Cloning And Expression Of CAST Gene And Morphological Characterization Analysis Of F₁ Hybrids Between Ctenopharyngodon Idellus And Squanliobarbus Curriculus

The F1 from distant hybridization between grass carp (Ctenopharyngodon idellus) and barbel chub (Squaliobarbus cunrriculus) are superiors to grass carp in terms of disease resistance and meat quality. To analyze the morphological characteristics of reciprocal cross F1 and molecular mechanism of meat quality, aiming to provided theoretical basis to identify and generalize the F1, the traditional shape and modern framework traits of four species: direct cross F1 (grass carp(?)×barbel chub(?)), reverse cross F1(barbel chub(?)×grass carp(?)), grass carp and barbel chub, were measured, and CAST gene’s bioinformatics features of three species:direct cross F1, grass carp and barbel chub, were analyzed. The main research results are as follows:(1) Direct cross F1 and reverse cross F1 had no beard inheriting grass carp, and they were of red spot in eye inheriting barbel chub. Three countable characters:soft ray of pectoral fin, lateral line scales and scales below lateral line, existed significant differences (P<0.05)between reciprocal cross F1 and barbel chub. The hybrid index indicated that body width of direct cross F1 was super phenotypic, five Measurable characters (snout length, mouth length, oral fissure length, interorbital distance and tail fin length) of reverse cross F1 were super phenotypic. The results of cluster analysis showed that reciprocal cross F1’s traditional morphometric characteristics was similar to barbel chub while its modern framework traits resembled grass carp. The identification key and discriminant functions (the discrimination accuracy was 100%) could distinguish effectively reciprocal cross F1 with their parents.(2) The full cDNA sequences of CAST gene in grass carp, barbel chub and direct cross F1 were obtained by using RACE-qPCR method. Accession numbers in GenBank were KT780366.1, KU994884, KU994883, respectively. The complete sequences of CAST cDNA in these three species were 3036bp,3165bp and 3086bp, respectively. Their ORF was 2706bp,2682bp and 2715bp, respectively, encoding 901 amino acids residues,893 amino acids residues and 904 amino acids residues, respectively. Molecular weight of these putative protein was 93.72 kDa,92.77 kDa and 94.02kDa, respectively, and their isoelectric point(PI) was 5.92,6.01 and 6.02, respectively; These protein had four calpain-inhib domains with the conserved seven-peptide sequences (TLPPXYR).The similarity of cDNA sequence from grass carp and barbel chub to direct corss F1 was 99%,96%, respectively. The similarity of amino acids residues from grass carp and barbel chub to direct corss F1 was 97%、93.7%, respectively.(3) Tested the relative expression of CAST mRNA in different tissues of direct cross F1 and muscle tissue of their parents by using RT-PCR method. The results indicated that CAST mRNA expressed in all the thirteen tested tissues of direct cross F1, and the relative expression in muscle was significantly higher than other tested tissues (P<0.05). The relative expression in muscle of direct cross F1 was significantly higher than barbel chub (P<0.05), it has no significant difference with grass carp (P>0.05).