Detection of recombinant human lactoferrin and lysozyme produced in a bitransgenic cow

Autores
Kaiser, Germán Gustavo; Mucci, Nicolás Crescencio; González, Vega; Sánchez, Lourdes; Parrón, José A.; Pérez, María D.; Calvo, Miguel; Aller Atucha, Juan Florencio; Hozbor, Federico Andrés; Mutto, Adrián Angel
Año de publicación
2017
Idioma
inglés
Tipo de recurso
artículo
Estado
versión publicada
Descripción
Lactoferrin and lysozyme are 2 glycoproteins with great antimicrobial activity, being part of the nonspecific defensive system of human milk, though their use in commercial products is difficult because human milk is a limited source. Therefore, many investigations have been carried out to produce those proteins in biological systems, such as bacteria, yeasts, or plants. Mammals seem to be more suitable as expression systems for human proteins, however, especially for those that are glycosylated. In the present study, a bicistronic commercial vector containing a goat β-casein promoter and an internal ribosome entry site fragment between the human lactoferrin and human lysozyme genes was developed to allow the introduction of both genes into bovine adult fibroblasts in a single transfection. Embryos were obtained by somatic cell nuclear transfer, and, after 6 transferences to recipients, 3 pregnancies and 1 viable bitransgenic calf were obtained. The presence of the vector was confirmed by fluorescent in situ hybridization of skin cells. At 13 mo of life and after artificial induction of lactation, both recombinant proteins were found in the colostrum and milk of the bitransgenic calf. Human lactoferrin concentration in the colostrum was 0.0098 mg/mL and that in milk was 0.011 mg/mL, whereas human lysozyme concentration in the colostrum was 0.0022 mg/mL and that in milk was 0.0024 mg/mL. The molar concentration of both human proteins revealed no differences in protein production of the internal ribosome entry site upstream and downstream protein. The enzymatic activity of lysozyme in the transgenic milk was comparable to that of human milk, being 6 and 10 times higher than that of bovine lysozyme present in milk. This work represents an important step to obtain multiple proteins or enhance single protein production by using animal pharming and fewer regulatory and antibiotic-resistant foreign sequences, allowing the design of humanized milk with added biological value for newborn nutrition and development. Transgenic animals can offer a unique opportunity to dairy industry, providing starting materials suitable to develop specific products with high added value.
Fil: Kaiser, Germán Gustavo. Instituto Nacional de Tecnología Agropecuaria. Centro Regional Buenos Aires Sur. Estación Experimental Agropecuaria Balcarce. Laboratorio de Biotecnología de la Reproduccion; Argentina
Fil: Mucci, Nicolás Crescencio. Instituto Nacional de Tecnología Agropecuaria. Centro Regional Buenos Aires Sur. Estación Experimental Agropecuaria Balcarce. Laboratorio de Biotecnología de la Reproduccion; Argentina
Fil: González, Vega. Universidad de Zaragoza; España
Fil: Sánchez, Lourdes. Universidad de Zaragoza; España
Fil: Parrón, José A.. Universidad de Zaragoza; España
Fil: Pérez, María D.. Universidad de Zaragoza; España
Fil: Calvo, Miguel. Universidad de Zaragoza; España
Fil: Aller Atucha, Juan Florencio. Instituto Nacional de Tecnología Agropecuaria. Centro Regional Buenos Aires Sur. Estación Experimental Agropecuaria Balcarce. Laboratorio de Biotecnología de la Reproduccion; Argentina
Fil: Hozbor, Federico Andrés. Instituto Nacional de Tecnología Agropecuaria. Centro Regional Buenos Aires Sur. Estación Experimental Agropecuaria Balcarce. Laboratorio de Biotecnología de la Reproduccion; Argentina
Fil: Mutto, Adrián Angel. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Biotecnológicas. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas; Argentina
Materia
HUMAN LACTOFERRIN
RECOMBINANT
BI TRANSGENIC COW
ELISA
Nivel de accesibilidad
acceso abierto
Condiciones de uso
https://creativecommons.org/licenses/by-nc-sa/2.5/ar/
Repositorio
CONICET Digital (CONICET)
Institución
Consejo Nacional de Investigaciones Científicas y Técnicas
OAI Identificador
oai:ri.conicet.gov.ar:11336/102766
Acceder
id CONICETDig_f91e305bcce83fa3dd4e94341201aa25
oai_identifier_str oai:ri.conicet.gov.ar:11336/102766
network_acronym_str CONICETDig
repository_id_str 3498
network_name_str CONICET Digital (CONICET)
spelling Detection of recombinant human lactoferrin and lysozyme produced in a bitransgenic cowKaiser, Germán GustavoMucci, Nicolás CrescencioGonzález, VegaSánchez, LourdesParrón, José A.Pérez, María D.Calvo, MiguelAller Atucha, Juan FlorencioHozbor, Federico AndrésMutto, Adrián AngelHUMAN LACTOFERRINRECOMBINANTBI TRANSGENIC COWELISAhttps://purl.org/becyt/ford/4.4https://purl.org/becyt/ford/4Lactoferrin and lysozyme are 2 glycoproteins with great antimicrobial activity, being part of the nonspecific defensive system of human milk, though their use in commercial products is difficult because human milk is a limited source. Therefore, many investigations have been carried out to produce those proteins in biological systems, such as bacteria, yeasts, or plants. Mammals seem to be more suitable as expression systems for human proteins, however, especially for those that are glycosylated. In the present study, a bicistronic commercial vector containing a goat β-casein promoter and an internal ribosome entry site fragment between the human lactoferrin and human lysozyme genes was developed to allow the introduction of both genes into bovine adult fibroblasts in a single transfection. Embryos were obtained by somatic cell nuclear transfer, and, after 6 transferences to recipients, 3 pregnancies and 1 viable bitransgenic calf were obtained. The presence of the vector was confirmed by fluorescent in situ hybridization of skin cells. At 13 mo of life and after artificial induction of lactation, both recombinant proteins were found in the colostrum and milk of the bitransgenic calf. Human lactoferrin concentration in the colostrum was 0.0098 mg/mL and that in milk was 0.011 mg/mL, whereas human lysozyme concentration in the colostrum was 0.0022 mg/mL and that in milk was 0.0024 mg/mL. The molar concentration of both human proteins revealed no differences in protein production of the internal ribosome entry site upstream and downstream protein. The enzymatic activity of lysozyme in the transgenic milk was comparable to that of human milk, being 6 and 10 times higher than that of bovine lysozyme present in milk. This work represents an important step to obtain multiple proteins or enhance single protein production by using animal pharming and fewer regulatory and antibiotic-resistant foreign sequences, allowing the design of humanized milk with added biological value for newborn nutrition and development. Transgenic animals can offer a unique opportunity to dairy industry, providing starting materials suitable to develop specific products with high added value.Fil: Kaiser, Germán Gustavo. Instituto Nacional de Tecnología Agropecuaria. Centro Regional Buenos Aires Sur. Estación Experimental Agropecuaria Balcarce. Laboratorio de Biotecnología de la Reproduccion; ArgentinaFil: Mucci, Nicolás Crescencio. Instituto Nacional de Tecnología Agropecuaria. Centro Regional Buenos Aires Sur. Estación Experimental Agropecuaria Balcarce. Laboratorio de Biotecnología de la Reproduccion; ArgentinaFil: González, Vega. Universidad de Zaragoza; EspañaFil: Sánchez, Lourdes. Universidad de Zaragoza; EspañaFil: Parrón, José A.. Universidad de Zaragoza; EspañaFil: Pérez, María D.. Universidad de Zaragoza; EspañaFil: Calvo, Miguel. Universidad de Zaragoza; EspañaFil: Aller Atucha, Juan Florencio. Instituto Nacional de Tecnología Agropecuaria. Centro Regional Buenos Aires Sur. Estación Experimental Agropecuaria Balcarce. Laboratorio de Biotecnología de la Reproduccion; ArgentinaFil: Hozbor, Federico Andrés. Instituto Nacional de Tecnología Agropecuaria. Centro Regional Buenos Aires Sur. Estación Experimental Agropecuaria Balcarce. Laboratorio de Biotecnología de la Reproduccion; ArgentinaFil: Mutto, Adrián Angel. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Biotecnológicas. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas; ArgentinaAmerican Dairy Science Association2017-03info:eu-repo/semantics/articleinfo:eu-repo/semantics/publishedVersionhttp://purl.org/coar/resource_type/c_6501info:ar-repo/semantics/articuloapplication/pdfapplication/pdfhttp://hdl.handle.net/11336/102766Kaiser, Germán Gustavo; Mucci, Nicolás Crescencio; González, Vega; Sánchez, Lourdes; Parrón, José A.; et al.; Detection of recombinant human lactoferrin and lysozyme produced in a bitransgenic cow; American Dairy Science Association; Journal of Dairy Science; 100; 3; 3-2017; 1605-16170022-0302CONICET DigitalCONICETenginfo:eu-repo/semantics/altIdentifier/doi/10.3168/jds.2016-11173info:eu-repo/semantics/altIdentifier/url/https://www.journalofdairyscience.org/article/S0022-0302(17)30035-8/abstractinfo:eu-repo/semantics/openAccesshttps://creativecommons.org/licenses/by-nc-sa/2.5/ar/reponame:CONICET Digital (CONICET)instname:Consejo Nacional de Investigaciones Científicas y Técnicas2025-09-29T09:51:55Zoai:ri.conicet.gov.ar:11336/102766instacron:CONICETInstitucionalhttp://ri.conicet.gov.ar/Organismo científico-tecnológicoNo correspondehttp://ri.conicet.gov.ar/oai/requestdasensio@conicet.gov.ar; lcarlino@conicet.gov.arArgentinaNo correspondeNo correspondeNo correspondeopendoar:34982025-09-29 09:51:55.646CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicasfalse
dc.title.none.fl_str_mv Detection of recombinant human lactoferrin and lysozyme produced in a bitransgenic cow
title Detection of recombinant human lactoferrin and lysozyme produced in a bitransgenic cow
spellingShingle Detection of recombinant human lactoferrin and lysozyme produced in a bitransgenic cow
Kaiser, Germán Gustavo
HUMAN LACTOFERRIN
RECOMBINANT
BI TRANSGENIC COW
ELISA
title_short Detection of recombinant human lactoferrin and lysozyme produced in a bitransgenic cow
title_full Detection of recombinant human lactoferrin and lysozyme produced in a bitransgenic cow
title_fullStr Detection of recombinant human lactoferrin and lysozyme produced in a bitransgenic cow
title_full_unstemmed Detection of recombinant human lactoferrin and lysozyme produced in a bitransgenic cow
title_sort Detection of recombinant human lactoferrin and lysozyme produced in a bitransgenic cow
dc.creator.none.fl_str_mv Kaiser, Germán Gustavo
Mucci, Nicolás Crescencio
González, Vega
Sánchez, Lourdes
Parrón, José A.
Pérez, María D.
Calvo, Miguel
Aller Atucha, Juan Florencio
Hozbor, Federico Andrés
Mutto, Adrián Angel
author Kaiser, Germán Gustavo
author_facet Kaiser, Germán Gustavo
Mucci, Nicolás Crescencio
González, Vega
Sánchez, Lourdes
Parrón, José A.
Pérez, María D.
Calvo, Miguel
Aller Atucha, Juan Florencio
Hozbor, Federico Andrés
Mutto, Adrián Angel
author_role author
author2 Mucci, Nicolás Crescencio
González, Vega
Sánchez, Lourdes
Parrón, José A.
Pérez, María D.
Calvo, Miguel
Aller Atucha, Juan Florencio
Hozbor, Federico Andrés
Mutto, Adrián Angel
author2_role author
author
author
author
author
author
author
author
author
dc.subject.none.fl_str_mv HUMAN LACTOFERRIN
RECOMBINANT
BI TRANSGENIC COW
ELISA
topic HUMAN LACTOFERRIN
RECOMBINANT
BI TRANSGENIC COW
ELISA
purl_subject.fl_str_mv https://purl.org/becyt/ford/4.4
https://purl.org/becyt/ford/4
dc.description.none.fl_txt_mv Lactoferrin and lysozyme are 2 glycoproteins with great antimicrobial activity, being part of the nonspecific defensive system of human milk, though their use in commercial products is difficult because human milk is a limited source. Therefore, many investigations have been carried out to produce those proteins in biological systems, such as bacteria, yeasts, or plants. Mammals seem to be more suitable as expression systems for human proteins, however, especially for those that are glycosylated. In the present study, a bicistronic commercial vector containing a goat β-casein promoter and an internal ribosome entry site fragment between the human lactoferrin and human lysozyme genes was developed to allow the introduction of both genes into bovine adult fibroblasts in a single transfection. Embryos were obtained by somatic cell nuclear transfer, and, after 6 transferences to recipients, 3 pregnancies and 1 viable bitransgenic calf were obtained. The presence of the vector was confirmed by fluorescent in situ hybridization of skin cells. At 13 mo of life and after artificial induction of lactation, both recombinant proteins were found in the colostrum and milk of the bitransgenic calf. Human lactoferrin concentration in the colostrum was 0.0098 mg/mL and that in milk was 0.011 mg/mL, whereas human lysozyme concentration in the colostrum was 0.0022 mg/mL and that in milk was 0.0024 mg/mL. The molar concentration of both human proteins revealed no differences in protein production of the internal ribosome entry site upstream and downstream protein. The enzymatic activity of lysozyme in the transgenic milk was comparable to that of human milk, being 6 and 10 times higher than that of bovine lysozyme present in milk. This work represents an important step to obtain multiple proteins or enhance single protein production by using animal pharming and fewer regulatory and antibiotic-resistant foreign sequences, allowing the design of humanized milk with added biological value for newborn nutrition and development. Transgenic animals can offer a unique opportunity to dairy industry, providing starting materials suitable to develop specific products with high added value.
Fil: Kaiser, Germán Gustavo. Instituto Nacional de Tecnología Agropecuaria. Centro Regional Buenos Aires Sur. Estación Experimental Agropecuaria Balcarce. Laboratorio de Biotecnología de la Reproduccion; Argentina
Fil: Mucci, Nicolás Crescencio. Instituto Nacional de Tecnología Agropecuaria. Centro Regional Buenos Aires Sur. Estación Experimental Agropecuaria Balcarce. Laboratorio de Biotecnología de la Reproduccion; Argentina
Fil: González, Vega. Universidad de Zaragoza; España
Fil: Sánchez, Lourdes. Universidad de Zaragoza; España
Fil: Parrón, José A.. Universidad de Zaragoza; España
Fil: Pérez, María D.. Universidad de Zaragoza; España
Fil: Calvo, Miguel. Universidad de Zaragoza; España
Fil: Aller Atucha, Juan Florencio. Instituto Nacional de Tecnología Agropecuaria. Centro Regional Buenos Aires Sur. Estación Experimental Agropecuaria Balcarce. Laboratorio de Biotecnología de la Reproduccion; Argentina
Fil: Hozbor, Federico Andrés. Instituto Nacional de Tecnología Agropecuaria. Centro Regional Buenos Aires Sur. Estación Experimental Agropecuaria Balcarce. Laboratorio de Biotecnología de la Reproduccion; Argentina
Fil: Mutto, Adrián Angel. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Biotecnológicas. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas; Argentina
description Lactoferrin and lysozyme are 2 glycoproteins with great antimicrobial activity, being part of the nonspecific defensive system of human milk, though their use in commercial products is difficult because human milk is a limited source. Therefore, many investigations have been carried out to produce those proteins in biological systems, such as bacteria, yeasts, or plants. Mammals seem to be more suitable as expression systems for human proteins, however, especially for those that are glycosylated. In the present study, a bicistronic commercial vector containing a goat β-casein promoter and an internal ribosome entry site fragment between the human lactoferrin and human lysozyme genes was developed to allow the introduction of both genes into bovine adult fibroblasts in a single transfection. Embryos were obtained by somatic cell nuclear transfer, and, after 6 transferences to recipients, 3 pregnancies and 1 viable bitransgenic calf were obtained. The presence of the vector was confirmed by fluorescent in situ hybridization of skin cells. At 13 mo of life and after artificial induction of lactation, both recombinant proteins were found in the colostrum and milk of the bitransgenic calf. Human lactoferrin concentration in the colostrum was 0.0098 mg/mL and that in milk was 0.011 mg/mL, whereas human lysozyme concentration in the colostrum was 0.0022 mg/mL and that in milk was 0.0024 mg/mL. The molar concentration of both human proteins revealed no differences in protein production of the internal ribosome entry site upstream and downstream protein. The enzymatic activity of lysozyme in the transgenic milk was comparable to that of human milk, being 6 and 10 times higher than that of bovine lysozyme present in milk. This work represents an important step to obtain multiple proteins or enhance single protein production by using animal pharming and fewer regulatory and antibiotic-resistant foreign sequences, allowing the design of humanized milk with added biological value for newborn nutrition and development. Transgenic animals can offer a unique opportunity to dairy industry, providing starting materials suitable to develop specific products with high added value.
publishDate 2017
dc.date.none.fl_str_mv 2017-03
dc.type.none.fl_str_mv info:eu-repo/semantics/article
info:eu-repo/semantics/publishedVersion
http://purl.org/coar/resource_type/c_6501
info:ar-repo/semantics/articulo
format article
status_str publishedVersion
dc.identifier.none.fl_str_mv http://hdl.handle.net/11336/102766
Kaiser, Germán Gustavo; Mucci, Nicolás Crescencio; González, Vega; Sánchez, Lourdes; Parrón, José A.; et al.; Detection of recombinant human lactoferrin and lysozyme produced in a bitransgenic cow; American Dairy Science Association; Journal of Dairy Science; 100; 3; 3-2017; 1605-1617
0022-0302
CONICET Digital
CONICET
url http://hdl.handle.net/11336/102766
identifier_str_mv Kaiser, Germán Gustavo; Mucci, Nicolás Crescencio; González, Vega; Sánchez, Lourdes; Parrón, José A.; et al.; Detection of recombinant human lactoferrin and lysozyme produced in a bitransgenic cow; American Dairy Science Association; Journal of Dairy Science; 100; 3; 3-2017; 1605-1617
0022-0302
CONICET Digital
CONICET
dc.language.none.fl_str_mv eng
language eng
dc.relation.none.fl_str_mv info:eu-repo/semantics/altIdentifier/doi/10.3168/jds.2016-11173
info:eu-repo/semantics/altIdentifier/url/https://www.journalofdairyscience.org/article/S0022-0302(17)30035-8/abstract
dc.rights.none.fl_str_mv info:eu-repo/semantics/openAccess
https://creativecommons.org/licenses/by-nc-sa/2.5/ar/
eu_rights_str_mv openAccess
rights_invalid_str_mv https://creativecommons.org/licenses/by-nc-sa/2.5/ar/
dc.format.none.fl_str_mv application/pdf
application/pdf
dc.publisher.none.fl_str_mv American Dairy Science Association
publisher.none.fl_str_mv American Dairy Science Association
dc.source.none.fl_str_mv reponame:CONICET Digital (CONICET)
instname:Consejo Nacional de Investigaciones Científicas y Técnicas
reponame_str CONICET Digital (CONICET)
collection CONICET Digital (CONICET)
instname_str Consejo Nacional de Investigaciones Científicas y Técnicas
repository.name.fl_str_mv CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicas
repository.mail.fl_str_mv dasensio@conicet.gov.ar; lcarlino@conicet.gov.ar
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score 13.070432

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