GSE168821
Bulk RNA sequencing on sorted blood cells from Drosophila lymph glands [MZ IZ bulk RNA-seq]
We report the use of bulk mRNA sequencing on FACS-sorted blood cells to identify new and unique genetic markers for each major zone of the  Drosophila lymph gland. We used flow cytomotery in a wild-type genetic background whihc contains two fluorescent reporters that allow for the separation of 4 distinct populations.  Progenitors of the medullary zone (MZ) are marked by dome-meso-EGFP expression. Differentitating cels of the cortical zone (CZ) are marked by Hml-DsRed. These two sorted populations (MZ and CZ) are positive for the expected zone-specific markers (i.e. Tep4,  dome, and shg for MZ and Hml, Pxn, vkg, Col4a1, and NimC1 for plasmatocytes of the CZ), however, the CZ population also expresses crystal cell markers (hnt, lz, ProPO), indicating that it is a mix of plasmatocytes and crystal cells. Intermediate progenitors of the intermediate zone (IZ), which are not well understood and do not have specific markers, but are characterized by simultaneous expression of EGFP and DsRed (double positives) were also sorted. In addtiion, there is a population of lymph gland cells that is double negative for both EGFP and DsRed fluorescence. This double negative population contains a mix of cell types including posterior signaling center (PSC) cells, MZ and CZ cells that have lost fluorescence, late crystal cells that do not express Hml (early CCs do express), and non-blood cells (i.e. dorsal vessel and pericardial cells). These double negatives were excluded from the analysis for these reasons. We analyzed transcript data of these 4 sorted populations in the early third instart larvae (90-96 hours after eggs lay at 25 degrees C). We identified novel markers that are differentially expressed in each of the three major blood cell populations (MZ, IZ, CZ) that have been useful to definetely identify populations in scRNA-Seq and give information about transcript isoforms in each of the distinct populations. Overall design: Examination of mRNA expression in sorted  blood cell populations