| Clupeiformes belonging to the superorder Clupeomorphe including teleosts,head of the Cretaceous fossil taxa and clupeiformes,and clupeiformes mainly refers to all living groups.The Clupeiformes is a good taxonomic group for studying marine biogeography,and adaptive evolution of marine fish invading freshwaters.The Clupeiformes is a good taxonomic group for studying marine biogeography,and adaptive evolution of marine fish invading freshwaters and is an order of fish in the family of the most widely distributed today.The Clupeiformes(Osteichthys)is classified into 2 suborders,5 families,84 genera and about 402 species.However,the phylogenetic relationships among the clupeiforms are still controversial.The traditional classification is incongruent with phylogenies based on molecular data,which themselves are not consistent with each other either.At present,the research and development of clupeiformes as a whole system is very few,and the past research is mainly using morphological method,namely the use of molecular data,only a few sites of mitochondrial data.The results of several research of Lavou et al,systematic position dorab are different from each other,and clupeiformes in many lineages of the bootstrap support rate is very low,the relationship between internal departments clupeiformes cannot be determined.The common conclusion is the molecular classification of Clupeidae is no longer a monophyletic group,but is divided into several groups,but the relationship between these groups and clupeiformes subjects based on mitochondrial genome and nuclear genes with conflicting results,no definite conclusion.The confusion in the classification not only greatly influenced the study of clupeiformes system evolution,also cannot explain the contradiction between fossil data and biogeographic deduction conclusion,and adaptive evolution in false positive results.The phylogenetic relationships among the clupeiforms are still controversial.The traditional classification is incongruent with phylogenies based on molecular data,which themselves are not consistent with each other either.The major reasons could be that there were a limited number of loci used in the previous molecular studies,thus did not provide adequate phylogenetic signal to resolve those deep internal nodes.The other cause may be that many genes have highly biased GC content(50%-80%)among different Clupeiform species,thus,current analyses often mistakenly group unrelated taxa with similar GC content.In this research,we collected representative species(already collected samples from 5 families and 48 genera)and address the aforementioned problems in two ways:1)increasing the number of loci,through comparing genome sequences of model fish species,we would develop thousands of"single-copy" nuclear coding loci,applying cross-species gene capture and Illumina sequencing,we could get the sequences of those loci;2)designing new analytical methods,screen for loci with more balanced GC content to avoid the systematic error.Our goal is to reconstruct the phylogenetic relationship of the clupeiforms(form which we have already sampled 5 families and 48 genera)and provide a robust framework for studying the systematics and evolution of the clupeiforms in the futureMany clupeiform fishes have high economic value,whereas some are also very important ecologically due to their high biomass in the marine ecosystem.For example,anchovies,sardines,herrings and the slender shad(Ilisha elongata)always are on the top of Chinese marine catch list Nevertheless,there are few molecular markers available for these species,which hindered the studies on conservation and utilization of the genetic resource of these species.In our proposed work,we are going to collect sequence data of thousands of single-copy nuclear genes from over 60 representative Clupeiform species using the cross-species gene capture approach,which was developed recently in our lab.We will develop new analytic methods to deal with the problem of low informativeness and systematic errors found in the data.We aim to answer the unresolved questions in the current classification of the Clupeiformes.Meanwhile,we will collect sequence data from population samples of the economically important species found in China coast,to select a set of gene markers for future studies on the biogeography,population genetics,conservation and genetic resource of the clupeiform fishes.We will use the slender shad(I elongata)as a case study to test the efficacy of these markersThe wide distribution of the Ilisha elongate along Northwestern Pacific coasts has been documented since the year of 1938.However,despite copious documentation,analyses of the population structure and genetic profiles were limited to morphology or mitochondrial gene.The relationships among the different populations are still elusive,especially the population genetics of the Ilisha elongate are remained unresolved,according to the previous morphological and molecular research.This may be due to limited taxon sampling,as taxonomic density is known to affect population genetic.Herein,we used targeted high-throughput sequencing of over 1000 nuclear genes,collected 144 representative specimens from 18 populations(Yellow Sea 2populations,East China Sea 4 populations,Taiwan Strait 2 populations,South China Sea 6 populations,Japan 1 population,Korea 2populations)and 1 sister taxa from the Northwesten Pacific coasts.In our research,we use such a large database to assess the population genetics of the Ilisha elongate.The nuclear gene markers revealed high polymorphisms,with89 haplotypes.This study mainly includes two aspects,which are also the main two aspects of genetics.The first is the phylogeny of Clupeiformes,and the second is the population genetics of elongate ilisha in the western Pacific Ocean.The main results of this study are as follows:1.For 62 species of Clupeiformes and 7 species of skeletal sole fish,48 genera and 5 families of Clupeiformes are included.A total of 4408 genes are enriched.RaxML gene tree and RSTRAL species tree of all genes are constructed respectively.The GC content of herring fish is relatively special,among which the GC content of herring is relatively uniform,but the GC content of other families is generally higher,up to 70%.In order to detect whether the gene sequence with uneven GC content will lead to the deviation of phylogenetic tree,a Perl script program was written to sort and screen the GC content according to the coding position,and 2416 GC content is relatively uniform.The gene tree and species tree were constructed by 1346 genes with uniform and complete GC content.The results showed that the results were almost identical with those of all gene trees and species trees.In addition,4408 genes were used to construct trees and those with uniform GC content were selected to construct trees.The results showed that both gene trees and species trees were highly consistent.It proved that the GC content in this study had no effect on the tree-building effect.2.The gene tree and species tree of 4408 nuclear genes show that the oldest member of the Clupeiformes is the family of Denticeps,which is clustered in the outer group of calcaneal bladder fish at the root of phylogenetic tree and has a high support rate.3.The gene tree and species tree of 4408 nuclear genes clearly reflect that the family of Engraulidae is a single lineage,which is consistent with the phylogenetic tree constructed by many molecular studies,whether mitochondrial data or a few nuclear gene data.4.In this study,Chirocentrus and Clupeidae were mixed together and did not differentiate independently,which is consistent with many previous studies.Chirocentrus can no longer be separated into one family.5.The results show that the family of Clupeidae is no longer a single lineage.The time of differentiation of Clupeidae is earlier.The taxonomic status of Clupeidae family is consistent with the phylogenetic tree constructed by ragl,rag2,SLC and cytb by Joshua P.The relationship between Clupeinae family and Dussumieriinae is very close and the support rate is very high.Many studies show that Clupeidae is no longer a single lineage.So we need to redefine the family of Clupeidae.6.In previous studies,the position of Sundasalax family was not clear enough,and there were different taxonomic positions in different studies.In this study,the taxonomic status of Sanda silver fishes has been stable,clustered with Clupeidae,and in the outermost part of Clupeidae family.This is consistent with recent research.Joshua P.et al.used ragl,rag2,slc,zicl,cytb,16S 6 genes and ragl,rag2,slc,cytb four genes to support that the genus Herring is a sister group of all other herring families.Bloom et al.also strongly supported that the silverband herring with Spratelloides is a sister group of all other herring families,using the results of 16s,cytb,RAG1 and RAG2 genes.7.The family of Pristigasteridae can not continue to be defined as a single family.Pristigasteridae family and Chirocentrus family are gathered together and mixed in the family of Clupeidae,and bootstrap support rate is very high.This is consistent with the fact that researchers Joshua P no longer support the taxonomic status of Sawfish family by using six genes ragl,rag2,slc,zicl,cytb,16s.In Lavoue' S and Bloom's study,the low sisterhood support rate for Sawfish and Herring was also consistent.8.In this study,the genes enriched in herring fish and freshwater fish were also compared and analyzed.GO function analysis was also carried out.56 genes with significant functions were screened out,including 11 genes with the same function and 34 genes with different functions in freshwater fish and freshwater fish.Of the 18 genes related to biological processes,only one is unique to freshwater fish,and the other 17 are unique to seawater fish.10.More than 1000 nuclear genes were obtained by target gene enrichment technology.In order to select loci with high polymorphism,one SNP locus was randomly selected at each locus in this study.A total of 1877 SNPs were obtained.SNPs combined with mitochondrial data provided a solid basis for exploring the genetic structure of the western Pacific sole population.11.144 sole fishes were from 18 populations in the western Pacific Ocean.The analysis of D-loop data of 16 sole fishes(125)and SNPs loci of nuclear genes of 18 sole fishes(144)showed that there were significant genetic differences between Dandong and other populations.The genetic structure of other populations was not obvious. |
PDF Full Text Request