**2. Material and methods**

#### **2.1 Study site**

The research site is located in Dengkou County, Bayannur City, Inner Mongolia, with an altitude of about 1050 m. This region is located in the eastern edge of Ulan Buh Desert, which belongs to temperate continental monsoon climate. The average annual temperature is 7.6°C, and the accumulated temperature during the growing period is about 3100°C. The annual average rainfall is 144.5 mm, and the precipitation is unevenly distributed throughout the year, mainly from June to September. The annual average evaporation is 2397.6 mm, and the frost-free period is 136 days. The main soil type is eolian sandy soil, whose mechanical composition is dominated by fine sand (0.05–0.25 mm), accounting for more than 84%, with little physical clay and coarse sand. The experimental site is located at 106<sup>o</sup> 50'E, 40o 30'N.

### **2.2 Field survey, lab examination and data analysis**

The indoor algae culture experiment began on November 20, 2019. The field algae culture experiment began on January 2, 2020 in Southeastern margin of Ulan Buh Desert, but due to the sand burial, the field experiment did not succeed. After the algae spraying, water was sprayed once a day to keep the surface layer moist. On January 2, 2020, the 0–5 cm soil was sampled and divided into three parts. One part of the fresh soil was sealed in a self-sealing bag and sent to the laboratory at a low

temperature in an ice box. The fresh soil was stored in a refrigerator at −80°C for the determination of soil microbial carbon and nitrogen. Part of the fresh soil was sealed in a self-sealing bag and brought back to the laboratory at a low temperature in an ice box. It was stored in a 4°C refrigerator for the determination of ammonium and nitrate nitrogen content. The other parts of the soil were used to determine the organic matter, total carbon, total nitrogen and total phosphorus of the soil after natural air drying, in order to quantify the effects of different ratios of algae crusting on the basic physical and chemical properties of the soil.

Using concentrated sulfuric acid - distillation titration method to measure the total soil nitrogen, using elemental Analyzer to measure the total soil carbon. Soil total phosphorus was determined by alkali fusion - molybdenum antimony anti-spectrophotometry, and microbial carbon and nitrogen were determined by chloroform fumigation extraction method.

One-way ANOVA was used to study the change degree of soil chemical and biochemical indicators of soil samples sprayed with different ratios of algae under 95% confidence interval. If there were significant differences, the least significant difference method (LSD) was used for multiple comparisons. P < 0.05 indicated significant differences. Data were collected using SPSS (version 17.0, SPSS Inc., Chicago, IL, USA) and charted using Origin (version 2019, OriginLab Inc., Northampton, MA, USA).

#### **2.3 Artificial culture of algae crust**

Algae are the pioneer species for the formation of BSC. As pioneer colonizers, cyanobacteria can grow and reproduce under harsh environmental conditions, such as drought, ultraviolet radiation, poor nutrition, etc. Cyanobacteria can form a fixed quicksand algae crust on the sand surface.

Sand-fixing algae crusts can be formed in a short time by artificial culture. With the development and succession of algeal-sand crusts, and the material input brought by the deposition of fine particles and the accumulation of atmospheric dust, the nutrient enrichment in the sand surface layer was promoted, which created conditions for the reproduction of micro-soil organisms and the colonization of herbaceous plants, and then promoted the desert ecosystem into a virtuous cycle [28].

Algae crust artificial cultivation, is using the principle of algae ecology, physiology, and the theory of biological crust, separating, choosing a natural development formation of the excellent algal crust, after large-scale artificial cultivation, incubated on the surface of the sand to make the rapid formation of crust with algae, bacteria, fungi, algae. The technology mainly includes five aspects [1]:

(1) Isolation, purification and breeding of fine algae species in algae crusts; (2) the scale culture of alga species; (3) Factory/scale production; (4) Incubation in the field; (5) Management and maintenance. The main process and steps of artificial culture of algae crust are as follows (**Figure 1**).

① The algae crusts developed well under natural conditions were adopted. During sampling, the sampling frame is first pressed into the soil forcefully. When the sampling frame is completely put into the soil, the algae crust of about 50 cm2 in the stainless steel frame is gently taken out with a shovel disinfected with 75% alcohol and put into an envelope for use.

② Sample sieving. Disinfect the gloves with 75% alcohol before operation, and wear gloves; The collected algae crust samples were crushed, passed through a 0.2 mm sieve, and set aside.

③ Cleaning. 20 g of the screened algae crust sample was soaked in 50 mL distilled water and stood for 20 min. The soaked algae crust sample was placed on an ordinary medical gauze and rinsed slowly in distilled water until all the impurities and soil in the sample were removed. The remaining liquid was a mixed algal suspension.

*Cultivation of Artificial Algal Crust and Its Effect on Soil Improvement in Sandy Area DOI: http://dx.doi.org/10.5772/intechopen.98716*

**Figure 1.**

*Flow diagram of artificial algae crust cultivation. (a) Algae cultivation (b) algae propagation (c) field inoculation.*

④ The separation of algae species. The algal suspension in step ③ is examined under a microscope. If it is found that a large number of algae need to be separated, it can be separated immediately. If the quantity is small, it should be pre-cultured first, and then separated after increasing.

⑤ Species of algae culture. 2 ~ 5 mL of isolated single or mixed seaweed suspension was taken and added with 200 mL BGI1. The nutrient solution (BG11 culture solution composition and dosage are shown in **Table 1** was placed in a triangular flask. Place the triangular flask on the shaker (Rpm is 140rmin-1) for initial culture, indoor temperature was controlled at 25 ~ 30°C, light intensity was controlled at 600 lx. Indoor culture after 7–10 days, the algae suspension was transferred to a 2-L volumetric flask for preliminary propagation. After propagation for 7 days, the culture medium was transferred into a 50 L plastic bucket, each bucket was equipped with 50 L BG11 culture medium and a set of oxygenation facilities (such as oxygenation pump for domestic ornamental fish); The dosage requirement of algal factory production can be reached after about 7 days of cultivation outdoors.

⑥ The large-scale production of algae. Add the species algae cultured in step ⑤ into the production pool (the fresh weight of species algae added in each pool is about 200 g, and the dry weight is about 2 g). The culture medium used in the incubator was BG11 culture medium, and the water depth was about 0.5 m. In the process of cyanobacteria culture, the growth of cyanobacteria is the best when the water temperature is 25–30°C, and cooling measures should be taken when the water temperature is >40°C (because cyanobacteria will stop growing when the water temperature exceeds


#### **Table 1.**

*Composition and dosage of BG11 medium.*

40°C). The light intensity is controlled at 15000 lx. Harvest will be carried out after 7 to 12 days of culture of cyanobacteria (about 7 days in July and August in summer, 10–12 days in April and June in spring and 10–12 days in September and October in autumn). Determination of harvest period: it was determined according to the growth curve of cyanobacteria. When the growth rate or increase amount of cyanobacteria began to decline (the growth amount began to decline after reaching the maximum), the harvest was carried out. Before harvest, the liquid in the culture tank was left standing for 6 ~ 8 h. The culture solution is then discharged into an empty pool. (After one culture, the algae can still grow, but the yield is reduced.)30% ~ 40%. During the culture process, green algae from the air will enter the culture pond, leading to increased competition between algae and cyanobacteria. When the water depth in the pool is about 0.5 cm, the algal fluid is collected into a plastic bucket for use.
