Screening of universal DNA barcodes for grass forages’ eight species

DNA barcoding is an emerging technology, which is used as a tool to indentify species accurately and quickly using the conservative DNA segments. Our research has established and optimized target fragments’ the amplification conditions for eleven samples (Sorghum bicolor×Sorghum sudanense, Zea mays, Stipa capillata, Lolium perenne ‘Plxie’, Lolium perenne ‘Caddieshack’, Festuca kansuensis, Festuca rubra ‘Bargena’, F. rubra ‘Maxima’, Poa pratensis ‘Barvictor’, P. pratensis ‘Diamond’, Achnatherum splendens) from seven major genera (Sorghum, Zea, Stipa, Lolium, Festuca, Poa, Achnatherum) and eight species according to nucleotide sequence of grass matK and rbcL genes in GenBank, and designed four universal primers. 5' end and 3' end conservative fragments in four marked fragments were selected by amplification products sequencing and analyzing. Single nucleotide polymorphism (SNP) haplotype analysis of conservative fragments nucleotide sequences of each maker loci showed there were 6 haplotypes(H1A, H1B, H1C, H1D, H1E and H1F), 7 haplotypes (H2A, H2B, H2C, H2D, H2E, H2F and H2G) and 3 (H3A, H3B and H3C) haplotypes among matK1, matK2, matK3 respectively, by the same token, rbcL gene has 5 haplotypes (H4A, H4B, H4C, H4D and H4E). According to the haplotypes about matK (matK1, matK2 and matK3) and rbcL, we have constructed the DNA barcoding for eight grass species. The research results provide a scientific basis for identification of mixed feed grasses including eight species of the seven generas.