ES2525796T3 - Nuevos microorganismos para la producción de 1,2-propanodiol obtenidos mediante una combinación de evolución y diseño racional - Google Patents
Nuevos microorganismos para la producción de 1,2-propanodiol obtenidos mediante una combinación de evolución y diseño racional Download PDFInfo
- Publication number
- ES2525796T3 ES2525796T3 ES08714246.9T ES08714246T ES2525796T3 ES 2525796 T3 ES2525796 T3 ES 2525796T3 ES 08714246 T ES08714246 T ES 08714246T ES 2525796 T3 ES2525796 T3 ES 2525796T3
- Authority
- ES
- Spain
- Prior art keywords
- strain
- gene
- evolution
- production
- propanediol
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Active
Links
Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/20—Bacteria; Culture media therefor
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
- C12N15/01—Preparation of mutants without inserting foreign genetic material therein; Screening processes therefor
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
- C12N15/09—Recombinant DNA-technology
- C12N15/11—DNA or RNA fragments; Modified forms thereof; Non-coding nucleic acids having a biological activity
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
- C12N9/90—Isomerases (5.)
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P7/00—Preparation of oxygen-containing organic compounds
- C12P7/02—Preparation of oxygen-containing organic compounds containing a hydroxy group
- C12P7/04—Preparation of oxygen-containing organic compounds containing a hydroxy group acyclic
- C12P7/18—Preparation of oxygen-containing organic compounds containing a hydroxy group acyclic polyhydric
-
- C—CHEMISTRY; METALLURGY
- C40—COMBINATORIAL TECHNOLOGY
- C40B—COMBINATORIAL CHEMISTRY; LIBRARIES, e.g. CHEMICAL LIBRARIES
- C40B10/00—Directed molecular evolution of macromolecules, e.g. RNA, DNA or proteins
Landscapes
- Life Sciences & Earth Sciences (AREA)
- Health & Medical Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Engineering & Computer Science (AREA)
- Genetics & Genomics (AREA)
- Zoology (AREA)
- Wood Science & Technology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Biotechnology (AREA)
- General Engineering & Computer Science (AREA)
- General Health & Medical Sciences (AREA)
- Biochemistry (AREA)
- Molecular Biology (AREA)
- Biomedical Technology (AREA)
- Microbiology (AREA)
- Medicinal Chemistry (AREA)
- Chemical Kinetics & Catalysis (AREA)
- General Chemical & Material Sciences (AREA)
- Physics & Mathematics (AREA)
- Biophysics (AREA)
- Plant Pathology (AREA)
- Virology (AREA)
- Tropical Medicine & Parasitology (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
- Preparation Of Compounds By Using Micro-Organisms (AREA)
Abstract
Método para la preparación de una cepa de microorganismo evolucionada para la producción de 1,2-propanodiol a partir de una fuente de carbono, comprendiendo dicho método: - hacer crecer una cepa inicial bajo presión selectiva en un medio de crecimiento apropiado, comprendiendo dicha cepa bacteriana una atenuación de la expresión del gen tpiA y una atenuación de la expresión de al menos uno de los genes involucrados en la conversión de metilglioxal en lactato, con el objetivo de promover la evolución de dicha cepa inicial, - seleccionar y aislar la cepa evolucionada que presenta una tasa de producción de 1,2-propanodiol incrementada, - reconstruir un gen tpiA funcional en la cepa evolucionada.
Description
E08714246
11-12-2014
El cassette de resistencia a cloramfenicol se eliminó de la cepa E. coli MG1655 lpd*, ∆tpiA, ∆pflAB, ∆adhE, ldhA::km, ∆gloA, ∆aldA, ∆aldB, ∆edd::cm (Ver WO2005073364) de acuerdo con el Protocolo 1.
Protocolo 1: Eliminación de cassettes de resistencia
5 Los cassettes de resistencia a cloramfenicol y/o a kanamicina se eliminaron de acuerdo con la siguiente técnica. El plásmido pCP20 portador de la FLP recombinasa que actúa en los sitios FRT de los cassettes de resistencia a cloramfenicol y/o a kanamicina se introdujo en la cepa mediante electroporación. Después de un cultivo seriado a 41ºC, la pérdida de los cassettes de resistencia a antibiótico se verificó mediante un análisis PCR con los
10 oligonucleótidos detallados en la Tabla 1.
La presencia de modificaciones previamente construidas en la cepa se verificó utilizando los oligonucleótidos detallados en la Tabla 1.
15 La cepa obtenida se nombró E. coli MG1655 lpd*, ∆tpiA, ∆pflAB, ∆adhE, ldhA::km, ∆gloA, ∆aldA, ∆aldB, ∆edd.
Tabla 1: Oligonucleótidos utilizados para la verificación de la inserción de un cassette de resistencia o la pérdida de un cassette de resistencia
- Nombre de la región
- Nombre de los oligos SEQ ID Homología con la región cromosómica
- gen tpiA
- cdh N°1 Ver WO2005073364
- (deleción)
- YIIQ N°2
- gen pflAB
- pflABF N°3 Ver WO2005073364
- pflABR
- N°4
- gen adhE
- ychGf N°5 Ver WO2005073364
- adhECr
- N°6
- gen ldhA
- hsIJC N°7 Ver WO2005073364
- (inserción cassette)
- ldhAC2 N°8
- gen gloA
- NemACd N°9 Ver WO2005073364
- Rnt Cr
- N°10
- gen aldA
- Ydc F C f N°11 Ver WO2005073364
- gapCCr
- N°12
- gen aldB
- aldB C f N°13 Ver WO2005073364
- YiaYCr
- N°14
- gen edd
- Edad N°15 Ver WO2005073364
- Zwf r
- N°16
- gen ldhA gene
- ldhAF N°17 1439724 hasta 1439743
- (deleción)
- ldhAR N°18 1441029 hasta 1441007
- gen arcA
- arcAF N°19 4638292 hasta 4638273
- arcAR
- N°20 4636854 hasta 4636874
- gen ndh
- ndhF N°21 1164722 hasta 1164742
- ndhR
- N°22 1167197 hasta 1167177
- gen tpiA
- YIIQ N°2 4109599 hasta 4109580
- (reconstrucción)
- tpiA R N°23 4108953 hasta 4108973
- promotor gapA
- yeaAF N°24 1860259-1860287
- (Ptrc16-gapA)
- gapAR N°25 1861068-1861040
- gen pykA
- pykAF N°26 1935338 hasta 1935360
- pykAR
- N°27 1937425 hasta 1937401
- gen pykF
- pykFF N°28 1753371 hasta 1753392
11 E08714246
11-12-2014
- Nombre de la región
- Nombre de los oligos SEQ ID Homología con la región cromosómica
- pykFR
- N°29 1755518 hasta 1755495
- genes ackA-pta
- B2295 N°30 2410900 hasta 2410919
- YfcCR
- N°31 2415164 hasta 2415145
- genes poxB
- poxBF N°32 908475 hasta 908495
- poxBR
- N°33 910375 hasta 910352
5 Para eliminar el cassette de resistencia a kanamicina y para desactivar el gen ldhA, el cassette de resistencia a cloramfenicol se insertó en el gen ldhA delecionando la mayor parte del gen en cuestión de acuerdo con el Protocolo
2.
Protocolo 2: Introducción de un producto de PCR para recombinación y selección de los recombinantes
10 Los oligonucleótidos elegidos para la sustitución de un gen o de una región intergénica y detallados en la Tabla 2 se utilizaron para amplificar ya sea el cassette de resistencia a cloramfenicol a partir del plásmido pKD3 o el cassette de resistencia a kanamicina a partir del plásmido pKD4 (Datsenko, K.A. & Wanner, B.L. (2000)). El producto de PCR obtenido se introdujo posteriormente mediante electroporación en la cepa receptora portadora del plásmido pKD46
15 en el que el sistema λ Red (y, β, exo) expresado favorece enormemente la recombinación homóloga. Los transformantes resistentes a antibiótico fueron entonces seleccionados y la inserción del cassette de resistencia fue verificada por análisis PCR utilizando los oligonucleótidos apropiados que se detallan en la Tabla 1.
Las demás modificaciones de la cepa fueron verificadas con los oligonucleótidos detallados en la Tabla 1. 20 La cepa resultante se nombró E. coli MG1655 lpd*, ∆ldhA::cm, ∆tpiA, ∆pflAB, ∆adhE. ∆gloA, ∆aldA, ∆aldB, ∆edd.
Tabla 2: Oligonucleótidos utilizados para la sustitución de una región cromosómica por recombinación con un producto de PCR en la cepa E. coli MG1655 25
- Nombre de la región
- Nombre de los oligos SEQ ID Homología con la región cromosómica
- gen ldhA
- DldhAF N°34 1440865-1440786
- DldhAR
- N°35 1439878-1439958
- gen arcA
- DarcAF N°36 4637868-4637791
- DarcAR
- N°37 4637167-4637245
- gen ndh
- DndhF N°38 1165071-1165149
- DndhR
- N°39 1166607-1166528
- gen tpiA
- tpiA::kmF N°40 4109264 -4109195
- (reconstrucción)
- tpiA::kmR N°41 4109109 -4109193
- promotor gapA
- Ptrc-gapAF N°42 1860478-1860536
- (Ptrc 16-gapA)
- Ptrc-gapAR N°43 1860762-1860800
- gen pykA
- DpykAF N°44 1935756-1935836
- DpykAR
- N°45 1937055-1937135
- gen pykF
- DpykFF N°46 1753689-1753766
- DpykFR
- N°47 1755129-1755051
- genes ackA-pta
- DackAF N°48 2411494-2411573
- DptaR
- N°49 2414906-2414830
- genes poxB
- DpoxBF N°50 908557-908635
- DpoxBR
- N°51 910262-910180
12
Claims (1)
-
imagen1
Applications Claiming Priority (3)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
IB2007001680 | 2007-03-23 | ||
WOPCT/IB2007/001680 | 2007-03-23 | ||
PCT/EP2008/053445 WO2008116852A1 (en) | 2007-03-23 | 2008-03-21 | New micro-organisms for the production of 1,2-propanediol obtained by a combination of evolution and rational design. |
Publications (1)
Publication Number | Publication Date |
---|---|
ES2525796T3 true ES2525796T3 (es) | 2014-12-30 |
Family
ID=38814484
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
ES08714246.9T Active ES2525796T3 (es) | 2007-03-23 | 2008-03-21 | Nuevos microorganismos para la producción de 1,2-propanodiol obtenidos mediante una combinación de evolución y diseño racional |
Country Status (14)
Country | Link |
---|---|
US (1) | US8298807B2 (es) |
EP (1) | EP2109681B1 (es) |
JP (1) | JP5570821B2 (es) |
KR (1) | KR101528943B1 (es) |
CN (1) | CN101641446A (es) |
AR (1) | AR066196A1 (es) |
BR (1) | BRPI0808972B1 (es) |
CA (1) | CA2679989C (es) |
DK (1) | DK2109681T3 (es) |
ES (1) | ES2525796T3 (es) |
IL (1) | IL200705A0 (es) |
MX (1) | MX2009010229A (es) |
TW (1) | TW200904984A (es) |
WO (1) | WO2008116852A1 (es) |
Families Citing this family (22)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US8647642B2 (en) | 2008-09-18 | 2014-02-11 | Aviex Technologies, Llc | Live bacterial vaccines resistant to carbon dioxide (CO2), acidic PH and/or osmolarity for viral infection prophylaxis or treatment |
BRPI0823256B8 (pt) * | 2008-11-07 | 2020-05-26 | Metabolic Explorer Sa | uso de sacarose como substrato para produção fermentativa de 1,2-propanodiol |
EP2233562A1 (en) | 2009-03-24 | 2010-09-29 | Metabolic Explorer | Method for producing high amount of glycolic acid by fermentation |
MY187676A (en) | 2009-06-04 | 2021-10-08 | Genomatica Inc | Microorganisms for the production of 1,4-butanediol and related methods |
WO2011012702A1 (en) | 2009-07-30 | 2011-02-03 | Metabolic Explorer | Mutant glycerol dehydrogenase (glydh) for the production of a biochemical by fermentation |
BR112012002153A2 (pt) | 2009-07-30 | 2015-09-15 | Metabolic Explorer Sa | metilglioxal sintase mutante (mgs) para a produção de material bioquímico por fermentação |
WO2011012697A2 (en) | 2009-07-30 | 2011-02-03 | Metabolic Explorer | Mutant yqhd enzyme for the production of a biochemical by fermentation |
EP2532751A1 (en) * | 2011-06-10 | 2012-12-12 | Metabolic Explorer | Use of inducible promoters in the fermentative production of 1,2-propanediol |
EP2721167A1 (en) | 2011-06-15 | 2014-04-23 | B.R.A.I.N. Biotechnology Research And Information Network AG | New means and methods for producing propanediol |
EP3050970B1 (en) | 2015-01-28 | 2019-09-18 | Metabolic Explorer | Modified microorganism for optimized production of 1,4-butanediol |
US10415062B2 (en) | 2015-04-07 | 2019-09-17 | Metabolic Explorer | Modified microorganism for the optimized production of 2,4-dihydroxybutyrate |
CN107771214B (zh) | 2015-04-07 | 2022-01-18 | 代谢探索者公司 | 用于具有增加的2,4-二羟基丁酸外排物的优化的2,4-二羟基丁酸产生的修饰的微生物 |
PL3331998T3 (pl) | 2015-08-07 | 2020-12-28 | Evonik Operations Gmbh | Wytwarzanie przez fermentację l-metioniny zależne od tiokarboksylanu białka |
EP3347478B1 (en) | 2015-09-10 | 2022-09-21 | Metabolic Explorer | New lactaldehyde reductases for the production of 1,2-propanediol |
EP3365427B1 (en) | 2015-10-23 | 2020-08-19 | Metabolic Explorer | Microorganism modified for the assimilation of levulinic acid |
MX2018006434A (es) | 2015-11-27 | 2018-09-28 | Evonik Degussa Gmbh | Metodo para producir l-metionina. |
KR102430878B1 (ko) | 2016-07-08 | 2022-08-09 | 메타볼릭 익스플로러 | 당 포스포트랜스퍼라제 시스템 (pts)을 코딩하는 유전자를 포함하는 미생물에 의한 관심 분자의 발효적 생산을 위한 방법 |
US11129906B1 (en) | 2016-12-07 | 2021-09-28 | David Gordon Bermudes | Chimeric protein toxins for expression by therapeutic bacteria |
US11180535B1 (en) | 2016-12-07 | 2021-11-23 | David Gordon Bermudes | Saccharide binding, tumor penetration, and cytotoxic antitumor chimeric peptides from therapeutic bacteria |
EP3342873A1 (en) | 2016-12-29 | 2018-07-04 | Metabolic Explorer | Conversion of methylglyoxal into hydroxyacetone using enzymes and applications thereof |
EP3470512A1 (en) | 2017-10-10 | 2019-04-17 | Metabolic Explorer | Mutant phosphoserine aminotransferase for the conversion of homoserine into 4-hydroxy-2-ketobutyrate |
KR102245274B1 (ko) | 2019-01-28 | 2021-04-27 | 재단법인 지능형 바이오 시스템 설계 및 합성 연구단 | 최소 유전체를 갖는 신규 미생물 및 이의 제조 방법 |
Family Cites Families (4)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US6087140A (en) | 1997-02-19 | 2000-07-11 | Wisconsin Alumni Research Foundation | Microbial production of 1,2-propanediol from sugar |
ES2356526T3 (es) * | 2002-10-04 | 2011-04-08 | E.I. Du Pont De Nemours And Company | Procedimiento para la producción biológica de 1,3-propanodiol con elevado rendimiento. |
JP2006517796A (ja) | 2003-02-18 | 2006-08-03 | メタボリック エクスプローラー | 代謝経路の生成または改変を可能とする進化した微生物の生産方法 |
FR2864967B1 (fr) | 2004-01-12 | 2006-05-19 | Metabolic Explorer Sa | Microorganisme evolue pour la production de 1,2-propanediol |
-
2008
- 2008-03-21 WO PCT/EP2008/053445 patent/WO2008116852A1/en active Application Filing
- 2008-03-21 ES ES08714246.9T patent/ES2525796T3/es active Active
- 2008-03-21 MX MX2009010229A patent/MX2009010229A/es active IP Right Grant
- 2008-03-21 KR KR1020097022123A patent/KR101528943B1/ko active IP Right Grant
- 2008-03-21 CN CN200880009616A patent/CN101641446A/zh active Pending
- 2008-03-21 TW TW097110003A patent/TW200904984A/zh unknown
- 2008-03-21 BR BRPI0808972A patent/BRPI0808972B1/pt not_active IP Right Cessation
- 2008-03-21 JP JP2009554045A patent/JP5570821B2/ja not_active Expired - Fee Related
- 2008-03-21 US US12/532,405 patent/US8298807B2/en active Active
- 2008-03-21 DK DK08714246.9T patent/DK2109681T3/en active
- 2008-03-21 CA CA2679989A patent/CA2679989C/en active Active
- 2008-03-21 EP EP08714246.9A patent/EP2109681B1/en not_active Not-in-force
- 2008-03-25 AR ARP080101213A patent/AR066196A1/es active IP Right Grant
-
2009
- 2009-09-03 IL IL200705A patent/IL200705A0/en active IP Right Grant
Also Published As
Publication number | Publication date |
---|---|
WO2008116852A1 (en) | 2008-10-02 |
TW200904984A (en) | 2009-02-01 |
BRPI0808972B1 (pt) | 2016-11-29 |
CA2679989C (en) | 2015-07-07 |
CN101641446A (zh) | 2010-02-03 |
EP2109681B1 (en) | 2014-10-15 |
DK2109681T3 (en) | 2014-12-08 |
US8298807B2 (en) | 2012-10-30 |
CA2679989A1 (en) | 2008-10-02 |
JP2010521958A (ja) | 2010-07-01 |
IL200705A0 (en) | 2010-05-17 |
US20100285547A1 (en) | 2010-11-11 |
BRPI0808972A2 (pt) | 2014-08-26 |
MX2009010229A (es) | 2009-10-26 |
EP2109681A1 (en) | 2009-10-21 |
JP5570821B2 (ja) | 2014-08-13 |
KR101528943B1 (ko) | 2015-06-15 |
KR20100015810A (ko) | 2010-02-12 |
AR066196A1 (es) | 2009-08-05 |
Similar Documents
Publication | Publication Date | Title |
---|---|---|
ES2525796T3 (es) | Nuevos microorganismos para la producción de 1,2-propanodiol obtenidos mediante una combinación de evolución y diseño racional | |
US20220347271A1 (en) | Sequence-specific antimicrobials by blocking dna repair | |
Vandecraen et al. | The impact of insertion sequences on bacterial genome plasticity and adaptability | |
Doublet et al. | Antibiotic marker modifications of λ Red and FLP helper plasmids, pKD46 and pCP20, for inactivation of chromosomal genes using PCR products in multidrug-resistant strains | |
Thomason et al. | Recombineering: genetic engineering in bacteria using homologous recombination | |
JP6621464B2 (ja) | 欠失突然変異 | |
Ellermeier et al. | Construction of targeted single copy lac fusions using λ Red and FLP-mediated site-specific recombination in bacteria | |
Murphy | Phage recombinases and their applications | |
Huguet et al. | A toxin antitoxin system promotes the maintenance of the IncA/C-mobilizable Salmonella Genomic Island 1 | |
Cooper et al. | Determining the specificity of cascade binding, interference, and primed adaptation in vivo in the Escherichia coli type IE CRISPR-Cas system | |
Riccobono et al. | Characterization of IncI1 sequence type 71 epidemic plasmid lineage responsible for the recent dissemination of CTX-M-65 extended-spectrum β-lactamase in the Bolivian Chaco region | |
US20130164790A1 (en) | Self-deleting plasmid | |
Xin et al. | Identification and functional analysis of potential prophage-derived recombinases for genome editing in Lactobacillus casei | |
EP2532751A1 (en) | Use of inducible promoters in the fermentative production of 1,2-propanediol | |
Kalivoda et al. | New vector tools with a hygromycin resistance marker for use with opportunistic pathogens | |
Wang et al. | IcgA is a virulence factor of Rhodococcus equi that modulates intracellular growth | |
Brantl et al. | An antisense RNA-mediated transcriptional attenuation mechanism functions in Escherichia coli | |
Jacoby et al. | Properties of IncP-2 plasmids of Pseudomonas spp | |
van Aartsen et al. | An optimized method for suicide vector-based allelic exchange in Klebsiella pneumoniae | |
DE102014212675A1 (de) | T7-Expressionssystem, Verfahren zu seiner Herstellung und seine Verwendung zur Herstellung von rekombinanten Proteinen | |
Hossain et al. | Genome modifications and cloning using a conjugally transferable recombineering system | |
Xin et al. | Targeted and repetitive chromosomal integration enables high-level heterologous gene expression in Lactobacillus casei | |
Ji et al. | Development of a dual-antimicrobial counterselection method for markerless genetic engineering of bacterial genomes | |
Tark et al. | Dual role of NER in mutagenesis in Pseudomonas putida | |
US7919322B2 (en) | Targeted deletions using retroelement-mediated placement of recombination sites |