Automated whole mount localisation techniques for plant seedlings.
Plant biology is currently experiencing a growing demand for easy and reliable mRNA and protein localisation techniques. Here, we present novel whole mount in situ hybridisation and immunolocalisation protocols, suitable to localise mRNAs and proteins in Arabidopsis seedlings. We demonstrate that these methods can be used in different organs of Arabidopsis seedlings as well as in other plant species. In order to achieve better reproducibility and higher throughput, we modified these protocols for automation to be performed by a liquid handling robot. In addition, we show that other procedures such as reporter enzyme assays and tissue clearing can be similarly automated. We present examples of application of our protocols including mRNA localisation and proteins and epitope tag (co)localisations which demonstrate that these methods provide reliable and versatile tools for expression, localisation and anatomical studies in plants.
Friml
Jirí
J
Zentrum für Molekularbiologie der Pflanzen, Universität Tübingen, Auf der Morgenstelle 3, 72076 Tübingen, Germany. jiri.friml@zmbp.uni-tuebingen.de
Benková
Eva
E
Mayer
Ulrike
U
Palme
Klaus
K
Muster
Gerhard
G
D017360:Arabidopsis / Q000235:genetics / Q000378:metabolism; D001331:Automation / Q000295:instrumentation / Q000379:methods; D020869:Gene Expression Profiling / Q000379:methods; D018507:Gene Expression Regulation, Developmental; D018506:Gene Expression Regulation, Plant; D007150:Immunohistochemistry / Q000379:methods*; D017403:In Situ Hybridization / Q000379:methods*; D028528:Petroselinum / Q000235:genetics / Q000378:metabolism; D010940:Plant Proteins / Q000032:analysis* / Q000235:genetics*; D012333:RNA, Messenger / Q000032:analysis / Q000235:genetics; D015203:Reproducibility of Results; D036226:Seedlings / Q000235:genetics* / Q000378:metabolism*; D014026:Nicotiana / Q000235:genetics / Q000378:metabolism
D010940:Plant Proteins; D012333:RNA, Messenger