Article Title: Cerebral organoids model human brain development and microcephaly
Figure Lengend Snippet: Human cerebral organoids recapitulate various brain region identities a, RT-PCR for forebrain markers (BF1 and Six3) and hindbrain markers (Krox20 and Isl1) at 12, 16 and 20 days of differentiation. Human fetal brain cDNA was used as positive control. b, Immunohistochemistry in serial sections for the forebrain marker Pax6 (red, first panel) and the hindbrain markers Krox20 (green, first panel) and Pax2 (red, second panel) at 16 days of differentiation. Note the juxtaposition reminiscent of the mid-hindbrain boundary (arrows). DAPI marks nuclei (blue). c,-i, Staining for various brain region identities: forebrain, FoxG1 ( c ); dorsal cortex, Emx1 ( d ); prefrontal cortex (note the discrete boundary, arrow), Auts2 ( e ); hippocampus, Nrp2, Fzd9, Prox1 ( f ); ventral forebrain, Nkx2.1 ( g ) and choroid plexus, TTR ( i ). g, Staining for adjacent ventral (arrow) and dorsal (Pax6, arrowhead) forebrain and for calretinin (green) in a serial section revealing cortical interneurons in the ventral region (arrow). Calretinin interneurons within dorsal cortex ( h ) exhibit typical morphology of tangential migration (arrows). j, Hematoxylin-eosin staining of retinal tissue exhibiting stereotypical layering: retinal pigment epithelium (RPE), outer nuclear layer (ONL) and inner nuclear layer (INL). Scale bars: 100 μm.
Article Snippet: The CDK5RAP2 expression construct was generated using the Gateway system (Invitrogen) by PCR amplification of CDK5RAP2 from MGC human CDK5RAP2 cDNA (clone ID: 9052276) using the primers with AttB sites: Forward: GGGGACAAGTTTGTACAAAAAAGCAGGCTTCATGATGGACTTGGTGTTGGAAGA, Reverse: GGGGACCACTTTGTACAAGAAAGCTGGGTCAGCTTTATTGGCTGAAAGTTCTTCTC.
Techniques: Reverse Transcription Polymerase Chain Reaction, Positive Control, Immunohistochemistry, Marker, Staining, Migration