## 2 Results
### Molecular Evolution of HpGGPPS Genes  
To predict the function of isolated _HpGGPPS_ candidates, the domain of their putative amino acids was analyzed. According to SMART online prediction, the deduced amino acids of seven out of eight candidates contained the full domain of Polyphenyl_syn (Pfam accession: PF00348), which is associated with isoprenoid compounds synthesis. The excluded candidate, _HpGGPPS3-4_, has incomplete Polyphenyl_syn domain, because of the missing exon in comparison with _HpGGPPS3-3_. By aligning the deduced amino acids of eight _HpGGPPS_ candidates from this study and four from Genbank (KX236181, KX236182, and KX236183 from _H. pluvialis_; XM001703117 from _Chlamydomonas reinhardtii_), motifs analysis was performed and results revealed that all 12 genes contained the signature aspartate rich motifs, SARM (DDxxxD, which x refers to any amino acids) and FARM (DDxxxD, which x refers to any amino acids), commonly found in _GGPPS_ gene (Figure 2). Unlike the SARM domain conserved among aligned genes, the variation was found in the FARM motifs. In detailed, _HpGGPPS1-1, HpGGPPS2-1_, KX236181, and XM_001703117 have the traditional motifs of DDxxxD, while rest genes have a modified motifs of DDxx-D. Moreover, _HpGGPPS1-1, HpGGPPS2-1_, KX236181, and XM_001703117 have the additional CxxxC motifs, indicating they might have different bio-functions from the rest of genes. Besides, _GGPPS_ genes from _H. pluvialis_ (_HpGGPPS1-1, HpGGPPS2-1_, and KX236181) have the modified TxxxC motifs, comparing with the _GGPPS_ gene from _C. reinhardtii_.  
Figure 1: Schematic diagram of _HpGGPPS_ gene structure, which was predicted by aligning with corresponding genome sequences. The slash indicates incomplete genome sequences resulting partial gene structure.  
To further evaluate the evolutionary pattern of _HpGGPPS_ genes, a phylogenetic tree was constructed using _GGPPS_ genes obtained in this study and retrieved from Genbank which originally from other species including Planta, Bacteria, Archaea, and Fungi (Figure 3). Eight _HpGGPPS_ genes were clustered into three distinguish groups, paralleled with the three classes identified according to sequence similarity. The Group1 genes, composing of _HpGGPPS1-1_ and _HpGGPPS1-2_, were clustered with _GGPPS_ genes from Plantae and were more close to _Euglena gracillis_ and _Thermospheococcus elongatus_ belonging to photosynthetic unicellular species, compared with other high plant species. The Group2 genes, composing of _HpGGPPS2-1_ and _HpGGPPS2-2_, were in a distinct cluster apart from the cluster containing _HpGGPPS3-1_, _HpGGPPS3-2_, _HpGGPPS3-3_, and _HpGGPPS3-4_ (Group3). It is  
Figure 2: Alignment of Polyprenyl_syn domain region in the deduced amino acids of _GGPPS_ genes. The motifs for CxxxC, FARM, and SARM are red squared. The sequence of XM_001713117 is from _Chlamydomonas reinhardtii_, while the rest of 11 sequences are from _Haematococcus pluvialis_.  
interesting that six putative _HpGGPPS_ genes from Group2 and Group3 were more likely clustered with _GGPPS_ genes from Bacteria, which implied that their function might different from _HpGGPPS1-1_ and _HpGGPPS1-2_ that are close to Planta evolutionarily.