My PROJECT

Amoa project

A2,A3  amoa sample sequencing

(1) use geneious to deal with the sequence raw data,  get 635bp DNA sequence

(2)use mafft to get the alignment results

(3)mothur deal pipline

mothur > dist.seqs(fasta=AMOA_A2A3.mafft.align.shortname.fasta, output=lt)

0 0
59 0

Output File Name:
AMOA_A2A3.mafft.align.shortname.phylip.dist

It took 0 to calculate the distances for 60 sequences.

mothur > cluster(phylip=AMOA_A2A3.mafft.align.shortname.phylip.dist, cutoff=0.05)

********************#****#****#****#****#****#****#****#****#****#****#
Reading matrix: ||||||||||||||||||||||||||||||||||||||||||||||||||||
***********************************************************************
unique 4 49 2 1 1
0.01 11 22 4 3 0 2 0 0 0 0 0 1
0.02 16 15 4 2 0 0 0 0 0 0 0 0 0 0 0 1 1
0.03 36 10 3 1 0 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 00 0 0 0 0 0 0 0 0 0 0 0 0 0 1
0.04 38 9 1 0 1 0 0 1 0 0 0 0 0 0 0 0 0 0 0 0 0 00 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1
changed cutoff to 0.0352006

Output File Names:
AMOA_A2A3.mafft.align.shortname.phylip.an.sabund
AMOA_A2A3.mafft.align.shortname.phylip.an.rabund
AMOA_A2A3.mafft.align.shortname.phylip.an.list

It took 0 seconds to cluster

mothur > unique.seqs(fasta=AMOA_A2A3.mafft.align.shortname.fasta)

60 53

Output File Names:
AMOA_A2A3.mafft.align.shortname.unique.fasta
AMOA_A2A3.mafft.align.shortname.names

mothur > parse.list(list=AMOA_A2A3.mafft.align.shortname.phylip.an.list, group=AMOA_A2A3.mafft.align.shortname.group)

unique
0.01
0.02
0.03
0.04

Output File Names:
AMOA_A2A3.mafft.align.shortname.phylip.an.A2.list
AMOA_A2A3.mafft.align.shortname.phylip.an.A3.list

mothur > rarefaction.single(list=AMOA_A2A3.mafft.align.shortname.phylip.an.A3.list, iters=10000, freq=0.10, calc=ace-sobs-chao, processors=3)

unique
0.01
0.02
0.03
0.04

Output File Names:
AMOA_A2A3.mafft.align.shortname.phylip.an.A3.r_ace
AMOA_A2A3.mafft.align.shortname.phylip.an.A3.rarefaction
AMOA_A2A3.mafft.align.shortname.phylip.an.A3.r_chao
mothur > rarefaction.single(list=AMOA_A2A3.mafft.align.shortname.phylip.an.A2.list, iters=10000, freq=0.10, calc=ace-sobs-chao, processors=3)

unique
0.01
0.02
0.03
0.04

Output File Names:
AMOA_A2A3.mafft.align.shortname.phylip.an.A2.r_ace
AMOA_A2A3.mafft.align.shortname.phylip.an.A2.rarefaction
AMOA_A2A3.mafft.align.shortname.phylip.an.A2.r_chao

mothur > rarefaction.single(list=AMOA_A2A3.mafft.align.shortname.phylip.an.A3.list, iters=10000, freq=0.10, calc=shannon, processors=3)

unique
0.01
0.02
0.03
0.04

Output File Names:
AMOA_A2A3.mafft.align.shortname.phylip.an.A3.r_shannon
mothur > rarefaction.single(list=AMOA_A2A3.mafft.align.shortname.phylip.an.A2.list, iters=10000, freq=0.10, calc=shannon, processors=3)

unique
0.01
0.02
0.03
0.04

Output File Names:
AMOA_A2A3.mafft.align.shortname.phylip.an.A2.r_shannon

 

mothur > get.oturep(phylip=AMOA_A2A3.mafft.align.shortname.phylip.dist, fasta=AMOA_A2A3.mafft.align.shortname.fasta, list=AMOA_A2A3.mafft.align.shortname.phylip.an.A2.list, group=AMOA_A2A3.mafft.align.shortname.group)

********************#****#****#****#****#****#****#****#****#****#****#
Reading matrix: ||||||||||||||||||||||||||||||||||||||||||||||||||||
***********************************************************************
unique 27
0.01 17
0.02 12
0.03 7
0.04 6

Output File Names:
AMOA_A2A3.mafft.align.shortname.phylip.an.A2.unique.rep.names
AMOA_A2A3.mafft.align.shortname.phylip.an.A2.0.01.rep.names
AMOA_A2A3.mafft.align.shortname.phylip.an.A2.0.02.rep.names
AMOA_A2A3.mafft.align.shortname.phylip.an.A2.0.03.rep.names
AMOA_A2A3.mafft.align.shortname.phylip.an.A2.0.04.rep.names
AMOA_A2A3.mafft.align.shortname.phylip.an.A2.0.01.rep.fasta
AMOA_A2A3.mafft.align.shortname.phylip.an.A2.0.02.rep.fasta
AMOA_A2A3.mafft.align.shortname.phylip.an.A2.0.03.rep.fasta
AMOA_A2A3.mafft.align.shortname.phylip.an.A2.0.04.rep.fasta
AMOA_A2A3.mafft.align.shortname.phylip.an.A2.unique.rep.fasta
mothur > get.oturep(phylip=AMOA_A2A3.mafft.align.shortname.phylip.dist, fasta=AMOA_A2A3.mafft.align.shortname.fasta, list=AMOA_A2A3.mafft.align.shortname.phylip.an.A3.list, group=AMOA_A2A3.mafft.align.shortname.group)

********************#****#****#****#****#****#****#****#****#****#****#
Reading matrix: ||||||||||||||||||||||||||||||||||||||||||||||||||||
***********************************************************************
unique 29
0.01 22
0.02 16
0.03 13
0.04 11

Output File Names:
AMOA_A2A3.mafft.align.shortname.phylip.an.A3.unique.rep.names
AMOA_A2A3.mafft.align.shortname.phylip.an.A3.0.01.rep.names
AMOA_A2A3.mafft.align.shortname.phylip.an.A3.0.02.rep.names
AMOA_A2A3.mafft.align.shortname.phylip.an.A3.0.03.rep.names
AMOA_A2A3.mafft.align.shortname.phylip.an.A3.0.04.rep.names
AMOA_A2A3.mafft.align.shortname.phylip.an.A3.0.01.rep.fasta
AMOA_A2A3.mafft.align.shortname.phylip.an.A3.0.02.rep.fasta
AMOA_A2A3.mafft.align.shortname.phylip.an.A3.0.03.rep.fasta
AMOA_A2A3.mafft.align.shortname.phylip.an.A3.0.04.rep.fasta
AMOA_A2A3.mafft.align.shortname.phylip.an.A3.unique.rep.fasta

R studio 画图

> setwd("/home/shenzy/Desktop/amoA_work")
> data0<-read.table(file="AMOA_A2A3.mafft.align.shortname.phylip.an.A2.rarefaction",header=T)
> data1<-read.table(file="AMOA_A2A3.mafft.align.shortname.phylip.an.A3.rarefaction",header=T)
> plot(x=data0$numsampled, y=data0$X0.03, xlab="Number of clones analyzed",ylab="Number of OTUs observed", type="l", col="green", xlim=c(0,30),ylim=c(0,15))
> points(x=data1$numsampled, y=data1$X0.03, type="l", col="blue")
> legend(x=2, y=14, c("A2","A3"),c("green","blue")