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农艺性状表型数据与环境关联互作分析
local({r <- getOption("repos")
r["CRAN"] <- "http://mirrors.tuna.tsinghua.edu.cn/CRAN/"
options(repos=r)})
options(BioC_mirror="https://mirrors.tuna.tsinghua.edu.cn/bioconductor")
#install.packages("GGEBiplots")
#install.packages("GGEBiplotGUI")
library(GGEBiplotGUI)
library("GGEBiplots")
library("reshape2")
library(vegan)
library(corrplot)
library("psych")
library(Cairo)
setwd("D:/potato/")
data1<-read.table("张北data.txt",header = F,row.names = 1,comment.char = "",sep = "\t",encoding = "UTF-8")
data.t<-as.data.frame(t(data1))
data.l<-melt(data.t,id.vars = c('年度', '性状'),variable.name = "基因型")
mydata1<-dcast(data.l, `年度` + `基因型` ~ `性状`)
#`单株产量` + `淀粉含量(%)` + `干物质含量(%)` + `平均出苗率(%)`+ `生育日数(天)`+ `株高`
data2<-read.table("康保data.txt",header = F,row.names = 1,comment.char = "",sep = "\t",encoding = "UTF-8")
data.t<-as.data.frame(t(data2))
data.l<-melt(data.t,id.vars = c('年度', '性状'),variable.name = "基因型")
mydata2<-dcast(data.l, `年度` + `基因型` ~ `性状`)
#`单株产量` + `淀粉含量(%)` + `干物质含量(%)` + `平均出苗率(%)`+ `生育日数(天)` + `株高`
#inter<-intersect(colnames(mydata1),colnames(mydata2))
#去掉一些数据
inter<-c("年度" , "基因型" , "单株产量" ,
"淀粉含量(%)" , "干物质含量(%)" , "平均出苗率(%)" ,
"生育日数(天)" , "株高" )
mydata1.c<-mydata1[,inter]
mydata2.c<-mydata2[,inter]
mydata1.c$`环境`="张北"
mydata2.c$`环境`="康保"
mydata=rbind(mydata1.c,mydata2.c)
########################计算平均值
#traits<-c("单株产量" , "淀粉含量(%)" , "干物质含量(%)" , "平均出苗率(%)", "生育日数(天)" , "小区产量" ,"折合单位产量kg/hm", "株高")
traits=c( "单株产量" , "淀粉含量(%)" ,
"干物质含量(%)" , "平均出苗率(%)" ,
"生育日数(天)" ,
"株高" )
for (i in traits){
mydata[,i]=as.double(mydata[,i])
}
SD=round(apply(mydata[,traits],2,sd),2)
MEAN=round(apply(mydata[,traits],2,mean),2)
RANGE=round(apply(mydata[,traits],2,range),2)
CV=round(SD/MEAN*100,2)
H=round(diversity(mydata[,traits], index = "shannon", MARGIN = 2),2)
trait.mean=cbind(`平均值`=MEAN,`标准差`=SD,`变异系数(%)`=CV,`变幅`=paste(range.min=RANGE[1,],range.max=RANGE[2,],sep="-"),`多样性指数 H’`=H)
write.csv(trait.mean,file="01.马铃薯表型性状统计.csv")
#################################相关性分析
occor = corr.test(mydata[,traits],use="pairwise",method="spearman",adjust="BH",alpha=.05)
occor.r = occor$r # 取相关性矩阵R值
occor.p = occor$p # 取相关性矩阵p值
#设置色板
palette_2 <- colorRampPalette(c("yellow","red"))(100)
#绘图
#png(file="traits_cor.png",w=10*300,h=10*300,res = 300)
pdf(file="traits_cor.pdf",w=10,h=10,family="GB1")
corrplot(occor.r, method = "square", type = "lower", order="AOE",
col = palette_2, tl.pos = "tp", tl.col = "blue", cl.pos = "r",
title = "Traits corr", mar = c(2,4,4,1))
corrplot(occor.r, method = "number", type = "upper",order="AOE",
col = palette_2,add = TRUE, diag = FALSE, tl.pos = "n", cl.pos = "n",
number.cex=1,mar = c(2,4,4,1))
dev.off()
write.csv(occor.p,file="02.相关性P值.csv")
write.csv(occor.r,file="03.相关性R值.csv")
########################分布范围柱状图
#########################求平均
genotype.mean=aggregate(mydata[,traits],by=list(`基因型`=mydata$`基因型`),mean)
for(t in traits){
#t="平均出苗率(%)"
d<- genotype.mean[,t]
ff=gsub("\\(\\S+\\)", "", t)
ff=gsub("kg/hm", "", ff)
pdf(paste0(ff,"-表型性状值分布.pdf"),w=5,h=5,family="GB1")
par(mar=c(6,4.1,2,4))
xhist <- hist(d,breaks=seq(from=floor(min(d)),to=ceiling(max(d)),length.out=10),
plot=F,xlab =t ,ylim=c(0,),col="white",xaxt="n", yaxs="i",fill=NULL,ylab = "样本数",freq=T,main="")
xhist <- hist(d,breaks=seq(from=floor(min(d)),to=ceiling(max(d)),length.out=10),
plot=T,xlab ="" ,ylim=c(0,max(xhist$counts)*1.1),col="white",xaxt="n", yaxs="i",fill=NULL,ylab = "样本数",freq=T,main="")
mtext(t,side=1,line=4.5)
axis(side=1,xhist$mids,labels=F)
b=(xhist$mids[2]-xhist$mids[1])/2
s=round(xhist$mids-b,2)
s[1]=0
e=round(xhist$mids+b,2)
lab=paste(s,e,sep="-")
text(x=xhist$mids, y=-3, labels=lab,cex=1, xpd=TRUE, srt=45,adj=1)
dd=density(d)
par(new=T)
plot(dd, ylim=c(0,max(dd$y)*1.1), col ='red',lwd=2,yaxs="i",xlab = "",ylab="",xaxt="n",yaxt="n",main = "")
axis(4, las=1,at=seq(from = 0, to = max(xhist$density), length.out = 6), labels=round(seq(0, max(xhist$density),length.out =6)/sum(xhist$density),2), col = 'red', col.axis = 'red')
mtext("频率",side=4,line=3,col="red")
box(bty="l")
dev.off()
}
##############GGEbioplots
ggdata=mydata[,c("环境","基因型","单株产量")]
ggdata1=aggregate(`单株产量` ~ 基因型 + 环境, data = mydata, mean)
gg.f=dcast(ggdata1,基因型 ~环境)
row.names(gg.f)=gg.f[,1]
gg.f=gg.f[,-1]*10
#GGEBiplot(Data = gg.f)
library(ggplot2)
GGE1<-GGEModel(gg.f,centering = "tester",
SVP = "symmetrical",
scaling = "none")
g1=GGEPlot(GGE1,type=6,sizeGen=4,sizeEnv = 2,largeSize=3)
g1=g1+ labs(title = "")
pdf(paste0("适应性分析bioplot.pdf"),w=5,h=5,family="GB1")
print(g1)
dev.off()
g2=GGEPlot(GGE1,type=9,sizeGen=2,sizeEnv = 3,largeSize=3)
g2=g2+ labs(title = "")
pdf(paste0("丰产性-稳定性分析bioplot.pdf"),w=5,h=5,family="GB1")
print(g2)
dev.off()
#################################表型主成分分析
p=genotype.mean
row.names(p)=p[,1]
p=p[,-1]
pca = prcomp(t(p), scale=TRUE)
#pca = prcomp(t(decostand(p, "hellinger")), scale=TRUE)
ss=summary(pca)
pc=pca$x
xx=ss$importance
row.names(xx)<-c("特征值E","贡献率CR","累计贡献率CCR")
pca.res=rbind(pc,xx)
write.csv(pca.res,file="04.PCA分析结果.csv")
#表型数据聚类分析
dist_mat <- dist(genotype.mean[,2:5], method = "euclidean")
clustering <- hclust(dist_mat, method = "complete")
plot(clustering,labels=genotype.mean$基因型,xlab="sample",ylab="Height" ,main="")
#根据进化树分组
group.data=genotype.mean
group.data$group=as.factor(cutree(clustering,h=15))
names=c("基因型" , "单株产量" , "淀粉含量" , "干物质含量",
"平均出苗率" ,"生育日数" ,"株高" , "group" )
colnames(group.data)=names
library("ggpubr")
phenotype=names[2:(length(names)-1)]
test.res=data.frame()
for(i in phenotype){
f=as.formula(paste0("`",i,"`~group"))
print(f)
res=compare_means(f, data = group.data,method="t.test")
print(res)
test.res=rbind(test.res,res)
}
write.csv(test.res,file="聚类树分组差异统计检验.csv")
write.csv(group.data,file="聚类树分组信息表.csv")
g1.res=aggregate(group.data[,2:7],by=list(group.data$group),FUN=mean)
g2.res=aggregate(group.data[,2:7],by=list(group.data$group),FUN=sd)
write.csv(g1.res,file="聚类树分组统计平均值.csv")
write.csv(g2.res,file="聚类树分组统计标准差.csv")
#表型数据在不同环境下变异
for (i in traits){
mydata1[,i]=as.double(mydata1[,i])
}
SD=round(apply(mydata1[,traits],2,sd),2)
MEAN=round(apply(mydata1[,traits],2,mean),2)
RANGE=round(apply(mydata1[,traits],2,range),2)
CV=round(SD/MEAN*100,2)
H=round(diversity(mydata1[,traits], index = "shannon", MARGIN = 2),2)
trait.mean=cbind(`平均值`=MEAN,`标准差`=SD,`变异系数(%)`=CV,`变幅`=paste(range.min=RANGE[1,],range.max=RANGE[2,],sep="-"),`多样性指数 H’`=H)
write.csv(trait.mean,file="01.马铃薯表型性状统计(张北).csv")
#表型数据在不同环境下变异
for (i in traits){
mydata2[,i]=as.double(mydata2[,i])
}
SD=round(apply(mydata2[,traits],2,sd),2)
MEAN=round(apply(mydata2[,traits],2,mean),2)
RANGE=round(apply(mydata2[,traits],2,range),2)
CV=round(SD/MEAN*100,2)
H=round(diversity(mydata2[,traits], index = "shannon", MARGIN = 2),2)
trait.mean=cbind(`平均值`=MEAN,`标准差`=SD,`变异系数(%)`=CV,`变幅`=paste(range.min=RANGE[1,],range.max=RANGE[2,],sep="-"),`多样性指数 H’`=H)
write.csv(trait.mean,file="01.马铃薯表型性状统计(康保).csv")
######关联分析
mydata$年度=as.factor(mydata$年度)
mydata$环境=as.factor(mydata$环境)
F.res=data.frame()
for (i in traits){
print(i)
fl=c(as.formula(paste0("`",i,"`~基因型")),
as.formula(paste0("`",i,"`~环境")),
as.formula(paste0("`",i,"`~年度")),
as.formula(paste0("`",i,"`~基因型*环境")),
as.formula(paste0("`",i,"`~基因型*年度"))
#as.formula(paste0("`",i,"`~基因型*年度*环境"))
)
for(f in fl){
blp=lm(f,data=mydata)
aa=summary(blp)
#
#print(aa)
print(aa$df)
print(f)
fstatistic = aa$fstatistic
p_value = pf(as.numeric(fstatistic[1]), as.numeric(fstatistic[2]), as.numeric(fstatistic[3]), lower.tail = FALSE)
print(p_value)
aa=data.frame(`公式`=as.character(f)[3],df=aa$df[1],F=fstatistic[1],P=p_value,`性状`=i)
F.res=rbind(F.res,aa)
# question1 <- readline("Would you like to proceed untill the loop ends? (Y/N)")
# if(regexpr(question1, 'y', ignore.case = TRUE) == 1){
# continue = TRUE
# next
# } else{
# break
# }
}
}
write.csv(F.res,file="05.表型与基因型环境年度互作效应.csv")
第二版版本,注意PCA分析升级,输入数据行为样本,列为表型;
local({r <- getOption("repos")
r["CRAN"] <- "http://mirrors.tuna.tsinghua.edu.cn/CRAN/"
options(repos=r)})
options(BioC_mirror="https://mirrors.tuna.tsinghua.edu.cn/bioconductor")
#install.packages("GGEBiplots")
#install.packages("GGEBiplotGUI")
library(GGEBiplotGUI)
library("GGEBiplots")
library("reshape2")
library(vegan)
library(corrplot)
library("psych")
library(Cairo)
setwd("D:/potato/第二次分析")
mydata<-read.table("data.txt",header = T,row.names = 1,comment.char = "",sep = "\t",encoding = "UTF-8")
########################计算平均值
traits=colnames(mydata)
for (i in traits){
mydata[,i]=as.double(mydata[,i])
}
SD=round(apply(mydata[,traits],2,sd),2)
MEAN=round(apply(mydata[,traits],2,mean),2)
RANGE=round(apply(mydata[,traits],2,range),2)
CV=round(SD/MEAN*100,2)
H=round(diversity(mydata[,traits], index = "shannon", MARGIN = 2),2)
J=round(H/log(specnumber(t(mydata[,traits]))),2)
trait.mean=cbind(`平均值`=MEAN,`标准差`=SD,`变异系数(%)`=CV,`变幅`=paste(range.min=RANGE[1,],range.max=RANGE[2,],sep="~"),`H’`=H,J=J)
write.csv(trait.mean,file="01.马铃薯表型性状统计.csv")
########################分布范围柱状图
#########################求平均
for(t in traits){
##t="顶小叶宽"
#t="出苗率"
d<- mydata[,t]
pdf(paste0(t,"-表型性状值分布.pdf"),w=5,h=5,family="GB1")
par(mar=c(6,4.1,2,4))
MIN=min(d)
MAX=max(d)
step=(MAX-MIN)/(length(d)^(1/2)+1)
xhist <- hist(d,breaks=seq(from=floor(min(d)),to=ceiling(max(d))+step,by=step),
plot=F,xlab =t ,ylim=c(0,),col="white",xaxt="n", yaxs="i",fill=NULL,ylab = "样本数",freq=T,main="")
xhist <- hist(d,breaks=seq(from=floor(min(d)),to=ceiling(max(d))+step,by=step),
plot=T,xlab ="" ,ylim=c(0,max(xhist$counts)*1.1),col="green",xaxt="n", yaxs="i",fill=NULL,ylab = "样本数",freq=T,main="")
mtext(t,side=1,line=4.5)
axis(side=1,xhist$mids,labels=F)
s=round(xhist$mids-step,2)
if (s[1]<0) {s[1]=0}
e=round(xhist$mids+step,2)
lab=paste(s,e,sep="-")
text(x=xhist$mids, y=-3, labels=lab,cex=1, xpd=TRUE, srt=45,adj=1)
axis(4, las=1,at=seq(from = 0, to = max(xhist$counts), length.out = 4), labels=paste0(round(seq(0, max(xhist$counts),length.out =4)/sum(xhist$counts),4)*100,"%"), col = 'red', col.axis = 'red')
dd=density(d)
par(new=T)
plot(dd, ylim=c(0,max(dd$y)*1.1), col ='red',lwd=2,yaxs="i",xlab = "",ylab="",xaxt="n",yaxt="n",main = "")
#mtext("频率",side=4,line=3,col="red")
box(bty="l")
l.at="topright"
if(t=="出苗率" || t=="叶缘形状"){
l.at="topleft"
}
legend(l.at,legend = c("直方图"),
fill = c('green'),
bty='n')
legend(l.at,legend = c("正态图"),
col = c("red"), lty=1,
bty='n',inset = c(0,0.08), lwd=3)
dev.off()
}
################相关性分析###################
#################################相关性分析
occor = corr.test(mydata,use="pairwise",method="spearman",adjust="BH",alpha=.05)
occor.r = occor$r # 取相关性矩阵R值
occor.p = occor$p # 取相关性矩阵p值
#设置色板
palette_2 <- colorRampPalette(c("yellow","red"))(100)
#绘图
#png(file="traits_cor.png",w=10*300,h=10*300,res = 300)
pdf(file="traits_cor.pdf",w=10,h=10,family="GB1")
corrplot(occor.r, method = "square", type = "lower", order="AOE",
col = palette_2, tl.pos = "tp", tl.col = "blue", cl.pos = "r",
title = "Traits corr", mar = c(2,8,4,1))
corrplot(occor.r, method = "number", type = "upper",order="AOE",
col = palette_2,add = TRUE, diag = FALSE, tl.pos = "n", cl.pos = "n",
number.cex=1,mar = c(2,4,4,1))
dev.off()
write.csv(occor.p,file="02.相关性P值.csv")
write.csv(occor.r,file="03.相关性R值.csv")
###############因子分析###############
mydata.l=melt(mydata)
bt=bartlett.test(value~variable,data=mydata.l)
kmo=KMO(mydata)
sink("04.因子分析.txt")
bt
kmo
sink()
#################################表型主成分分析
#pca = prcomp(mydata, scale=TRUE)
pca = princomp(mydata, scores=TRUE, cor=TRUE)
#pca = prcomp(t(mydata), scale=TRUE)
fa1 = factanal(mydata, factor=2, rotation="varimax", scores="regression")
fa1
#write.csv(abs(fa1$loadings), "loadings.csv")
pdf(file="04.PCA分析特征根碎石图.pdf",w=8,h=4,family="GB1")
screeplot(pca, type="line", main="Scree Plot")
dev.off()
sink(file="04.PCA分析summary.txt")
summary(pca)
sink()
pc=loadings(pca)[1:21,]
pc=as.data.frame(pc)
write.csv(pc,file="04.PCA分析结果.csv")
library(RColorBrewer)
mycolor=c(brewer.pal(9, "Set1"),brewer.pal(8, "Set2"),brewer.pal(9, "Paired"))
pdf(file="04.PCA分析表型分布.pdf",w=7,h=6,family="GB1")
par(mar=c(5,4,4,10))
plot(x=pc[,1],y=pc[,2],cex=1.5,col=mycolor,pch=16,xlab="factor 1",ylab = "factor 2")
abline(h=0,lty=1)
abline(v=0,lty=1)
legend("topright",legend =rownames(pc) ,pch=16,col=mycolor,xpd =T,inset = c(-0.3,0),ncol=1,bty='n',cex=1)
dev.off()
#表型数据聚类分析
dist_mat <- dist(mydata, method = "euclidean")
clustering <- hclust(dist_mat, method = "complete")
pdf(file="05.聚类图.pdf",w=15,h=4,family="GB1")
plot(clustering,labels=rownames(mydata),cex=0.55,xlab="sample",ylab="Height" ,main="")
abline(h=40,col="red")
dev.off()
#根据聚类树分组
group.data=mydata
group.data$group=as.factor(cutree(clustering,h=40))
###################分组PCA图################
#install.packages("ggfortify")
library(ggfortify)
pdf(file="04.PCA分析表型样本双序图.pdf",w=5,h=4,family="GB1")
#par(mar=c(5,4,4,10))
#pcs=pca$scores
#plot(x=pcs[,1],y=pcs[,2],cex=1.5,col=mycolor[group.data$group],pch=1,xlab="factor 1",ylab = "factor 2")
#par(new=T)
#plot(x=pc[,1],y=pc[,2],cex=1.5,col="blue",pch=16,xlab="factor 1",ylab = "factor 2")
#legend("topright",legend =rownames(pc) ,pch=16,col=mycolor,xpd =T,inset = c(-0.3,0),ncol=1,bty='n',cex=1)
#legend("topright",legend =1:6 ,pch=16,col=mycolor[1:6],xpd =T,inset = c(-0.3,0),ncol=1,bty='n',cex=1)
#biplot(pca)
g=autoplot(pca, data = group.data, colour = 'group',
loadings = TRUE, loadings.colour = 'blue',
loadings.label = TRUE, loadings.label.size = 3,size=2)#,frame = TRUE, frame.type = 'norm')
g=g+scale_color_manual(name="group",values =mycolor)+
scale_fill_manual(name="group",values =mycolor)+
theme_bw()+ theme(
panel.grid=element_blank(),
axis.text.x=element_text(colour="black"),
axis.text.y=element_text(colour="black"),
panel.border=element_rect(colour = "black"),
legend.key = element_blank(),
plot.title = element_text(hjust = 0.5))
g
dev.off()
library("ggpubr")
phenotype=traits
test.res=data.frame()
for(i in phenotype){
f=as.formula(paste0("`",i,"`~group"))
print(f)
fit=aov(f,data=group.data)
ss=summary(fit)
res=as.data.frame(ss[[1]])
res$`表型`=i
rownames(res)=c("组间","组内")
print(res)
test.res=rbind(test.res,res)
}
colnames(test.res)=c("Df","平方和SS","均方MS","F","显著性","性状trait")
write.csv(test.res,file="05.聚类树分组差异统计检验.csv")
write.csv(group.data,file="05.聚类树分组信息表.csv")
g1.res=aggregate(group.data[,traits],by=list(group.data$group),FUN=mean)
Average=apply(group.data[,traits],2,mean)
Average=c("Group.1"="总体平均值 Average",Average)
g.mean=rbind(g1.res,t(as.data.frame(Average)))
write.csv(t(g.mean),file="06.聚类树分组统计平均值.csv")
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