Structure and organization of the leghemoglobin genes in soybean

The organization and structure of the leghemoglobin (Lb) genes in soybean have been investigated. Using molecular cloning techniques, a Lb-cDNA recombinant molecule, pLb1, was prepared and characterized. Hybridization of this clone of genomic DNA revealed the presence of at least seven Lb genes in the soybean genome. The arrangement of these genes appeared to be the same in DNA isolated from infected or uninfected tissues. The plasmid pLb1 was also used to isolate three Lb sequences from a genomic library constructed in (lamda) Charon4 vector. Nucleotide sequence analysis showed that one Lb gene, present on an 11.5 kilobases (kb) Eco RI genomic fragment, spans about 1 200 nucleotides and appears to code for Lbc(,3). Its coding sequence is interrupted at amino acid positions 32 to 33, 68 to 69 and 103 to 104. The intervening sequences, as well as the 5' and 3' flanking regions of this gene, contain some consensus sequences found in other eukaryotic genes. The length of the 5'-untranslated region is 49 bases as determined by nuclease S1 mapping. R-loop analysis of the recombinant phage containing the 11.5 kb Eco RI fragment showed that another Lb gene is located 2.5 kb away. The nucleotide sequences of the second gene showed that this gene is incomplete, containing only two exons. The deduced amino acid sequence of this gene, although showing 78% homology with the corresponding region of the other Lb gene, is not represented in any of the known Lb proteins. Both genes are oriented in the same direction with respect to the coding strand. Analysis of the sequence present in a second genomic clone containing a 4.2 kb Eco RI fragment revealed a truncated Lb gene showing homology with the last exon and the non-coding region at the 3' end of the two other Lb genes. A very high homology is found among the nucleotide sequence of the Lbc(,3), Lba and Lbc(,1) genes. Comparison of the intervening sequences of these genes indicated that they diverged mainly through the creati