335 Tyr Cys He Glu Thr Asp Leu Asp Leu Tyr Cys Thr Arg He Val Thr 340 345 350 9MX Phe Pro Met Ser Pro Gly He Tyr Ser Cys Leu Ser Gly Asn Thr Ser 355 360 365 Wing Cys Met Tyr Ser Lys Thr Glu Gly Wing Leu Thr Thr Pro Tyr Met 370 375 380 Cys Val Asn Pro Pro Gly He He Ser Gln Asn Tyr Gly Glu Wing Val 405 410 415 Ser Leu He Asp Lys Gln Ser Cys Asn Val Leu Ser Leu Gly Gly He 420 425 430 Thr Leu Arg Leu Ser Gly Glu Phe Asp Val Thr Tyr Gln Lys Asn He 435 440 445 Ser He Gln Asp Ser Gln Val He He Thr Gly Asn Leu Asp He Ser 450 455 460 Thr Glu Leu Gly Asn Val Asn Asn Ser He Ser Asn Ala Leu Asn Lys 465 470 475 480 Leu Glu Glu Ser Asn Arg Lys Leu Asp Lys Val Asn Val Lys Leu Thr 485 490 495 Ser Thr Ser Wing Leu He Thr Tyr He Val Leu Thr He He Ser Leu 500 505 510 Val Phe Gly He Leu Ser Leu He Leu Wing Cys Tyr Leu Met Tyr Lys 515 520 525 Gln Lys Wing Gln Gln Lys Thr Leu Leu Trp Leu Gly Asn Asn Thr Leu 530 535 540 Asp Gln Met Arg Wing Thr Thr Lys Met 545 550 (2) INFORMATION FOR SEQ ID NO: 12: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 2681 base pairs (B) TYPE: nucleic acid (C) TYPE OF CHAIN: double (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: DNA (genomic) (iii) HYPOTHETICAL: NO (Ivan IT-DIRECTION: NO (ix) CHARACTERISTICS: (A) NAME / KEY: CDS (B) LOCATION: 146..481 (ix) FEATURES: (A) NAME / KEY: CDS (B) LOCATION: complement (602. . 1402) P1501 / 99MX (ix) FEATURES: (A) NAME / KEY: CDS (B) LOCATION: 1599.2135 (ix) FEATURES: (A) NAME / KEY: CDS (B) LOCATION: add-on (2308..2634) (xi) DESCRIPTION OF THE SEQUENCE: SEQ ID NO: Í12: TTTATCGGAC CTTGGGTATT CAGGGGAACC CATCTGGTTG AAATGCATCC GACCCTGCAC 60 TTGATCCTGG TTACCCCGAC CCAANTTTTA AGCCGGCTGG CGCGGTCCCT AGATAACCCC 120 CCGCTTAAAA CTAGCCCCAA TATTGATGTG CAGATATAAC ACAGNNANCC GATCAATGGA 180 AGACATGCTA CGGCGGTCAT CTCCCGAAGA CATCACCGAT TCCCTAACAA TGTGCCTGAT 240 TATGTTATCG CGCATTCGTC G? ACCATGCG CACCGCAGGA AATAAATATA GCTATATGAT 300 AGATCCAATG AATCGTATGT CTAATTACAC TCCAGGCGAA TGTATGACAG GTATATTGCG 360 ATATATTGAC GAACATGCTA GAAGGTGTCC TGATCACATA TGTAATTTGT ATATCACATG 420 TACACTTATG CCGATGTATG TGCACGGGCG ATATTTCTAT TGTAATTCAT TTTTTTGKTA 80 GTAAACTACC ACAGGCTGTC CGGAAATCTA AGTTAATGAA TAAAGTAGAT GGTTAATACT 540 CATTGCTTAG AATTGGACTA CTTTTAATYC TCTTTAATGT TCGTATTAAA TAAAAACATC 60O TTTAATAAAC TTCAGCCTCT TCGCTTATTG TAGAAATTGA GTATTCAMAA TCATGTTCAA 660 AGCCGTCTTC GGAGAGTGTA CTCGCCACGG TGGTTGGAAC ATCACTATGT CTACACGTCA 720 AATTTAAGCA CGTCAGGTCT GTCGAGGACA AGAAATGGTT AACTAGTGTT TCAATTATTC 780 TTATAAACGT TAAGCATTGT AAGCCCCCCG GCCGTCCGCA GCAACAATTT ACTAGTATGC 840 CGTGGGCTCC GGGACTATCA CGGATGTCCA ATTCGCACAT GCATATAATT TTTCTAGGGT 900 CTCTCATTTC GAGAAATCTT CGGGGATCCA TCAGCAATGC GGGCTGTAGT CCCGATTCCC 960 GTTTCAAATG AAGGTGCTCC AACACGGTCT TCAAAGCAAC CGGCATACCA GCAAACACAG 1020 ACTGCAACTC CCCGCTGCAA TGATTGGTTA TAAACAGTAA TCTGTCTTCT GGAAGTATAT 1080 TTCGCCCGAC AATCCACGGC GCCCCCAAAG TTAAAAACCA TCCATGTGTA TTTGCGTCTT 1140 CTCTGTTAAA AGAATATTGA CTGGCATTTT CCCGTTGACC GCCAGATATC CAAAGTACAG 1200 CACGATGTTG CACGGACGAC TTTGCAGTCA CCAGCCTTCC TTTCCACCCC CCCACCAACA 1260 AAATGTTTAT CGTAGGACCC ATATCCGTAA TAAGGATGGG TCTGGCAGCA ACCCCATAGG 1320 CGCCTCGGCG TGGTAGTTCT CGAGGATACA TCCAAAGAGG TTGAGTATTC TCTCTACACT 1380 TCTTGTTAAA TGGAAAGTGC ATTTGCTTGT TCTTACAATC GGCCCGAGTC TCGTTCACAG 1440 CGCCTCGTTC ACACTTAAAC CACAAATAGT CTACAGGCTA? ATGGGAGCC AGACTGAAAC 1500 • TCACATATGA CTAATATTCG GGGSTGTTAG TCACGTGTAG CCCATTGTGT GCATATAACG 1560 1 / 99MX ATGTTGGACG CGTCCTTATT CGCGGTGTAC TTGATACTAT GGCAGCGAGC ATGGGATATT 1620 CATCCTCGTC ATCGTTAACA TCTCTACGGG TTCAGAATGT TTGGCATGTC GTCGATCCTT 1680 TGCCCATCGT TGCAAATTAC AAGTCCGATC GCCATGACCG CGATAAGCCT GTACCATGTG 1740 GCATTAGGGT GACATCTCGA TCATACATTA TAAGACCAAC GTGCGAGTCT TCCAAAGACC 1800 TGCACGCCTT CTTCTTCGGA TTGTCAACGG GTTCTTCAGA ATCTATGCCC ATATCTGGCG 1860 TTGAGACCAT TGTGCGTTTA ATGAACAATA AAGCGGCATG CCATGGAAAG GAGGGCTGCA 1920 í GATCTCCATT TTCTCACGCC ACTATCCTGG ACGCTGTAGA CGATAATTAT ACCATGAATA 1980 TAGAGGGGGT ATGTTTCCAC TGCCACTGTG ATGATAAGTT TTCTCCAGAT TGTTGGATAT 2040 CTGCATTTTC TGCTGCCGAA CAAACTTCAT CGCTATGCAA AGAGATGCGT. GTGTACACGC 2100 GCCGGTGGAG TATACGGGAA ACTAAATGTT CATAGAGGTC TTTGGGCTAT ATGTTATTAA 2160 ATAAAATAAT TGACCAGTGA ACAATTTGTT TAATGTTAGT TTATTCAATG CATTGGTTGC 2220 AAATATTCAT TACTTCTCCA ATCCCAGGTC ATTCTTTAGC GAGATGATGT TATGACATTG 2280 CTGTGAAAAT TACTACAGGA TATATTTTTA AGATGCAGGA GTAACAATGT GCATAGTAGG 2340 CGTAGTTATC GCAGACGTGC AACGCTTCGC ATTTGAGTTA CCGAAGTGCC CAACAGTGCT 2400 GCGGTTATGG TTTATGCGCA CAGAATCCAT GCATGTCCTA ATTGAACCAT CCGATTTTTC 2460 TTTTAATCGC GATCGATGTT TGGGCAACTG CGTTATTTCA GATCTAAAAA ATTTACCCTY 2520 TATGACCATC ACATCTCTCT GGYTCATACC CCGCTTGGGN TAAGATATCA TGTAGATTCC 2580 GCCCCTAAGA AATTGCAAAC TAACATNATT GNCGGGTTCC ATATACAATC CCATCTTGTC 2640 CNCTCGAAAT TACAAACTCG CGCAATAGAC CCCCGTACAT T 2681 (2) INFORMATION FOR SEQ ID NO: 13: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 111 amino acids (B) TYPE: amino acids (C) TYPE OF CHAIN: double (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: DNA (genomic co) (iii) HYPOTHETICAL: NO (iv) ANTI-SENSE: NO (xi) DESCRIPTION OF THE SEQUENCE: SEQ ID NO: 13 Met Cys Arg Tyx Asn Thr Xaa Xaa Arg Ser Met Glu Asp Met Leu Arg 1 5 10 15 Arg Ser Ser Pro Glu Asp He Thr Asp Ser Leu Thr Met Cys Leu He 20 25 30 P1501 / 99 X Mßt Leu Ser Atg 11, Arg Arg 'Thr Met Arg thr "Al» ßy Aan Lya Tyr • so 0 45. Ser Tyr Met He Aap Pro Mßt Aan Arg Mac Sar Aan Tyr Thr Pro Gly 50 35 go Glu Cys Mßt Thr Gly lie Ltu Arg Tyr Ilß Asp Glu His Wing Arg Arg 69 70 75 80 Cyß Pro Aap His He Cys Asn Leu Tyr lie Thr Cyß Thr Leu Mßt Pro 85. 90 95 Met Tyr Val His Gly Arg Tyr Phe Tyr Cys Aan Ser Phe Phe Xaa 100 ios * no) INFORMATION FOR SEQ ID NO: 14: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 266 amino acids (B) TYPE : amino acids (C) TYPE OF CHAIN: double (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: DNA (genomic) (iii) HYPOTHETICAL: NO (iv) ANTI-SENSE: NO (xi) DESCRIPTION OF THE SEQUENCE: SEQ ID NO: 14 Mer Hia Phe Pro Phe Asn Lys Cys Arg Glu Asn Thr Gln Pro Leu 1 5 10 13 Trp Met Tyr Pro Arg Glu Leu Pro Arg Arg Gly Wing Tyr Gly Val Wing 20 25 30 Wing Arg Pro He Leu He Thr Asp Mßt Gly Pro Thr Xla Asn Xle Leu 35 40 45 Leu Val Gly Gly Trp Lya Gly Arg Lßu Val Thr Ala Lys Ser Ser Val 50 55 60 C'ln His Arg Ala Val Leu Txp xle Ser Gly Gly Gln Arg Glu Asn? The fi5 70 75 80 Ser Gn Tyr Ser Phe Asn Arg Glu Asp? Asn Thr His Gly Trp Phe 85 90 95 Leu Thr Leu Gly Ala Pro Trp lio Val Gly Arg Asn Xle Leu Pro Glu 100 105 110 Asp? Rg Leu Leu Phe Xle Thr Asn His Cys Ser Gly Glu Leu Gln Ser 115 120 125 Val Phß Ala Gly Met Pro Val Ala Leu Lys Thr Val Leu Glu His Leu 130 135 140 His Leu Lys Arg Glu Ser Gly Leu Gln Pro Wing Leu Leu Met Asp Pro 145 150 155 160 Áxg Are Phe Leu Glu MBC Arg Asp Pro Arg Lys Xle Xle Cys Met Cys P1501 / B9MX 165 170 175 Glu Leu Asp lia Arg Asp Ser Pro Gly Wing Hia Gly He Leu Val Asn 180 185 190 Cys Cys Cys Gly Arg Pro Gly Gly Leu Gln Cys Leu Thr Phß He Arg 195 200 205 Ilß lt Glu Thr Lßu Val Asn Hia phß Lßu S r Ser Thr Asp Leu Thr 210 215 220 Cys Leu Asn Leu Thr Cys Arg His Ser Asp Val Pro Thr Thr Val Wing 225 230 235 240 Ser Thr Leu Ser Glu Asp Gly Phe Glu His Asp Xaa Glu Tyr Ser He 245 • 250 255 Ser Thr He Ser Glu Glu Wing Glu Val Tyr 260 265 (2) INFORMATION FOR SEQ ID NO: 15: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 178 amino acids (B) TYPE: amino acids ( C) CHAIN TYPE: double '(D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: DNA (genomic) (iii) HYPOTHETICAL: NO (iv) ANTI-SENSE: NO (xi) DESCRIPTION OF SEQUENCE: SEQ ID NO : fifteen Met Wing Wing Being Met Gly Tyr Being Ser Being Being Leu Thr Being Leu 1 5 10 15 Arg Val Gln Asn Val Trp His Val Val Asp Pro Leu Pro He Val Wing 20 25 30 Asn Tyr Lys Ser Asp Arg His Asp Arg Asp Lys Pro Val Pro Cys Gly 35 40 45 Xle Arg Val hr Ser Arg Ser Tyr Xle Xle Arg Pro Thr Cys Glu Ser 50 55 60 Ser Lys Asp Lßu His Wing Phe Phe Phe Gly Leu Ser Thr Gly Be Ser 65"70 75 80 Glu Ser Met Pro Xle Ser Gly Val Glu Thr Xlß Val Arg Leu Met Asn 85 90 95 Asn Lys Ala Ala Cys His Gly Lys Glu Gly Cys Arg Ser Pro Phß Sar 100 105 110 His Ala Thr Xle Leu Asp Ala Val Asp Asp Asn Tyr Thr Met Asn Xla 115 120 125 Glu Gly Val Cys Phß His Cys His Cys Asp Asp Lys Phß Ser Pro Asp 130- 135 140"cys Trp Xls Ser Ala 'Phe Ser Ala Ala Glu Gln Thr Ser Ser Leu Cys P1S01 / 99MX 145 150 155 160 Lys Glu Met Arg Val Tyr Thr Arg Arg Trp Ser lie Arg Glu Thr Lys 165 170 175 Cys Ser (2) INFORMATION FOR SEQ ID NO: 16: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 108 amino acids (B) TYPE: amino acids (C) TYPE OF CHAIN: double (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: DNA (genomic) (iii) HYPOTHETICAL: NO (iv) ANTI-SENSE: NO (xi) DESCRIPTION OF THE SEQUENCE: SEQ ID NO: 16 Met Gly Leu Tyr Met Glu Pro Xaa Asn Xaa to Ser Leu Gln Phe Lßu 1 5 10 15 Arg Gly Gly He Tyr Met He Ser Xaa Pro Lys Arg Gly Met Xaa Gln 20 25 30 Arg Asp Val Mee Val He Xaa Gly Lys Phß Phß Arg Ser Glu He Thr 35 40 45 Gln Leu Pro Lys His Arg Ser Arg Leu Lys Glu Lys Ser Asp Gly Ser 50 55 60 He Arg Thr Cys Mss Asp Ser Val Arg He Asn His Asn Arg Ser Thr 65 70 75 80 Val Gly His Phe Gly Asn Be Asn Wing Lys Arg Cys Thr Be Wing He 85 90 95 Thr Thr Pro Thr Met His He Val Thr Pro Ala Ser 100 105 (2) INFORMATION FOR SEQ ID NO: 17: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 57 base pairs (B) TYPE: nucleic acid (C) TYPE OF CHAIN: double (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: DNA (genomic) (iii) HYPOTHETICAL: NO (iv) ANTI-SENSE: NO P1S01 / 99HX (xi) DESCRIPTION OF THE SEQUENCE: SEQ ID NO: 17: CTCGGCGTGG TAGTTCTCGA GGCCTTAATT AAGGCCCTCG AGGATACATC CAAAGAG 57 (2) INFORMATION FOR SEQ ID NO: 18: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 63 base pairs (B) TYPE: nucleic acid? (C) CHAIN TYPE: double (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: DNA (genomic) (üi) HYPOTHETICAL: NO (iv) ANTI-SENSE: NO (xi) DESCRIPTION OF SEQUENCE: SEQ ID NO : 18: CGGCGTGGTA GTTCTCGAGG CCTTAAGCGG CCGCTTAAGG CCCTCGAGGA TACATCCAAA 60 GAG 63 (2) INFORMATION FOR SEQ ID NO: 19: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 35 base pairs (B) TYPE: nucleic acid (C) TYPE OF CHAIN: double (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: DNA (genomic) (iii) HYPOTHETICAL: NO (iv) ANTI-SENSE: NO (xi) DESCRIPTION OF SEQUENCE: SEQ ID NO: 19: CGCAGGATCC GGGGCGTCAG AGGCGGGCG AGGTG 35 (2) INFORMATION FOR SEQ ID NO: 20: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 33 base pairs (B) TYPE: nucleic acid (C) CHAIN TYPE: double (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: DNA (genomic) (iii) HYPOTHETICAL: NO (iv) ANTI-SENSE: NO (xi) DESCRIPTION OF SEQUENCE: SEQ ID NO: 20: P1501 / 99MX GAGCGGATCC TGCAGGAGGA GACACAGAG CTG 33 (2) INFORMATION FOR SEQ ID NO: 2 1: (i) CHARACTERISTICS OF SECUENC IA: (A) LENGTH: 34 base pairs (B) TYPE: nucleic acid (C) ) TYPE OF CHAIN: double í (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: DNA (genomic) (iii) HYPOTHETICAL: NO (iv) ANTI-SENSE: NO (xi) DESCRIPTION OF THE SEQUENCE: SEQ ID NO: twenty-one : TGTAGAGATC TGGCTAAGTG CGCGTGTTG CCGT 34 (2) INFORMATION FOR SEQ ID NO: 22: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 34 base pairs (B) TYPE: nucleic acid (C) TYPE OF CHAIN: double (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: DNA (genomic) (iii) HYPOTHETICAL: NO (iv) ANTI-SENSE: NO (xi) DESCRIPTION OF SEQUENCE: SEQ ID NO: 22: TGTACAGATC TCACCATGGC TGTGCCTGC AAGC 34 (2) INFORMATION FOR SEQ ID NO: 23: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 387 base pairs (B) TYPE: nucleic acid (C) CHAIN TYPE: double (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: DNA (genomic) (iii) HYPOTHETICAL: NO. { iv) ANTI-SENSE: NO (xi) DESCRIPTION OF SEQUENCE: SEQ ID NO: 23: GAATTCCGAG TGGTTACTAT TCCATCACCA TTCTAGCCTG TACACAGAAA GTCAAGATGG 60 ACGAATCGCT CGACTTCGCT CGCGATTCGT CGAAGGCGGG GGGCCGGAGG CCCCCCGGTG 120 GCCCCCCTCC AACGAGTGGA GCACGTACAG GGGGGTACG? CATCCGTACA GGGGGGTACG 180 P1501 / 99MX TCATCCGTAC AGGGGGGTAC GTCACAAAGA GGCGTTCCCG TACAGGGGGG TACGTCACGC 240 GTACAGGGGG GTACGTCACA GCCAATCAAA AGCTGCCACG TTGCGAAAGT GACGTTTCGA 300 AAATGGGCGG CGCAAGCCTC TCTATATATT GAGCGCACAT ACCGGTCGGC AGTAGGTATA 360 387 CGCAAGGCGG TCCGGGAGGA TGGATCC (2) INFORMATION FOR SEQ ID NO: 24: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 26 base pairs (B) TYPE: nucleic acid (C) CHAIN TYPE: double (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: DNA (genomic) (iii) HYPOTHETICAL: NO (iv) ANTI-SENSE: NO (xi) DESCRIPTION OF SEQUENCE: SEQ ID NO: 24: ATCGAATTCC GAGTGGTTAC TATTCC 26 (2) INFORMATION FOR SEQ ID NO: 25: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 26 base pairs (B) TYPE: nucleic acid (C) TYPE OF CHAIN: double (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: DNA (genomic) (iii) HYPOTHETICAL: NO (iv) ANTI-SENSE: NO (xi) DESCRIPTION OF SEQUENCE: SEQ ID NO: 25: CGTGGATCCA TCTTACAGTC TTATAC 26 (2) INFORMATION FOR SEQ ID NO: 26: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 29 base pairs (B) TYPE: nucleic acid (C) TYPE OF CHAIN: double (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: DNA. { genomic) (iii) HYPOTHETICAL: NO (iv) ANTI-SENSE: NO (xi) DESCRIPTION OF SEQUENCE: SEQ ID NO: 26 P1S01 / 99MX GTTCGGATCC ATCCTCCCGG ACCGCCTTG 29 (2) INFORMATION FOR SEQ ID NO: 27: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 19 base pairs (B) TYPE: nucleic acid (C) TYPE OF CHAIN: double í (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: DNA (genomic) (iii) HYPOTHETICAL: NO (iv) ANTI-SENSE: NO (xi) DESCRIPTION OF SEQUENCE: SEQ ID NO: 27: GCGGAAGAGC GCCAATACG 19 (2) INFORMATION FOR SEQ ID NO: 28: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 30 base pairs (B) TYPE: nucleic acid (C) CHAIN TYPE: double (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: DNA (genomic) (iii) HYPOTHETICAL: NO (iv) ANTI-SENSE: NO (xi) DESCRIPTION OF SEQUENCE: SEQ ID NO: 28: GTTCGGATCC ATCCACCCG GACCGCCTTG 30 (2) INFORMATION FOR SEQ ID NO: 29: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 33 base pairs (B) TYPE: nucleic acid (C) TYPE OF CHAIN: double (D) ) TOPOLOGY: linear (ii) TYPE OF MOLECULE: DNA (genomic) (iii) HYPOTHETICAL: NO (iv) ANTI-SENSE: NO (xi) DESCRIPTION OF SEQUENCE: SEQ ID NO: 29: TGTACAGATC TCACCATGGC TGTGCCTGCA AGC 33 (2) INFORMATION FOR SEQ ID NO: 30: (i) CHARACTERISTICS OF THE SEQUENCE: P1501 / 99MX (A) LENGTH: 28 base pairs (B) TYPE: nucleic acid (C) CHAIN TYPE: double (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: DNA (genomic) (iii) HYPOTHETICAL: NO f (iv) ANTI-SENSE: NO (xi) DESCRIPTION OF THE SEQUENCE: SEQ ID NO: 30: GGCGAATTCG GCTAAGTGC GCGTGTTG 28 (2) INFORMATION FOR SEQ ID NO: 31: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 594 base pairs (B) TYPE: nucleic acid (C) CHAIN TYPE: double (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: DNA (genomic) (iii) HYPOTHETICAL: NO (iv) ANTI-SENSE: NO (xi) DESCRIPTION OF SEQUENCE: SEQ ID NO: 31: (ix) FEATURES: (A) NAME / KEY : CDS (B) LOCATION: 1..594 (D) OTHER INFORMATION: / product = "Quail Interferon Type 1" ATG GCT GTG CCT GCA AGC CCA CAG CAC CCA CGG GGG TAC GGC ATC CTG CTC 51 MAVPASPQHPRGYGILL 17 CTC ACG CTC CTT CTG AAA GCT CTC GCC ACC ACC GCC ACC GCC TCC GCC TGC 102 LTLLLKALATTATASAC 34 AGC CAC CTT CGC CCC CAC GAC GCC ACC TTC TCT CGC GAC AGC CTC CAG CTC 153 SHLRPHDATFSRDSLQ 51 CTA GGG GAC ATG GCT CCC AGC CCA CCC CAG CTG TGC CCA CAG CAC AGC GCG 204 LGDMAPSPPQLCPQHSA 68 TCG CCT TGC TCC TTC AAC GAC ACC ATC CTG GAC ACC AGC AAC ATC TGG CAA 255 SPCSF N D T I L D T S N I W Q 85 ACT GAC AAA ACC ACC CAC GAC ATT CTT CAG GAC CTC TTC AGT ATC CTC AGC 306 T D K T T H. D I L Q D L F S I L S 102 GGA CCA AGC ACT CCA CCC CAC TGG ATC GAA AGC CAA CGC CAA AGC CTC CTC 357 G P S T P H W I E S Q R Q S L L 119 AGC CAC ATC CAG CGC TAC ACC CAG CAC CTC GAG CAG TGC CTG GAA AAA AAC 08 S H I Q R Y T Q H E C C E L N K 136 AGC GAC ACG CGC TCC CGG ACA CGA CGG CCT CGA AAC CTT CAC CTC ACC ATC 459 S D T R S T R R P L H L T I 153 P1501 / 99MX? CC ??? C? C TTC? CC TCC CTC CCC? CC TTC CTC? CC C T ?? C C? C T? C? CC ,,.

C? C «CCC TCC C? C CTC CTC CTC CTC C ?? CCT CCT «CC TTC CCC ^ V L Q A E H F R R I87 ATC AAC AAC CTC ACA GGC AAC ACG CGC ACT TAG ± N N L T G N T R T í 594 197 (2) INFORMATION FOR SEQ ID NO: 32: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 197 amino acids (B) TYPE: amino acids (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: protein (xi) DESCRIPTION OF THE SEQUENCE: SEQ ID NO: 32 M A V P A S P 0 H P R G Y G I L L 17 T T L L L K A L T T A S C C 34 s H L R P H D A T. F S R D S L Q L 51 L G D M A P S P P Q L C P Q H S A 68 s P C S F N D T I L D T s H I W Q 85 T D T T H D I L Q D L F S I L S 102 G P S T P P H W I E S 0 R Q S L L 119 S K I Q R Y T Q H L E Q * C L E K N 136 s D T R S R T P R N L H L T I 153 s K H F S C L R T F L S D N D Y S 170 D C A W D L V L L Q A R E W F R R 187 I N N L T G N T R T 197 P1501 / 99MX

Claims (20)

1. NOVELTY OF THE INVENTION Having described the present invention, it is considered as a novelty and, therefore, the content of the following CLAIMS is claimed as property: 1. A recombinant herpesvirus of Marek-turkey disease virus chimera comprising a unique long viral genome region of turkey herpesvirus and a unique short viral genome region of Marek's disease virus.
2. 2. The Marek-turkey disease virus chimera recombinant herpes virus according to claim 1, wherein the foreign DNA sequence is inserted into the non-essential region of the viral genome of the Marek-turkey disease virus chimera, and is able to express itself in a host.
3. 3. The Marek-turkey disease virus chimera recombinant herpes virus according to claim 2, wherein the foreign DNA sequence is inserted into an EcoRl # 9 fragment of the unique long region of the viral genome of the virus chimera. Marek's disease-turkey herpesvirus.
4. 4. The Marek-turkey disease virus chimera recombinant herpes virus according to claim 2, wherein the foreign DNA sequence encodes a polypeptide.
5. 5. Chimera recombinant herpesvirus Marek-turkey disease virus P1501 / 99MX according to claim 2, wherein the foreign DNA sequence codes for a cytokine.
6. 6. The Marek-turkey disease virus chimera recombinant herpesvirus according to claim 5, wherein the cytokine is a chicken myelomonocyte growth factor (cMGF), chicken interferon (cIFN) or quaternary interferon type 1 (qlFN) ).
7. 7. The Marek-turkey disease virus chimera recombinant herpesvirus according to claim 2, wherein the foreign DNA sequence encodes an antigenic polypeptide selected from the group consisting of: Marek's disease virus, Nephropathy virus castle, infectious laryngotracheitis virus, infectious bronchitis virus and infectious bursal disease virus.
8. 8. The Marek-turkey disease virus chimera recombinant herpes virus according to claim 2, wherein the polypeptide is β-galactosidase of E. col i.
9. 9. The Marek-turkey disease virus chimera recombinant herpes virus according to claim 7, wherein the antigenic polypeptide is glycoprotein A (gA) of Marek's disease virus, glycoprotein B (gB) of Marek's disease virus or Glycoprotein D (gD) virus P1501 / 99MX Marek's disease.
10. 10. The Marek-turkey disease virus chimera recombinant herpes virus according to claim 7, wherein the antigenic polypeptide is Ne castle disease virus fusion protein or Ne castle disease virus hemagglutinin-neuraminidase.
11. 11. The Marek-turkey disease virus chimera recombinant herpes virus according to claim 7, wherein the antigenic polypeptide is glycoprotein B (gB) of infectious laryngotracheitis virus, glycoprotein I (gl) of infectious laryngotracheitis virus or glycoprotein D (gD).
12. 12. The recombinant herpesvirus of Marek-turkey disease virus chimera according to claim 7, wherein the antigenic polypeptide is spike protein of infectious bronchitis virus or matrix protein of infectious bronchitis virus.
13. 13. The Marek-turkey disease virus chimera recombinant herpes virus according to claim 7, wherein the antigenic polypeptide is VP2 of the infectious bursal disease virus, VP3 of the infectious bursal disease virus or VP4 of the infectious bursal disease virus.
14. 14. Recombinant herpesvirus of Marek-turkey disease virus chimera according to P1501 / 99MX claim 7, wherein the antigenic polypeptide is selected from the group consisting of: avian encephalomyelitis virus, avian reovirus, avian paramyxovirus, avian influenza virus, avian adenovirus, avi4ari4a pustulatory virus, avian coronavirus, avian rotavirus, chicken anemia virus (agent), Salmonella spp. , E. coli, Pas teurella spp. , Bordetella spp. , Eimeria spp. , His tomonas spp. , Tri chomonas spp. , poultry nematodes, cestodes, trematodes and mites / poultry poultry, protozoa poultry.
15. 15. The Marek-turkey disease virus chimera recombinant herpes virus according to claim 2, wherein the foreign DNA sequence is under the control of an upstream endogenous herpesvirus promoter.
16. 16. The Marek-turkey disease virus chimeric recombinant herpes virus according to claim 2, wherein the foreign DNA sequence is under the control of a heterologous upstream promoter.
17. 17. The Marek-turkey disease virus chimera recombinant herpes virus according to claim 16, wherein the promoter is selected from the group consisting of: chicken anemia virus promoter, pseudorabies virus gX promoter, alpha 4 promoter of herpes simplex virus 1, immediate early promoter of P1501 / 99MX human cytomegalovirus, gA promoter of Marek's disease virus, gB promoter, gD promoter of Marek's disease virus, gB promoter of infectious laryngotracheitis, VP8 promoter of bovine herpesvirus-1.1 and gD promoter of infectious laryngotracheitis.
18. 18. A recombinant turkey herpesvirus-Marek's disease virus chimera designated S-HVY-149.
19. 19. A recombinant turkey herpesvirus-Marek's disease virus chimera designated S-HVY-152. 20. A vaccine comprising an effective immunizing amount of the recombinant turkey herpesvirus-Marek's disease virus chimera of claim 2 and a suitable carrier. 21. A multivalent vaccine comprising an effective immunizing amount of the recombinant turkey herpesvirus-Marek's disease virus chimera of claim 2 and a suitable carrier. 22. A method for immunizing a bird against an avian pathogen comprising administering to the bird an effective immunizing dose of the vaccine of claim
20. 20. P1501 / 99MX