PMID- 32116304
OWN - NLM
STAT- MEDLINE
DCOM- 20200828
LR  - 20200828
IS  - 1940-087X (Electronic)
IS  - 1940-087X (Linking)
IP  - 156
DP  - 2020 Feb 14
TI  - Agrobacterium-Mediated Immature Embryo Transformation of Recalcitrant Maize 
      Inbred Lines Using Morphogenic Genes.
LID - 10.3791/60782 [doi]
AB  - Demonstrated here is a detailed protocol for Agrobacterium-mediated genetic 
      transformation of maize inbred lines using morphogenic genes Baby boom (Bbm) and 
      Wuschel2 (Wus2). Bbm is regulated by the maize phospholipid transferase gene 
      (Pltp) promoter, and Wus2 is under the control of a maize auxin-inducible (Axig1) 
      promoter. An Agrobacterium strain carrying these morphogenic genes on transfer 
      DNA (T-DNA) and extra copies of Agrobacterium virulence (vir) genes are used to 
      infect maize immature embryo explants. Somatic embryos form on the scutella of 
      infected embryos and can be selected by herbicide resistance and germinated into 
      plants. A heat-activated cre/loxP recombination system built into the DNA 
      construct allows for removal of morphogenic genes from the maize genome during an 
      early stage of the transformation process. Transformation frequencies of 
      approximately 14%, 4%, and 4% (numbers of independent transgenic events per 100 
      infected embryos) can be achieved for W22, B73, and Mo17, respectively, using 
      this protocol.
FAU - Masters, Alicia
AU  - Masters A
AD  - Department of Agronomy, Iowa State University; Department of Applied Science and 
      Technology, Corteva Agriscience.
FAU - Kang, Minjeong
AU  - Kang M
AD  - Department of Agronomy, Iowa State University; Crop Bioengineering Center, Iowa 
      State University; Interdepartmental Plant Biology Major, Iowa State University.
FAU - McCaw, Morgan
AU  - McCaw M
AD  - Department of Agronomy, Iowa State University; Crop Bioengineering Center, Iowa 
      State University.
FAU - Zobrist, Jacob D
AU  - Zobrist JD
AD  - Department of Agronomy, Iowa State University; Crop Bioengineering Center, Iowa 
      State University; Interdepartmental Genetics and Genomics Major, Iowa State 
      University.
FAU - Gordon-Kamm, William
AU  - Gordon-Kamm W
AD  - Department of Applied Science and Technology, Corteva Agriscience.
FAU - Jones, Todd
AU  - Jones T
AD  - Department of Applied Science and Technology, Corteva Agriscience.
FAU - Wang, Kan
AU  - Wang K
AD  - Department of Agronomy, Iowa State University; Crop Bioengineering Center, Iowa 
      State University; kanwang@iastate.edu.
LA  - eng
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
PT  - Research Support, U.S. Gov't, Non-P.H.S.
PT  - Video-Audio Media
DEP - 20200214
PL  - United States
TA  - J Vis Exp
JT  - Journal of visualized experiments : JoVE
JID - 101313252
RN  - 0 (DNA, Bacterial)
RN  - 0 (T-DNA)
SB  - IM
MH  - Agrobacterium tumefaciens/*genetics
MH  - DNA, Bacterial/genetics
MH  - *Genes, Plant
MH  - *Inbreeding
MH  - Morphogenesis/*genetics
MH  - Plants, Genetically Modified
MH  - Plasmids/genetics
MH  - Pollination
MH  - *Transformation, Genetic
MH  - Zea mays/*embryology/*genetics/growth & development
EDAT- 2020/03/03 06:00
MHDA- 2020/08/29 06:00
CRDT- 2020/03/03 06:00
PHST- 2020/03/03 06:00 [entrez]
PHST- 2020/03/03 06:00 [pubmed]
PHST- 2020/08/29 06:00 [medline]
AID - 10.3791/60782 [doi]
PST - epublish
SO  - J Vis Exp. 2020 Feb 14;(156). doi: 10.3791/60782.