2/23/16

Class Notes FPKM-->should be entering .expected_count

Dr. Heyer finished code to get supervised clustering, larger data set is not really graphing, Dr. Heyer set up a way to filter out if row for gene if average of six genes is greater than 10

From missed class

Looking for housekeeping intestine genes so that we can make sure our samples are accurate over-expressed sequences match:

Snake 1: matches with an mRNA sequence from an intestine sample of an elephant fish, SLC15A1 (FPKM 14366.5) , SLC6A19 (FPKM 7306.48), Aminopeptidase (FPKM 494.97) sucrase/maltase (FPKM 935.83)

Snake 2: SLC15A1 (FPKM 13466.62), SLC6A19 (FPKM 4132.78), sucrase/maltase (FPKM 1133.27), Aminopeptidase (FPKM 432.93)

Snake 3: SLC15A1 (FPKM 14322.98), SLC6A19 (FPKM 5695.09), sucrase/maltase (FPKM 1430.56), Aminopeptidase (FPKM 426.59)

Snake 4: SLC6A19 (FPKM 13213.57), SLC15A1 (FPKM 14316.17), maltase/sucrase (FPKM 747.19) aminopeptidase (FPKM 556.07)

Snake 5: SLC15A1 (FPKM 11347.62), SLC6A19 (FPKM 4130.4), sucrase/maltase (FPKM 1054.27) , aminopeptidase (FPKM 513.37)

Snake 6: SLC15A1 (FPKM 9310.97), SLC6A19 (FPKM 6632.26), sucrase/maltase (FPKM 735.21), aminopeptidase (FPKM 488.85)

Housekeeping genes

BOAT1 (SLC6A19) solute carrier family 6 (neutral amino acid transporter), member 19, mRNA (housekeeping gene BOAT1) found in intestine and kidney

SLC15A1 protein coding gene: encodes an intestinal hydrogen peptide cotransporter that is a member of the solute carrier family 15

Amino peptidase (Aspartyl_aminopeptidase_Homo_sapiens): source small intestine, products are amino acids and peptides

maltase/sucrose (Si_Sucrase-isomaltase,_intestinal_Rattus_norvegicus) : source small intestine, products are glucose and fructose

Housekeeping genes that weren't highly expressed

Important to note than many housekeeping genes for intestines were not extremely highly expressed at all in our samples, not as great evidence as was found for the liver samples

GATA6 (Regulates proximal-distal identity in the intestines) 1) 32.2 2) 33.96 3) 19.17 4) 37.87 5) 31.67 6) 31.98

MYBL2 (Regulates commitment of colon stem cells to differentiate) 1) 0.04 2) 0 3) 0.05 4) 0 5) 0 6) 0

ASCT2 (SLC1A5 expressed in the kidney but expression is high in the jejunum and colon but lower in duodenum and ileum) 1) 2.29 2) 2.96 3) 2.65 4) 1.39 5) 3.75 6) 3.07

STX2, COL3A1, GPBAR1: all from Castoe et al paper; examples of genes that have experienced positive selection (P < 0.001) on snake lineages and are related to prominent phenotypic or cellular traits of snakes- HOWEVER NOT HIGHLY EXPRESSED IN ANY OF OUR SNAKES (range of FPKM from 0 - 4)

Began process of creating new supervised clusters using R code

Here's some interesting info: Total Reads-->4i 7,061,976 5i 11,282,296 6i 11,912,035

We're having trouble with the supervised clustering because of the fewer reads in Snake 4, tried again using clustering